Liver fibrosis is the consequence of activation of hepatic stellate cells mediated by persistent or recurrent liver injury, where oxidative stress or inflammatory response resulting from immune cells and cytokines are involved. Targeting of hepatic stellate cells could be an important strategy for the therapy of liver fibrosis. In this study, we showed a tropism of recombinant adeno-associated virus (rAAV, serotype 2) with high efficiency in transduction of a homeostatic gene, heme oxygenase-1 (HO-1), to activated stellate cells. The binding of rAAVs to stellate cells increased significantly after serum-stimulated activation compared with quiescent status. Portal injection of rAAVs to normal or carbon tetrachloride (CCl(4))-induced liver fibrosis showed a distinct distribution of rAAV binding. The majority of injected rAAVs bound to the cells in fibrotic areas that were associated with higher expression levels of fibroblast growth factor receptor-1alpha at 2 hours after administration. Isolation of different types of cells from CCl(4)-induced fibrotic livers showed predominant expression of transgene in stellate cells after rAAV/HO-1 administration on day 3 and remained stable for 12 weeks. In addition, HO-1-transduced stellate cells showed reduced transcript levels of type 1 collagen and impaired proliferative ability compared with controls. With this approach, the severity of established micronodular cirrhosis was markedly reduced. In conclusion, these findings suggest a new approach for the treatment of liver fibrosis using adeno-associated virus-mediated gene transfer.