Hepatitis C virus persistence and lymphotropism

MacParland, Sonya Ann (2010) Hepatitis C virus persistence and lymphotropism. Doctoral (PhD) thesis, Memorial University of Newfoundland.

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Abstract

Hepatitis C virus (HCV) leads to chronic liver disease in up to 80% of those infected. While thought to be mainly hepatotrophic, HCV has been considered to be able to replicate in cells of the immune system. The extent and implications of viral replication in immune cells are unknown, however, this site may represent an important reservoir from which the release of infectious virions, leading to viral reactivation may take place. -- We hypothesize that HCV-naive lymphocytes are susceptible to infection with wild-type HCV and are capable of supporting the full cycle of its replication, including the release of biologically competent virions. We also postulate that small amounts of virus, lingering for years after apparent complete clinical resolution of hepatitis C, retains its infectious potential. -- This thesis is comprised of three related, but also stand-alone studies. In the first study, we established an in vitro infection system in which wild-type HCV was used as inoculum and primary human lymphocytes served as infection targets. Employing this system, we demonstrated that HCV can infect T lymphocytes, as it became apparent via detection of HCV genome replicative intermediates, non-structural proteins, and the appearance of unique variants in infected cells. We further identified and characterized the biophysical properties of virion particles by sucrose gradient centrifugation and by immune electron microscopy with HCV-specific antibodies directed against virus envelope (E2) protein, and by examining the in vitro infectivity of secreted virions by their serial passage in virus- naive lymphocytes. -- In the second study, we revealed that in vitro infection of T cells with HCV can lead to an altered T cell subset distribution characterized by an enrichment of CD8⁺ T cells that appears to be a result of an inhibition of CD4⁺ cell proliferation but not virus-related apoptotic death of the cells. -- In the third study, by utilizing the system established in the first study, we revealed for the first time that small amounts of HCV persistently circulating after apparent complete clinical resolution of hepatitis C due to antiviral therapy can infect HCV naive human T cells. This finding may have significant pathogenic (residual liver disease) and epidemiological (infectivity) consequences. -- Overall, our findings provide conclusive evidence that cells of the immune system can act as reservoirs of HCV and that virus replication in this compartment leads to the production of infectious virions. Furthermore, we described that virus exposure can alter the proliferative ability of lymphoid cells, possibly impairing their response to virus and slowing viral clearance in vivo. Finally, we documented the infectious potential of residual virus that lingers in essentially asymptomatic patients after resolution of chronic hepatitis C. While the clinical significance of our findings are currently under investigation, the in vitro system established, capable of supporting the entire cycle of HCV replication in the natural cell milieu, may serve as a valuable tool to study poorly understood aspects of HCV infection, including factors determining cell susceptibility and virus-induced cytopathic mechanisms induced by the virus, as well as being applicable for evaluations of novel anti-HCV agents.

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