Document Type

Dissertation

Date of Degree

2009

Degree Name

PhD (Doctor of Philosophy)

Degree In

Molecular and Cellular Biology

First Advisor

Anton McCaffrey

Abstract

microRNAs (miRNAs) are newly discovered small RNAs that repress gene expression and are evolving as clinical predictors for diagnosis and prognosis of human disease. Persistent hepatitis C virus (HCV) infection causes chronic inflammation and can lead to fibrosis, cirrhosis and liver cancer. HCV-associated liver injury is characterized by increased hepatocyte proliferation. Studies have demonstrated interactions between HCV and miRNAs. We investigated the role of miRNAs during HCV infection. We used global expression analysis to measure the expression levels of 380 miRNAs comparing HCV infected human livers with uninfected livers. We correlated the altered miRNA expression levels with clinical patient data, such as stage of fibrosis, liver transaminases, and viral load. We identified several miRNAs that correlated with these parameters, including miR-21. miR-21 correlated with stage of fibrosis, liver transaminases, and viral load. We used the mouse carbon tetrachloride model to induce fibrosis and identified correlations with miR-21 expression and stage of fibrosis. Furthermore, we identified, SMAD7, an inhibitor of fibrosis, as a novel target of miR-21 providing further evidence of miR-21's possible involvement in fibrosis development. In our patient samples, miR-21 expression also correlated with viral load. Establishing a further connection between miR-21 and HCV, we measured miR-21 expression levels in HCV replicons and infectious HCVcc tissue culture models. Altered expression of miR-21 varied depending on replicon genotype and tissue culture model preventing us from establishing a consistent relationship between miR-21 expression level and HCV infection. Interestingly, miR-21 inhibition in replicon cells resulted in decreased viral replication suggesting a possible miR-21 anti-viral effect.

Hepatocyte replication is associated with chronic HCV-induced liver injury. To investigate whether miR-21 correlation with HCV and fibrosis was a consequence of hepatocyte proliferation caused by viral infection and fibrosis, we measured miR-21 expression during liver regeneration. miR-21 was up-regulated during the early proliferative phase of liver regeneration. We identified Pellino-1 (PELI1), an adapter molecule involved in Nuclear factor-ĸB (NF-κB) signaling, as a novel target of miR-21. During liver regeneration, PELI1 mRNA levels were reduced, consistent with miR-21 targeting. We also found miR-21 over-expression decreased the activity of a NF-κB reporter. These studies investigated the role of miR-21 during HCV viral infection, fibrosis, and liver regeneration. We found miR-21 correlates with fibrosis in both human patient samples infected with HCV and in a mouse carbon tetrachloride mouse model. miR-21 may be pro-fibrogenic by targeting SMAD7, an inhibitor of fibrosis. miR-21's correlation with HCV viral load may be an anti-viral response by the host, which was demonstrated by increased HCV replication upon inhibition of miR-21. Furthermore, miR-21 is associated with hepatocyte replication during liver regeneration and inhibits NF-κB signaling, possibly by targeting PELI1. These studies identified several new roles for miR-21 during HCV viral infection, fibrosis, and liver regeneration.

Keywords

hepatitis C, liver, microRNAs

Pages

xii, 129 pages

Bibliography

Includes bibliographical references (pages 112-129).

Copyright

Copyright 2009 Rebecca Therese Marquez

Included in

Cell Biology Commons

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