Methotrexate is an antimetabolic drug with a myriad of serious side effects including nephrotoxicity, which presumably occurs due to oxidative tissue damage. Here, we evaluated the potential protective effect of lycopene, a potent antioxidant carotenoid, given in two different pharmaceutical forms in methotrexate-induced kidney damage in rats. Serum biochemical (urea and creatinine) and tissue oxidative damage markers and histopathological kidney changes were evaluated after systemic administration of both lycopene dissolved in corn oil and lycopene encapsulated in nanoliposomes. Similar to previous studies, single dose of methotrexate induced severe functional and morphological alterations of kidneys with cell desquamation, tubular vacuolation, and focal necrosis, which were followed by serum urea and creatinine increase and disturbances of tissue antioxidant status. Application of both forms of lycopene concomitantly with methotrexate ameliorated changes in serum urea and creatinine and oxidative damage markers and markedly reversed structural changes of kidney tissue. Moreover, animals that received lycopene in nanoliposome-encapsulated form showed higher degree of recovery than those treated with free lycopene form. The findings of this study indicate that treatment with nanoliposome-encapsulated lycopene comparing to lycopene in standard vehicle has an advantage as it more efficiently reduces methotrexate-induced kidney dysfunction.