Hepatitis C virus (HCV) infection is still a major global health issue despite decades of research. The liver-specific microRNA-122 (miR-122) can stimulate HCV replication/translation in vitro, indicating that miR-122 contributes to pathogenesis of HCV. However, it remains controversial whether interferon (IFN) inhibits HCV via modulating miR-122 expression. The underlying mechanism of ribavirin (RBV) in enhancing IFN treatment for HCV patients has yet to be explored. We investigated the relationship between miR-122 expression and anti-HCV activity of IFN beta in combination with RBV in vitro, due to difficulty accessing an HCV animal model. Upregulation of ISG54 mRNA or cytostatic effect was detected in Huh7 and HCV replicon cell lines in response to IFN beta or RBV stimulation, respectively. It was found that IFN beta and/or RBV suppressed miR-122 expression marginally, with a synergetic anti-HCV effect between IFN beta and RBV. Marginal modification of other miRNAs was also observed in these cell lines, using miRNA array following IFN beta and RBV treatment. Taken together, our data suggest that miRNAs are not crucial in anti-HCV action, following IFN beta and/or RBV stimulation in vitro.