Combination of stromal-derived factor-1α and vascular endothelial growth factor gene-modified endothelial progenitor cells is more effective for ischemic neovascularization

Background: Recruitment and entrapment of bone marrow-derived endothelial progenitor cells (EPCs) is important in vascular endothelial growth factor (V-EGF)-induced angiogenesis. EPC mobilization and differentiation are modulated by stromal-derived factor-1 alpha (SDF-1 alpha/CXCL12), another important chemokine. In this study, we investigated the hypothesis that SDF-1 alpha and VEGF might act synergistically on EPC-mediated vasculogenesis. Methods: EPCs were isolated and cultured from human peripheral blood, then transduced with retroviral vectors pBabe containing human VEGF(165) complimentary DNA (Td/V-EPCs) and pBabe wild-type (Td/p-EPCs). EPC migration activity was investigated with a modified Boyden chamber assay. EPC apoptosis induced by serum starvation was studied by annexin V assays. The combined effect of local administration of SDF-1 alpha and Td/V-EPC transplantation on neovascularization was investigated in a murine model of hind limb ischemia. Results. Over-expression of hV-EGF(165) increased SDF-1 alpha-mediated EPC migration. SDF-1 alpha-mediated migration was significantly increased when EPCs were modified with VEGF (Td/V-EPCs) vs when VEGF was not present (Td/p-EPCs) or when VEGF alone was present (Td/V-EPCs; 196.8 +/- 15.2, 81.2 +/- 9.8, and 67.4 +/- 7.4/mm(2), respectively P < .001). SDF-1 alpha combined with VEGF reduced serum starvation-induced apoptosis of EPCs more than SDF-1 alpha or VEGF alone (P < .001). To determine the effect of this combination in vivo, SDF-1 alpha was locally injected alone into the ischemic hind limb muscle of nude mice or combined with systemically injected Td/V-EPCs. The SDF-1 alpha plus VEGF group showed significantly increased local accumulation of EPCs, blood-flow recovery, and capillary density compared with the other groups. The ratio of ischemic/normal blood flow in Td/V-EPCs plus SDF-1 alpha group was significantly higher (P < .01), as was capillary density (capillaries/mm(2)), an index of neovascularization (Td/V-EPCs plus SDF-1 alpha group, 863 31; no treatment, 395 +/- 13; SDF-1 alpha, 520 +/- 29; Td/p-EPCs, 448 +/- 28; Td/p-EPCs plus SDF-1 alpha., 620 +/- 29; Td/V-EPCs, 570 +/- 30; P < .01). To investigate a possible mechanistic basis, we showed that VEGF up-regulated the receptor for SDF-1 alpha, CXCR4, on EM in Nitro. Conclusion: The combination of SDF-1 alpha and VEGF greatly increases EPC-mediated angiogenesis. The use VEGF and SDF-1 alpha together, rather than alone, will be a novel and efficient angiogenesis strategy to provide therapeutic neovascularization. (J Vase Surg 2009;50:608-16.)