Viral Hepatitis

Proteomic profiling of human liver biopsies: Hepatitis C virus–induced fibrosis and mitochondrial dysfunction

Diamond, Deborah L.^1*; Jacobs, Jon M.²; Paeper, Bryan¹; Proll, Sean C.¹; Gritsenko, Marina A.²; Carithers, Robert L. Jr.³; Larson, Anne M.³; Yeh, Matthew M.⁴; Camp, David G. II²; Smith, Richard D.²; Katze, Michael G.^1,5

Author Information

¹Department of Microbiology, University of Washington, Seattle, WA

²Biological Sciences Division, Environmental Molecular Sciences Laboratory, Pacific Northwest National Laboratory Richland, WA

³Hepatology Section, Department of Medicine, University of Washington, Seattle, WA

⁴Department of Pathology, School of Medicine, University of Washington, Seattle, WA

⁵Washington National Primate Research Center, University of Washington, Seattle, WA

^*Department of Microbiology, University of Washington, Box 358070, Seattle, WA 98195-8070

Email:[email protected]

Received 21 December 2006; Accepted 28 March 2007

Published online in Wiley InterScience (www.interscience.wiley.com).

Grant sponsor: National Institute on Drug Abuse; Grant Number: 1P30DA01562501; Grant sponsor: National Center for Research Resource; Grant Number: RR18522; Grant sponsor: Battelle Memorial Institute for the DOE; Grant Number: DE-AC05-76RLO-1830.

Potential conflict of interest: Nothing to report.

fax: 206-732-6055

Hepatology 46(3):p 649-657, September 2007. | DOI: 10.1002/hep.21751

Abstract

Liver biopsies from hepatitis C virus (HCV)-infected patients offer the unique opportunity to study human liver biology and diseasein vivo. However, the low protein yields associated with these small samples present a significant challenge for proteomic analysis. In this study we describe the application of an ultrasensitive proteomics platform for performing robust quantitative proteomic studies on microgram amounts of HCV-infected human liver tissue from 15 patients at different stages of fibrosis. A high-quality liver protein database containing 5,920 unique protein identifications supported high throughput quantitative studies using ¹⁶O/¹⁸O stable isotope labeling in combination with the accurate mass and time (AMT) tag approach. A total of 1,641 liver biopsy proteins were quantified, and analysis of variance (ANOVA) identified 210 proteins exhibiting statistically significant differences associated with fibrosis stage. Hierarchical clustering showed that biopsies representative of later fibrosis stages (for example, Batts-Ludwig stages 3–4) exhibited a distinct protein expression profile, indicating an apparent down-regulation of many proteins when compared with samples from earlier fibrosis stages (for example, Batts-Ludwig stages 0–2). Functional analysis of these signature proteins suggests that impairment of key mitochondrial processes including fatty acid oxidation and oxidative phosphorylation, and response to oxidative stress and reactive oxygen species occurs during advanced stage 3 to 4 fibrosis.Conclusion:The results reported here represent a significant advancement in clinical proteomics providing to our knowledge, the first demonstration of global proteomic alterations accompanying liver disease progression in patients chronically infected with HCV. Our findings contribute to a generally emerging theme associating oxidative stress and hepatic mitochondrial dysfunction with HCV pathogenesis. (Hepatology 2007.)

Abbreviations: AMT, accurate mass and time; ANOVA, analysis of variance; ER, endoplasmic reticulum; FTICR, Fourier transform ion cyclotron resonance; HCV, hepatitis C virus; LC, liquid chromatography; MS/MS, tandem mass spectrometry; NET, LC-normalized elution time; Sp1, specificity protein 1.