Clustered distribution of virus infected cells in the liver of patients with chronic hepatitis CIntroduction: detection of hepatitis C virus RNA and proteins in the liver can not only contribute to the knowledge of the mechanisms of HCV replication and pathogenesis but also complement diagnostics and therapeutic studies.
Objective: to determine the presence of hepatitis C virus in the liver of patients with chronic hepatitis C that were immunized with a therapeutic vaccine candidate.
Methods: detection of hepatitis C virus RNA and proteins were studied by in situ hybridization and immunofluorescence assays in liver biopsies from 14 hepatitis C virus -infected individuals.
Results: hybridization signals for hepatitis C virus-RNA of both positive and negative polarity were detected in the cytoplasm of hepatocytes from most of the samples analyzed. However, the HCV structural antigens could not be detected in any of the samples. The hepatitis C virus RNA was observed on granular structures in the cytoplasm of hepatocytes. This staining pattern is similar to that described for stress granules and/or lipid droplets which are involved in hepatitis C virus replication. The proportion of cells showing positive reactions for hepatitis C virus RNA of negative polarity (which is a marker of ongoing viral replication) ranged from 4.47 % to 15.94 %.
Conclusions: results from this work suggest that hepatitis C virus infection occurs in groups of neighbouring hepatocytes that are distributed sporadically in the liver. This is consistent with the model of clustered spatial distribution of hepatitis C virus infected cells and cell to-cell spread of hepatitis C virus in the liver, and suggests that hepatitis C virus replication was constrained in these patients.
Viviana Falcón, Nelson Acosta-Rivero, Sirenia González, Emilio Acosta Medina, Rocio Garateix Suárez, José A. Silva, Liz Álvarez-Lajonchere, Gillian Martínez-Donato, Carmen M. Rosales, Santiago Dueñas-Carrera, Juan Kourí
Maria Teresa Martínez Echevarría, Raúl Pablo Ferreira Capote, Isis Amores Sánchez, Amarylis Martínez Piedra, Rita Rosa Santana Hernández, Ángela Rosa Gutiérrez Rojas, Gissel García Menéndez
Ultrastructural remodelling of e ndoplasmic reticulum-derived membranes and hepatitis C virus morphogenesis in human liver samples

Introduction: hepatitis C virus infection is considered as a leading cause of liver disease worldwide. Despite recent advances in understanding the hepatitis C virus life cycle using the highly replicative JFH1 strain in human hepatoma cells, little is known about hepatitis C virus morphogenesis. Low levels of hepatitis C virus assembly in this cell culture model as well as low levels and complexity of hepatitis C virus particles in infected humans and chimpanzees have hampered the study of hepatitis C virus morphogenesis in vivo.
Objetivo: to study the ultrastructural features and viral assembly events in hepatocytes from HCV hepatitis C virus-infected patients.
Methods: liver needle biopsies samples of patients with hepatitis C virus infection, specific antibodies against hepatitis C virus and transmission electron microscopy and immunoelectron microscopy analyses were used in this study.
Results: ultrastructural studies in liver biopsies from hepatitis C virus-infected patients revealed that hepatitis C virus infection was related with remodelling of endoplasmic reticulum-derived membranes and with a variety of cytoplasmic microenvironments in hepatocytes. Dilated endoplasmic reticulum and formation of various membrane vesicles are features that have been associated with the viral replication complex. Interestingly, hepatitis C virus-like particles and core-like particles budding and assembly were observed near convoluted electron-dense membranes and tubular structures. Particularly, hepatitis C virus structural proteins localize to the endoplasmic reticulum.
Conclusions: these events indicate that hepatitis C virus nucleocapsids and early virion assembly take place at endoplasmic reticulum membranes that are associated with these cytoplasmic microenvironments in human hepatocytes.

Viviana Falcón Cama, Nelson Acosta-Rivero, Sirenia González Pozos, Santiago Dueñas-Carrera, Emilio Acosta Medina, Juan Kourí Flores
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