Our goal is to dissect the mechanisms underlying the development of thyroiditis and diabetes in patients with chronic hepatitis C virus (HCV) infection in order to design new treatment and prevention modalities. Nearly 2.7 million individuals in the US have chronic hepatitis C, resulting in up to 13,000 deaths per year. Moreover, the prevalence of HCV infection is significantly higher among veterans; thus, chronic HCV is a major problem in VA hospitals. However, both HCV infection itself and its therapy (IFNa) frequently trigger thyroiditis and diabetes, thereby complicating the management of these patients. In the previous grant cycle, we made significant progress in dissecting the mechanisms by which thyroiditis and diabetes develop in HCV patients. We demonstrated that HCV and IFNa trigger thyroiditis by direct thyroid toxic effects and through epigenetic modifications in thyroiditis-susceptibility genes. In addition, we were able, for the first time, to infect human thyroid cells with HCV. Moreover, we recently discovered that IFNa also causes epigenetic changes in islet cells, suggesting a new mechanism for inducing diabetes in HCV patients (see Progress Report). In the current proposal, we will build on these findings and dissect the mechanisms by which HCV and IFNa trigger thyroiditis and diabetes. Our hypothesis is that patients with HCV infection treated with IFNa develop thyroiditis and/or diabetes due to direct epigenetic modifications of thyroiditis and diabetes-susceptibility genes in thyroid cells and islet beta cells, respectively. Our specific aims are: Specific Aim 1: we will test in vitro genome-wide epigenetic modifications induced in human thyroid cells infected with HCV and exposed to IFNa; we will also evaluate epigenetic effects in the thyroid in vivo by creating a new transgenic mouse with tetracycline-inducible IFNa expression in the thyroid. Specific Aim 2: we will test the hypothesis that HCV and IFNa have direct toxic effects on beta cells through epigenetic modifications, causing diabetes in HCV-infected patients. In collaboration with Dr. Jason Blackard's group from UC, we will examine the epigenetic effects of exposure of a human beta cell line to HCV viral proteins or infectious virions, as well as the epigenetic modifications induce by IFNa in human beta cells. Specific Aim 3: we will test systemic effects of IFNa in other tissues, in addition to the thyroid and pancreas, by treating mice with IFNa systemically, and examining its effects on global methylation, histone modifications, and miR expression in selected tissues (blood, liver, fat, and muscle). In summary, our proposed studies will dissect the molecular mechanisms causing thyroiditis and diabetes in chronic hepatitis C patients receiving IFNa. Chronic hepatitis C infection is a major health problem for veterans, and thyroiditis and diabetes are common co-morbid conditions in these patients that significantly interfere with effective treatment of chroni HCV. Understanding the etiology of thyroiditis and diabetes in chronic HCV patients will enable the development of novel treatment modalities and prevention strategies based on the mechanisms triggering diabetes and thyroiditis in these patients. These new prevention and treatment strategies for thyroiditis and diabetes in HCV patients will facilitate the treatment and cure of chronic hepatitis C in veterans and non-veterans.