Abstract 282: Human CD133+ Progenitor Cells Promote the Cicatrisation of Diabetic Ischemic Ulcers through Paracrine Stimulation of Reepithelization
Stem cells hold promises for the cure of diabetic ulcers, but overlaid ischemia may limit their efficacy. We aimed to test the healing property of fetal CD133+ progenitor cells in a new model that consists of causing limb excisional ulcers with overlaid ischemia (femoral artery ligation) in streptozotocin-induced diabetic mice. Ischemic ulcers showed delayed cicatrisation, being 3.72-fold larger than non-ischemic ulcers, 7 days after wounding (P<0.01). In further studies, ischemic wounds were covered with collagen, collagen-embedded CD133+CD34+CD31−(CD133+, 2×104) or CD133−CD34+CD31+ fetal cells (CD133−, 2×104), or adult endothelial progenitor cells (EPC, 8×104). Contralateral ischemic ulcer received collagen. Transplantation of CD133+ accelerated wound closure (treated/contralateral ratio: 0.95±0.05, 0.76±0.06 and 0.70±0.08 at day 0, 3 and 7) as compared with CD133− (0.98±0.06, 1.05±0.2, and 1.86±0.5), EPC (0.99±0.07, 1.38±0.2, and 1.22±0.2) or collagen (0.98±0.07, 1.04±0.1, and 1.6±0.5; p±0.05 for both treatment and time). Few human nuclear antigen-positive cells were detected in CD133+ or CD133− treated ulcers (5.4±1.8 and 4.3±2.4 cells/mm2, respectively). Epidermal cell proliferation (PCNA immunohistochemistry) was higher in wounds given CD133+ or CD133− (p<0.001 vs collagen), while endothelial cell (EC) proliferation was marginally increased by CD133+ only (p=0.09 vs collagen or CD133−). Cultured CD133+ release large amounts of VEGF-A in culture medium (CM) (1350±52 vs 72±15 pg/ml in the CD133− CM, p<0.01). In vitro, either CD133+ or CD133− conditioned CM promoted keratinocyte proliferation (p<0.001 and p<0.01, respectively), but only CD133+ conditioned CM stimulated EC proliferation (p<0.01). These effects were reduced by the PI3K inhibitor, wortmannin (p<0.01). Under high glucose condition, CD133+ conditioned CM stimulated keratinocyte migration and prevented both EC apoptosis and ROS production by EC. We have shown that transplantation of a low number of human fetal CD133+ stimulates the healing of ischemic diabetic ulcers, while a 4-fold larger number of adult EPC is ineffective. The therapeutic action is attributable to paracrine stimulation of reepithelization, possibly via a VEGF-A, PI3K-mediated mechanism.