Mast cell hyperplasia is a characteristic feature of many inflammatory and fibrotic conditions, including intestinal radiation injury (radiation enteropathy). This study used mast cell-deficient rats to define the role of mast cells in the mechanisms underlying early radiation-induced mucosal injury and delayed intestinal wall fibrosis. Mast cell-deficient (Ws/Ws) mutant rats and mast cell-competent (+/+) littermates were used. A 4-cm loop of ileum was exposed to 21 Gy single-dose radiation. Irradiated and unirradiated intestine were examined at 2 or 26 weeks using quantitative histology and morphometry. Quantitative immunohistochemistry was used to assess transforming growth factor beta (Tgfb), myeloperoxidase, and epithelial and smooth muscle cell proliferation. Collagen content was measured colorimetrically, and steady-state Tgfb1 mRNA was determined with fluorogenic probe RT-PCR. Compared to +/+ rats, Ws/Ws animals exhibited strikingly exacerbated mucosal injury but minimal reactive intestinal wall fibrosis. Ws/Ws rats exhibited less radiation-induced intestinal smooth muscle cell proliferation and collagen accumulation than +/+ littermates. Tgfb expression increased to a similar extent in Ws/Ws and +/+ rats. Unirradiated intestine from Ws/Ws and +/+ rats did not differ significantly. Mast cells protected the intestinal mucosa during the early phase of radiation enteropathy and promoted intestinal fibrosis after the breakdown of the mucosal barrier. Mast cells may be required for Tgfb to exert its full fibrogenic effect in radiation enteropathy.