Objective: The aim of our study was to examine whether irradiation combined with pemetrexed can exacerbate pulmonary injury. Methods: Two groups of male Wister Rats were subjected to bilateral apex of lungs irradiation(a single dose of 12 Gy), with or without pemetrexed(20 mg/kg) by intraperitoneal injection at the same time; a third group of weightand age- matched animals were treated with pemetrexed alone, as the same dose scheme, time and root of injection. The fourth group served as control. The whole lung mounts were dissected to histological evaluation, while serum cytokine transforming growth factor-β1(TGF-β1) analysis were compared at 1, 7, 21, 35, 49 days post-irradiation after irradiation. Results: Histological examination showed a thickening of alveolar septal, accumulation of inflammatory cells. The irradiation treatment group and the radiation-chemo treatment group showed a statistically significant higher level of TGF-β1(P < 0.05) than other two groups, but there were no differences between these two irradiation groups. Conclusion: These results demonstrated that pemetrexed can not aggravate pulmonary injury and it could be safely used in concurrent or sequential radio-chemotherapy in lung adenocarcinoma. Methods: Two groups of male Wister rats were subjected to bilateral apex of lungs irradiation (a single dose of 12 Gy), with or without pemetrexed ( 20 mg / kg) by intraperitoneal injection at the same time; a third group of weight and age- matched animals were treated with pemetrexed alone, as the same dose scheme, time and root of injection. The fourth group served as control. The whole lung mounts were dissected to histological evaluation, while serum cytokine transforming growth factor-β1 (TGF-β1) analysis were compared at 1, 7, 21, 35, 49 days post-irradiation after irradiation. Results: Histological examination showed thickening of alveolar septal , the treatment of inflammatory cells. The irradiation treatment group and the radiation-chemo treatment group showed a statistically significant higher level of TGF-β1 (P <0.05) than the other two groups, but there were no differences between these two irradiation groups. Conclusion: These results in that pemetrexed can not aggravate pulmonary injury and it could be safely used in concurrent or sequential radio-chemotherapy in lung adenocarcinoma.