The metabolic regulator PGC-1a links hepatitis C virus infection to hepatic insulin resistance Amir Shlomai 1 , Maya Mouler Rechtman 2 , Ela Olga Burdelova 1 , Alona Zilberberg 2 , Sarit Hoffman 3 , Irit Solar 3 , Sigal Fishman 1 , Zamir Halpern 1 , Ella H. Sklan 2,⇑ 1 The Research Center for Digestive Tract and Liver Diseases, Tel-Aviv Sourasky Medical Center and The Sackler Faculty of Medicine, Tel-Aviv University, Tel-Aviv, Israel; 2 Department of Clinical Microbiology and Immunology, Sackler Faculty of Medicine, Tel-Aviv University, Tel-Aviv 69978, Israel; 3 The Institute of Pathology and Cancer Research, Tel-Aviv Sourasky Medical Center, Tel-Aviv, Israel Background & Aims: Chronic hepatitis C virus (HCV) infection is strongly associated with insulin resistance and diabetes mellitus. Peroxisome proliferator-activated receptor-gamma co-activator 1a (PGC-1a) is a transcriptional co-activator involved in the initi- ation of gluconeogenesis in the liver. Increased hepatic expression of PGC-1a has been implicated in insulin resistance. We investi- gated whether modulation of PGC-1a levels following HCV infection underlies HCV-associated hepatic insulin resistance. Methods: HCV genomes were expressed in hepatoma cells fol- lowed by analysis of PGC-1a and gluconeogenesis levels. Results: PGC-1a was robustly induced in HCV infected cells. PGC-1a induction was accompanied by an elevated expression of the gluconeogenic gene glucose-6 phosphatase (G6Pase) and increased glucose production. The induction of gluconeogenesis is HCV dependent, since interferon treatment abolishes PGC-1a and G6Pase elevation and decreases glucose output. Moreover, PGC-1a knockdown resulted in a significant reduction of G6Pase levels in HCV full length replicon cells, emphasizing the central role of PGC-1a in the exaggerated gluconeogenic response observed in HCV patients. Treatment of HCV replicon cells with the antioxidant N-acetylcysteine resulted in reduction of PGC- 1a levels, suggesting that HCV-induced oxidative stress pro- moted PGC-1a upregulation. Finally, both PGC-1a and G6Pase RNA levels were significantly elevated in liver samples of HCV infected patients, highlighting the clinical relevance of these results. Conclusions: PGC-1a is robustly induced following HCV infec- tion, resulting in an upregulated gluconeogenic response, thereby linking HCV infection to hepatic insulin resistance. Our results suggest that PGC-1a is a potential molecular target for the treat- ment of HCV-associated insulin resistance. Ó 2012 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved. Introduction Chronic hepatitis C virus (HCV) infection is a major global health problem. Chronically infected patients may develop cirrhosis and liver cancer that are implicated in substantial morbidity and mor- tality [1]. Although the liver is the target organ for HCV infection, extrahepatic manifestations of HCV, including insulin resistance and diabetes mellitus, are frequently encountered in the clinical setting [2]. Insulin resistance was found to be a specific feature of chronic HCV associated with genotypes 1 and 4 and with high serum viral RNA levels [3]. Insulin resistance and diabetes melli- tus also correlated with poor response to anti-HCV therapy, while eradication of the virus correlates with a marked improvement in insulin resistance [4]. The molecular mechanism by which HCV promotes insulin resistance is still undetermined. HCV infection was found to cause a postreceptor defect in the insulin signaling cascade [5]. In addition, the activation of proinflammatory mediators, such as nuclear factor-kappa-B and tumor necrosis factor-a, caused by the chronic infection state, interferes with insulin signaling and contributes to both hepatic and peripheral insulin resistance [6,7]. In agreement with the evidence suggesting a central role for reactive oxygen species in the development of insulin resistance [8], HCV infection is known to promote cellular oxidative stress through multiple mechanisms, including chronic inflammation, iron overload, and liver injury. Some of the HCV proteins were reported to directly contribute to this process [9,10]. Peroxisome proliferator-activated receptor gamma co-activa- tor 1a (PGC-1a) is a transcription co-activator and a master reg- ulator of gluconeogenesis that functions via interaction with transcription factors located on the promoters of gluconeogenic genes [11–13]. Although PGC-1a is scarcely detectable in the Journal of Hepatology 2012 vol. 57 j 867–873 Keywords: Hepatitis C virus; Peroxisome proliferator-activated receptor gamma co-activator 1a; Diabetes; Insulin resistance; Oxidative stress. Received 17 December 2011; received in revised form 19 May 2012; accepted 14 June 2012; available online 23 June 2012 ⇑ Corresponding author. Tel.: +972 3 6408197; fax: +972 3 6409160. E-mail address: sklan@post.tau.ac.il (E.H. Sklan). Abbreviations: HCV, hepatitis C virus; PGC-1a, peroxisome proliferator-activated receptor gamma, co-activator 1a; G6Pase, glucose-6 phosphatase; RNA, ribonu- cleic acid; PI3, phosphatidylinositol 3-kinase; FLRP, full length replicon; SGR, subgenomic replicon; NS5A, non-structural protein 5A; SEAP, secreted alkaline phosphatase; NS, non-structural; SDS, sodium dodecyl sulfate; RIPA, radio im- munoprecipitation assay; SDS-PAGE, sodium dodecyl sulfate polyacrylamide gel electrophoresis; PBS, phosphate buffered saline; HPRT, hypoxanthine phospho- ribosyltransferase; HA, hemagglutinin; ECL, enhanced chemiluminescence; IFN, interferon; SOCS, suppressors of cytokine signaling; FOXO1, forkhead box protein O1; ER, endoplasmic reticulum; JNK, c-Jun N terminal kinase; NAC, N-acetylcys- teine; IRS-1, insulin receptor substrate-1. Research Article