Human malignant mesothelial cells: Variability of ultrastructural features in established and nude mice transplanted cell lines

The aim of this study was to determine, by transmission electron microscopy, the differentiation features of 21 human malignant mesothelioma cell lines (HMCLs) established from 13 specimens of 12 confirmed human malignant mesotheliomas, and of tumours induced in nude mice injected with 16 HMCLs. Fifty per cent of HMCLs showed typical mesothelial differentiation (long and slender microvilli, desmosomes, perinuclear intermediate filaments); 29 per cent did not show differentiation; and the remainder were poorly differentiated. Three human tumour specimens gave several different HMCLs; the cell lines obtained from a given tumour exhibited variable mesothelial differentiation. Eleven HMCLs were compared with the native tumour. Four were similar to the tumour and seven were less well differentiated, in most cases in relation to their microvilli. With six HMCLs, tumours induced in nude mice were less well differentiated than the corresponding cell lines, whereas with four HMCLs, tumours were equally or better differentiated. However, in most nude mice tumours, typical mesothelial microvilli were present. These results show that cell lines established from malignant mesothelioma may exhibit dedifferentiated features. However, while the variability in ultrastructural differentiation may result from the culture microenvironment, it could also be related to the state of differentiation, of the native tumour sample and to tumour cell heterogeneity.     

Cloning and expression of surface antigens from occult chronic hepatitis B virus infections and their recognition by commercial detection assays

Occult hepatitis B virus (HBV) infections show little or no serological markers of viral infection, including the absence of hepatitis B surface antigen (HBsAg) which is the main marker of ongoing HBV infection. Such infections can be important in the context of blood and/or organ donations. To study whether mutations contribute to HBsAg seronegativity, S gene sequences from such patients were amplified and cloned. Sequencing revealed 12 clones from seven different patients which contained potentially important mutations. The sequences were subcloned into an expression vector and mutant HBsAgs were expressed in cell culture. The capacity of three HBsAg detection assays to recognise the mutant HBsAgs was studied. Three categories were found: mutant HBsAgs that are not recognised by the assays, those that are recognised as well as wild-type (WT) antigen and an intermediate category where detection of the mutant HBsAgs is reduced with respect to WT. Most of the isolates fall into the second category. Mutations can therefore contribute to HBsAg seronegativity in occult HBV infections, but in most cases the explanation is probably the low level of viral replication.     

Electronic implementation of guidelines in the EsPeR system: a knowledge specification method

Despite initiatives to standardize methods for the development of clinical guidelines, several barriers hinder their integration in daily clinical practice: failure to fulfil quality criteria, poor effectiveness of their dissemination. Computerization of guidelines can favor their dissemination. The initial step of computerization is the knowledge specification from the text of the guideline. We describe the method of knowledge specification, which is used in EsPeR (Personalized Estimate of Risks), a web-based decision support system in preventive medicine, which allows, for a given person, to estimate risks and access recommendations, based on clinical profile. This method is based on a structured and systematic analysis of text allowing detailed specification of a decision tree. We use decision tables to validate the decision algorithm and decision trees to specify this algorithm, along with elementary messages of recommendation. Editing tools are used to facilitate the process of validation and the workflow between expert physicians and computer scientists. Applied to eleven different guidelines, the method allows a quick and valid computerization and integration in the EsPeR system. The method used for computerization could help to define a framework usable at the initial step of guideline development in order to produce guidelines ready for electronic implementation.     

Identification of dermatophyte species by 28S ribosomal DNA sequencing with a commercial kit

We have shown that dermatophyte species can be easily identified on the basis of a DNA sequence encoding a part of the large-subunit (LSU) rRNA (28S rRNA) by using the MicroSeq D2 LSU rRNA Fungal Sequencing Kit. Two taxa causing distinct dermatophytoses were clearly distinguished among isolates of the Trichophyton mentagrophytes species complex.     

A longitudinal and prospective study of Epstein-Barr virus load in AIDS-related non-Hodgkin lymphoma.

BACKGROUND: Epstein-Barr virus (EBV) may be causally associated with non-Hodgkin Lymphoma (NHL) in HIV-infected patients. OBJECTIVES: To compare EBV load in whole blood in AIDS-NHL patients, HIV non-AIDS patients and non-HIV-infected persons, and to prospectively measure EBV load in whole blood in AIDS-NHL patients. STUDY DESIGN: Longitudinal and prospective study. RESULTS: We observed no statistical difference in EBV load between AIDS-NHL (3.69log(10) copies/mL [interquartile range (IQR): 2.89-4.27]) and HIV non-AIDS patients (3.08log(10) copies/mL [IQR: 1.29-3.57]) but AIDS-NHL patients had significantly higher EBV loads than HIV-negative controls (1.19log(10) copies/mL [IQR: 0.00-3.29]). We noticed an inverse correlation between CD4+ lymphocytes count and EBV load in patients with AIDS-NHL (r(2)=0.41, P=0.01). In the longitudinal study, the mean EBV load three months after NHL diagnosis decreased significantly (mean difference=-1.69log(10) copies/mL [95% confidence interval: -0.32; -3.04]; P=0.03) under chemotherapy but was still elevated in patients with relapses or no response to chemotherapy. CONCLUSION: Although EBV load seems a suboptimal marker for the diagnosis of AIDS-NHL, we observed a significant decrease of EBV load in patients treated with chemotherapy and a strong association between NHL outcome and EBV load in whole blood.     

Interaction of enteropathogenic and Shiga toxin-producing Escherichia coli and porcine intestinal mucosa: role of intimin and Tir in adherence

The ileal in vitro organ culture (IVOC) model using tissues originating from colostrum-deprived newborn piglets has proven to be an effective way to study the attaching and effacing (A/E) phenotype of porcine enteropathogenic Escherichia coli (EPEC) ex vivo. The aim of this study was to investigate the role of intimin subtype and Tir in the adherence of EPEC and Shiga-toxin-producing E. coli (STEC), isolated from different animal species, to porcine intestinal IVOC. Moreover, the role of intimin in Tir-independent adherence of the human EPEC strain E2348/69 was investigated using intimin and Tir-deficient derivatives. Our results demonstrated that A/E E. coli strains (AEEC) from various animal species and humans induce the A/E phenotype in porcine ileal IVOC and that intimin subtype influences intestinal adherence and tropism of AEEC strains. We also showed that a tir mutant of EPEC strain E2348/69 demonstrates close adherence to the epithelial cells of porcine ileal IVOC segments, with microvillous effacement but with no evidence of actin polymerization or pedestal formation, and that intimin seems to be involved in this phenotype. Overall, this study provides further evidence for the existence of one or more host-cell-encoded intimin receptor(s) in the pig gut.     

Up-regulation of tRNA biosynthesis affects translational readthrough in maf1-delta mutant of Saccharomyces cerevisiae

Maf1p is a negative effector of RNA polymerase III in yeast. The maf1-delta mutation caused an increase in the level of cellular tRNAs, but a decrease of translational readthrough at nonsense codons. Using the lacZ- luc dual gene reporter system, we detected an almost twofold diminution of UAA and UAG readthrough in maf1-delta compared with the parental strain. The maf1-delta mutation did not affect the rate of protein biosynthesis and growth at standard conditions, but resulted in temperature-sensitive growth on non-fermentable carbon sources. We examined the correlation of the temperature sensitive and antisuppression phenotypes of maf1- Delta using a colour phenotype assay in the ade2-1 SUP11 strain. Antisuppression, but not the temperature-sensitive growth defect, was compensated either by increased dosage of SUP11or by [PSI(+)], the prion form of the translation termination factor Sup35p. Summarizing, the elevated tRNA levels in maf1- Delta increase translational fidelity and, independently, affect growth under special conditions.     

Levels of analysis in etiological research on schizophrenia

Etiological research spanning domains of inquiry as diverse as social psychology and molecular genetics has identified a number of potential factors that likely contribute to the development and clinical manifestation of schizophrenia. In this article, we first highlight the challenges inherent in developing cogent etiological models that represent both the diversity of suspected causal influences and their mechanisms of action. Then, using our own research program as a heuristic context, we present a general analytical framework for identifying and integrating multiple types of etiologic factors across different levels of analysis in the prediction of schizophrenia. In recognition of the myriad complexities of multifactorial causation, we argue that a multilevel causal perspective is required for the development and advancement of a fully nuanced theory of schizophrenia etiology and pathophysiology.     

Plasma RNA viral load in human immunodeficiency virus type 2 subtype A and subtype B infections

Human immunodeficiency virus type 2 (HIV-2) is much less pathogenic than HIV-1, and HIV-2 infection is associated with plasma viral loads significantly lower than those found in HIV-1 infection. We have developed a real-time quantitative PCR method for measuring the HIV-2 RNA load that covers the range of genetic diversity of HIV-2 isolates and that detects extremely low viral loads. Samples from 49 patients were studied. Proviral DNA was first detected and quantified. The strains that were detected were then genotyped: 21 patients were infected with HIV-2 subtype A and 15 patients were infected with HIV-2 subtype B; 1 patient was infected with a highly divergent strain. Env PCR failed for the remaining 12 patients, so subtypes could not be determined. For viral RNA quantification, a stock of HIV-2 strain NIHZ, which was counted by electron microscopy, was used as the standard. Several primer sets targeting the highly conserved gag region were evaluated. Various primer combinations failed to amplify subtype B strains. With the final primer pair selected, which detected both subtype A and subtype B strains, the sensitivity of the assay was 100% at a viral load of 250 copies/ml and 66% at a viral load of 125 copies/ml. We found a correlation between the CD4(+)-cell count, the clinical stage, and the plasma HIV-2 RNA level. The median plasma HIV-2 RNA value for the 33 asymptomatic patients was 2.14 log(10), whereas it was 3.1 log(10) for the 16 patients with AIDS (P < 0.01). Proviral DNA was detectable in 18 symptom-free patients with high CD4(+)-cell counts, in whom viral RNA was undetectable.