Ultrasensitive methods to measure very low levels of hepatitis C virus (HCV) RNA in biological samples may have diagnostic and prognostic significance and be useful to evaluate the response to antiviral treatment. A sensitive assay to quantify HCV RNA in peripheral blood mononuclear cells (PBMCs) was developed and validated using the iCycler iQ™ Detection System (Bio-Rad) coupled with TaqMan® chemistry. HCV was co-amplified with the endogenous control glyceraldehyde-3-phosphate dehydrogenase in a multiplex reaction. Calculated PCR amplification efficiencies for both target and control genes were used in a mathematical model for relative quantitation of HCV RNA. A linear relationship between input RNA and CT values over 6 log dilutions was observed for both HCV- and GAPDH-specific products (R2 ≥ 0.99). As few as 1.5 IU/reaction could be detected, with high accuracy (CV ≤ 3.94%) and reproducibility (CV ≤ 2.20%). Quantitation of HCV RNA levels ranging from 103 to 107 IU/ml as measured in 47 plasma samples was highly correlated with values obtained by the COBAS Amplicor™ HCV Monitor test, v2.0 (Roche) (R2 = 0.977). In conclusion, this assay provides an excellent tool to determine accurately HCV kinetics in PBMCs during antiviral therapy and to assess the long-term significance of different patterns of response to treatment.

Background/Aims: The hepatitis C virus (HCV) induces lipid accumulation in vitro and in vivo. Although clinical observations are consistent with a direct effect of HCV genotype 3a on lipid metabolism, experimental systems have focused on the expression of HCV proteins of genotype 1. To extend these observations, we established an in vitro model expressing the HCV core of different genotypes.Methods: The HCV core protein from patients with severe (genotype 3a) or no (genotypes 1b, 2a, 3h, 4h and 5a) liver steatosis was expressed in Huh7 cells. Core protein expression (by immunohistochemistry and immunoblot) and triglycerides accumulation (by Oil Red 0 stain and enzymatic measurement) were evaluated 48 h after transfection.Results: Although triglyceride accumulation occurred with genotypes 1b, 3a and 3h, the genotype 3a core protein expression resulted in the highest level of accumulation (i.e. about 3-fold with respect to 1b, and 2-fold with respect to 3h). This effect was not related to core protein expression levels and was abolished by culturing cells in lipid-free medium.Conclusions: Consistent with observations in chronic hepatitis C patients, the in vitro expression of HCV genotype 3a core protein is the ideal candidate model for studying the mechanisms of HCV-associated steatosis. (c) 2005 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.

This paper concerns the development of a scenario aiming at the improvement of teacher education in Geneva in the field of media, image and ICT (MITIC) in a media- and pedagogy-oriented process instead of technology-driven action as is usually the case. Among a lot of observations, experiments and constraints, the key factors we address are the lack of well-educated teachers of teachers (level F3), willingness to enhance coherence in teacher education together with pedagogical research (innovative projects), the need to take into account the evolution of the demography of teachers in professional development and the necessity to promote an attitude to and an awareness of practices in collaborative and cooperative work. In this process we have built a prototype of a new concept of teacher (F3-MITIC), and we have cooperated in a national incentive policy and proposed an agenda for the remaining problems to be solved. We conclude the presentation of our contribution by the assessment of the fundamental necessity to put in action a real e-piloting including a set of strategies, as well as projects and tools which will have to be elaborated with the actors in the field, in the four following directions: sharing data, re-defining teachers' competencies, taking care of innovative projects and designing pedagogical models of e-learning.