Peroxisome proliferator-activated receptor- γ (PPAR- γ) has been demonstrated to protect against non – alcoholic steatohepatitis, both cyclophosphamide and paracetamol-induced hepatotoxicity. The present study was carried out to evaluate the possible protective effect of a PPAR- γ agonist, pioglitazone (PIO), in a model of hepatotoxicity induced by methotrexate (MTX) in rats. Twenty-four male rats were divided randomly into four groups (6 rats each): control with vehicle; MTX with vehicle or PIO; or MTX with PIO plus GW9662 as a specific PPAR- γ antagonist. Vehicle or PIO or PIO plus GW9662 were administered orally for 2 weeks and on the 11th day MTX was administered. The results displayed the efficiency of PIO to restore the oxidative / antioxidative balance, which was disturbed in MTX treated rats, through attenuation of hepatic malondialdehyde (MDA) level as index for oxidative stress and enhancement of antioxidant; reduced glutathione (GSH), superoxide dismutase (SOD) and catalase (CAT). Additionally, PIO pretreatment mitigate the serum level of alanine aminotransferase(ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), γ- glutamyl transferase (γ-GT)and the pro - inflammatory cytokines; tumor necrosis factor –α (TNF- α) and interleukin- 1 beta(IL-1β)which were increased in MTX treated rats. Moreover, PIO was able to decrease the increased caspase -3 activity in MTX treated group. Histopathological evaluation confirmed the protective effect of PIO supplementation against MTX - induced liver injury. Co- administration of GW9662 with PIO prevents the hepatoprotective effect of PIO in MTX treated rats. It can be concluded from the aforementioned results that PIO might be valuable for protection against hepatotoxicity induced by MTX via its antioxidant, anti-inflammatory and antiapoptotic potential and this beneficial effect is PPAR –γ dependent.