Prolonged use of an antineoplastic agent methotrexate (MTX), can cause numerous side effects such as nephrotoxicity. The aim of this study was to examine the effects of MTX on kidneys and demonstrate the protective effects of gallic acid (GA). Twenty-four, male, rats distributed into three groups. Each groups consisted eight rats and only saline was administered to the control group. The MTX group received a single dose (20 mg/kg) MTX intraperitoneally. The MTX þ GA group received same dose MTX and 100 mg/kg GA orally during the 7 days. Renal functions, oxidative stress markers, histopathological and immunohistochemical changes were evaluated at the end of the experiment. Blood urea nitrogen, creatinine, uric acid levels and tissue oxidative stress markers, total oxidant status and oxidative stress index levels significantly increased and total antioxidant status levels significantly decreased in MTX group compared with the control group. At the histopathological examination hemorrhages, tubular cell necrosis, glomerulosclerosis, inflammatory cell infiltrations and proteinous materials in tubules were noticed in MTX group. Immunohistochemical examination revealed that increased expressions of serum amyloid A (SAA), tumor necrosis factor alpha (TNF-a), prostaglandin E2 (PGE-2) and Creactive protein (CRP) in tubular epithelial cells of kidneys in this group. There were no immunoreaction with SAA and CRP, only small number of PGE-2 and TNF-a positive tubular epithelial cells were observed in MTX þ GA group. In conclusion, all evidence suggested that oxidative stress caused MTX-induced nephrotoxicity and GA prevent the kidney from the nephrotoxicity due to its antioxidant and anti-inflammatory activities.