Non-neoplastic tissues around an abdomino-pelvic tumor can be damaged by the radiotherapy protocol, leading to chronic gastrointestinal complications that affect the quality of life with substantial mortality. Stem cell-based approaches using immunosuppressive bone marrow mesenchymal stem cells (MSCs) are promising cell therapy tools. In a rat model of radiation proctitis, we evidenced that a single MSC injection reduces colonic mucosa damages induced by ionizing radiation with improvement of the re-epithelization process for up to 21 days. Immune cell infiltrate and inflammatory molecule expressions in the colonic mucosa were investigated. We report that MSC therapy specifically reduces T-cell infiltration and proliferation, and increases apoptosis of radiation-activated T cells. We assessed the underlying molecular mechanisms and found that interleukin-10 and regulatory T lymphocytes are not involved in the immunosuppressive process in this model. However, an increased level of corticosterone secretion and HSD11b1 (11β-hydroxysteroid dehydrogenase type 1)-steroidogenic enzyme expression was detected in colonic mucosa 21 days after MSC treatment. Moreover, blocking the glucocorticoid (GC) receptor using the RU486 molecule statistically enhances the allogenic lymphocyte proliferation inhibited by MSCs in vitro and abrogates the mucosal protection induced by MSC treatment in vivo. Using the irradiation model, we found evidence for a new MSC immunosuppressive mechanism involving GCs.