Proteomic Characterization of In Vivo and In Vitro Model Systems of Hepatitis C Virus Infection: Global Quantitative Proteome AMT Tag Measurements of Cellular Protein Expression
EMSL Project ID
13792
Abstract
Approximately 170 million individuals worldwide are infected with HCV and in the United States, there are approximately 4 million infected individuals and over 10,000 HCV associated deaths annually. The most remarkable aspect of HCV is its striking ability to persist in the infected host, which often leads to progressive liver disease including fibrosis, cirrhosis, and increased risk of hepatocellular carcinoma. Unfortunately, HCV research has been hampered by the lack of an adequate tissue culture system, or an effective animal model for HCV infection. As a result, the mechanisms of HCV replication and pathogenesis remain poorly understood, and little is known about the underlying mechanisms leading to the liver injury that often results from HCV infection.To gain a better understanding of HCV replication and pathogenesis, we propose to use the AMT tag approach to study changes in the liver proteome during HCV infection and HCV-associated liver disease using in vivo and in vitro model systems. In our previous EMSL Proposal (ID 4393) we completed 1) the first large scale proteomic analysis of a full length HCV replicon model system for in vitro analysis of HCV replication and, 2) the first high-throughput characterization of the liver proteome from human liver biopsy specimens using the ultra-sensitive 'nano-proteomic" technologies at PNNL. These studies have been described in a manuscript entitled "Proteome Analysis of Liver Cells Expressing a Full-Length Hepatitis C Virus (HCV) Replicon and Biopsies of Post-Transplanted Liver from HCV-Infected Patients" by Jon M. Jacobs, Deborah L. Diamond, Eric Y. Chan, Marina A. Gritsenko, Weijun Qian, Miroslava Stastna, Tracey Baas, David G. Camp II, Robert L. Carithers, Jr., Richard D. Smith, and Michael G. Katze that has been accepted for publication in the Journal of Virology. The work proposed here represents an extension of these initial studies and is consistent with the overall goals of our Program Project Grant from the National Institute on Drug Abuse entitled "Functional Genomics and HCV-Associated Liver Disease" (grant 1P30DA01562501 to M.G.K.). This grant supports our proteomics collaboration with the Biological Systems Analysis and Mass Spectrometry group headed by Dr. Richard Smith at Pacific Northwest National Laboratory (PNNL) and the work outlined in this new EMSL proposal.
The major objectives of this EMSL user proposal are to expand our human liver AMT database utilizing recently acquired pieces of normal and cirrhotic human liver tissue and to perform quantitative proteomic analyses of human liver biopsy samples. In this regard, we propose to track HCV progression using serial liver biopsies acquired from patients after liver transplantation, perform cross-sectional studies of HCV-infected patients with various degrees of fibrosis in order to validate our findings in liver transplant recipients, and characterize the more rapid progression of liver disease in HCV/HIV co-infected individuals (a common occurrence among injection drug users who now represent the primary mode of HCV infection in the US). Although human liver biopsy studies remain a high priority, we anticipate that the analyses of certain components of these projects may need to be put on hold until adequate specimens become available (e.g. we do not yet have adequate numbers of specimens from patients with later stage 3 and 4 fibrosis in the transplant model). Thus we propose to incorporate complimentary proteomic analyses utilizing 1) additional clinical samples (human serum or peripheral blood mononuclear cells-PBMC's) for the identification of secreted biomarkers, etc and 2) serum and human liver tissue from mice containing chimeric human livers that support prolonged HCV infection with high viral titers for studying human HCV in vivo. Taken together, the final results of these studies should increase our understanding of the mechanisms of HCV replication and pathogenesis and may provide critical markers for both diagnostic and prognostic applications as well as suggest novel targets for therapeutic and prophylactic intervention.
Project Details
Start Date
2005-04-20
End Date
2008-04-20
Status
Closed
Released Data Link
Team
Principal Investigator
Team Members
Related Publications
Diamond D.L., A.J. Syder, J.M. Jacobs, C.M. Sorensen, K.A. Walters, S.C. Proll, J.E. McDermott, M.A. Gritsenko, Q. Zhang, R. Zhao, T.O. Metz, D.G. Camp 2nd, K.M. Waters, R.D. Smith, C.M. Rice, and M.G. Katze. 2010. Temporal proteome and lipidome profiles reveal hepatitis C virus-associated reprogramming of hepatocellular metabolism and bioenergetics. PLoS Pathog. 6:e1000719. Epub 2010 Jan 8
Proteomic profiling of human liver biopsies: hepatitis C virus-induced fibrosis and mitochondrial dysfunction.
Diamond DL, Jacobs JM, Paeper B, Proll SC, Gritsenko MA, Carithers RL Jr, Larson AM, Yeh MM, Camp DG 2nd, Smith RD, Katze MG.
Hepatology. 2007 Sep;46(3):649-57