Interferon (IFN) alpha inhibits hepatitis C virus (HCV) replication both clinically and in vitro; however, the complete spectrum of interferon-stimulated genes (ISGs) expressed in the HCV-infected liver or the genes responsible for control of HCV replication have not been defined. To better define ISG expression in the chronically infected HCV liver, DNA microarray analysis was performed on 9 individuals with chronic hepatitis C (CHC). A total of 232 messenger RNAs were differentially regulated in CHC compared with nondiseased liver controls. A significant proportion of these were potential ISGs that were transcriptionally elevated, suggesting an ongoing response to endogenous IFN and/or double-stranded RNA. One ISG significantly elevated in all patients was viperin, an evolutionary conserved ISG that has antiviral activity against human cytomegalovirus. Stimulation of Huh-7 and HepG2 cells with IFN-alpha or -gamma revealed viperin is predominantly a type I ISG. Furthermore, viperin expression could also be induced following transfection of Huh-7 cells with either poly(I:C) or HCV RNA. Transient expression of viperin in cells harboring the HCV genomic replicon resulted in a significant decrease in HCV replication, suggesting that viperin has anti-HCV activity. In conclusion, even in the face of a persistent HCV infection, there is an active ISG antiviral cellular response, highlighting the complexity of the host viral relationship. Furthermore, ISG viperin has anti-HCV activity in vitro; we postulate that viperin, along with other ISGs, acts to limit HCV replication.