Abstract BACKGROUND: Micromeria croatica (Pers.) Schott is an aromatic plant from Lamiaceae family previously found to possess potent in vitro antioxidant activity which is mainly attributed to the high level of polyphenolic substances. The aim of this study was to investigate the hepatoprotective activity and possible underlying mechanisms of Micromeria croatica ethanolic extract (MC) using a model of carbon tetrachloride (CCl4)-induced liver injury in mice. METHODS: Male BALB/cN mice were randomly divided into seven groups: control group received saline, MC group received ethanolic extract of M. croatica in 5% DMSO (100 mg/kg b.w., p.o.), and CCl4 group was administered CCl4 dissolved in corn oil (2 mL/kg, 10% v/v, ip). MC50, MC200 and MC400 groups were treated with MC orally at doses of 50, 200 and 400 mg/kg once daily for 2 consecutive days, respectively, 6 h after CCl4 intoxication. The reference group received silymarin at dose of 400 mg/kg. At the end of experiment, blood and liver samples were collected for biochemical, histopathological, immunohistochemical and Western blot analyses. In addition, major phenolic compounds in MC were quantified by HPLC-DAD. RESULTS: CCl4 intoxication resulted in liver cells damage and oxidative stress and triggered inflammatory response in mice livers. MC treatment decreased ALT activity and prevented liver necrosis. Improved hepatic antioxidant status was evident by increased Cu/Zn SOD activity and decreased 4-hydroxynonenal (4-HNE) formation in the livers. Concomitantly, nuclear factor erythroid 2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) were overexpressed. The hepatoprotective activity of MC was accompanied by the increase in nuclear factor-kappaB (NF-κB) activation and tumor necrosis factor-alpha (TNF-α) expression, indicating amelioration of hepatic inflammation. Additionally, MC prevented tumor growth factor-β1 (TGF-β1) and α-smooth muscle actin (α-SMA) expression, suggesting the potential for suppression of hepatic fibrogenesis. CONCLUSION: These results of the present study demonstrated that MC possesses in vivo antioxidant and anti-inflammatory activity and exhibits antifibrotic potential, which are comparable to those of standard hepatoprotective compound silymarin.