Several growth factors are trophic for the gastrointestinal tract and able to reduce the degree of intestinal damage caused by cytotoxic agents. However, studies of epidermal growth factor (EGF) for chemotherapy-induced intestinal injury are conflicting. The development of a transgenic mouse that specifically overexpresses EGF in the small intestine provided a unique opportunity to assess the contribution of EGF in mucositis. After a course of fluorouracil, transgenic mice fared no better than control mice. Weight recovery was inferior, and mucosal architecture was not preserved. Apoptosis was not decreased and proliferation was not increased in the crypts. To corroborate the findings in transgenic mice, ICR mice were treated with exogenous EGF after receiving fluorouracil. Despite ileal upregulation of native and activated EGF receptor, the mice were not protected from intestinal damage. No benefits were observed with different EGF doses or schedules or routes of EGF administration. Finally, mucositis was induced in mutant mice with specific defects of the EGF signaling axis. Compared with control mice, clinical and histological parameters of intestinal injury after fluorouracil were no different in waved-2 mice, which have functionally diminished EGF receptors, or waved-1 mice, which lack transforming growth factor-, another major ligand for the EGF receptor. These findings do not support a critical role for EGF or its receptor in chemotherapy-induced intestinal injury.