Quantitative analysis of hepatitis C virus in peripheral leukocytes and phenotypic analysis of liver-infiltrating lymphocytes in HCV infection.

摘要

Hepatitis C virus (HCV) is a hepatotropic virus with a worldwide incidence estimated at 3%. It has been suggested that HCV is able to establish a productive infection in extrahepatic reservoirs, namely peripheral lymphocytes. Furthermore, while much is known about the peripheral immune response, far less is known about the intrahepatic response against HCV. This two-part thesis addresses both of these issues.; The first goal of this work was to characterize the nature of HCV's association with peripheral blood mononuclear cells. Chronic HCV patients were recruited and viral load was measured in the following purified leukocyte subpopulations: polymorphonuclear cells (PMNs), T cells, B cells and monocytes. Replication was then assayed by the specific detection of negative sense viral RNA. A preferential association of HCV with B cells was found, while T cells were virtually negative and PMNs and monocytes displayed very low levels of viral RNA. Because peripheral leukocytes are in a resting state, it was thought that cellular activation might also stimulate HCV to go from a latent or non-replicating state, to an actively replicating state. Purified peripheral B cells from chronic HCV patients were stimulated in vitro and both total viral RNA and negative-strand viral RNA were measured. It was thus determined, that under these particular conditions, HCV replication is not occurring.; These results are significant because of their relevance not only to tissue tropism, but also to in vitro cell culture.; The second goal of this work was to phenotypically characterize the lymphocytic infiltrate in end-stage and chronic HCV infection. Lymphocytes were isolated from liver tissue samples of HCV patients. These liver-infiltrating lymphocytes were evaluated by flow cytometry for a number of different cell surface markers, including subpopulation-defining molecules, T cell receptor, co-receptor, activation molecule and chemokine receptor expression patterns.; These results are significant because of the important role of the site-specific immune response, our emerging understanding of the role played by chemokines in such site-specific responses and our need for better understanding of what constitutes an effective immune response against HCV.; Collectively, these results constitute a valuable contribution to the field of HCV pathogenesis and immunology.