Abstract Abnormal or inadequate vasculogenesis, local inflammation and severe epithelial damage are common features of both inflammatory bowel disease (IBD) and irradiation injury after pelvic or abdominal cancer treatment. Several studies have shown that adult bone marrow-derived stem cells, upon transplantation, home to the damaged digestive tissue and facilitate mucosal repair in both IBD and radiation injury. However, despite increasing evidence that endothelial progenitor cells (EPC) represent a promising tool for ischemic cardiac and vascular repair, few have investigated whether transplanted EPC can contribute to the intestinal vasculogenic process and/or the stem or mature epithelial cell pool. In order to study the intrinsic ability of human EPC to contribute to the epithelial or vascular bed of the small intestine, we transplanted 13 pre-immune 55–60 day old fetal sheep with 0.5–2.6x106 human EPC/fetus and examined the contribution of these cells to the intestinal architecture. CB-derived EPC were obtained as previously described (Ingram et al. Blood:104,2004) and transduced with a retroviral vector expressing DsRed. Recipients were evaluated at 85 days post-transplant for the presence of donor (human)- specific cell types by confocal microscopy. We found that within the intestine, EPC, as detected by DsRed positivity, localized preferentially to the mucosal layer above the muscularis mucosa in the area of the crypts of Lieberkühn. The overall levels of EPC engraftment positively correlated with the cell dose administered (p<0.05) such that the levels of DsRed positive cells found within the mucosal layer of animals in each transplant group were as follows: 7.6±0.5% in the animal transplanted with 5x105cells; 8±0.3% in those transplanted with 1.5x106 cells (n=8), and 10.9±0.5% in sheep transplanted with 2.6x106cell/fetus (n=4). Immunostaining with vWF and CD31 demonstrated that only 0.7–1.7% of the DsRed cells retained a EPC phenotype, thus suggesting that that the majority of the transplanted cells had adopted an alternative fate. Double positivity for DsRed and Cytokeratin 20, a major cellular protein present in mature enterocytes, was found in 1.12±0.02% of the cells, all of which were found in the villi area. Colocalization of DsRed cells with expression of Musashi a putative marker for intestinal stem cells was also evaluated. These analyses revealed that 10.79 ±0.01% of the epithelial cells within the crypts were DsRed positive and thus donor derived, and all expressed Musashi. Expression of this marker was not observed in any DsRed positive cells in any other location within the intestine. In addition DsRed positive cells that did not co-express any of the markers tested thus far were found in the stromal layer adjacent to the crypts. Further characterization of these cells is currently underway. In conclusion, these are the first studies, to our knowledge, to show that EPC can contribute significantly to the developing intestinal mucosa and the intestinal stem cell pool, and EPC may thus represent a valuable cell source for intestinal regeneration and repair.