Vascular endothelial growth factor-C derived from CD11b+ cells induces therapeutic improvements in a murine model of hind limb ischemia

ObjectiveThe use of bone marrow cells (BMCs) in therapeutic angiogenesis has been studied extensively. However, the critical paracrine effects of this treatment are still unclear. Therefore, we studied autotransfusable cells that produce vascular endothelial growth factor (VEGF), especially VEGF-C.MethodsMale C57BL/6 mice with hind limb ischemia were administered intramuscular injections of phosphate-buffered saline as controls, or unsorted BMCs, sorted CD11b+, or CD11b− cells from BMCs, and recombinant VEGF-C. To evaluate the treatments, perfusion was measured by laser Doppler scanning performed on days 0, 1, 3, 7, 14, 21, and 28. A functional assay was performed in parallel, with mice traversing an enclosed walkway. Capillary density was determined by directly counting vessels stained positive with von Willebrand factor at individual time points. Lymphangiogenesis was assessed by LYVE-1 positive cells.ResultsPostischemic recovery of hind limb perfusion significantly improved in BMC, CD11b+, and VEGF-C treatment groups compared with the control groups, as assessed by laser Doppler scanning. On early operative days 1 and 3, the blood flow recovery ratio was higher in the CD11b+-treated group compared with BMC and VEGF-C treatment groups. In the functional assay, the VEGF-C group dramatically recovered compared with the control group. The capillary/myofiber ratio in the thigh muscle and number of LYVE-1 positive cells was higher in the CD11b+ and VEGF-C groups than in controls. Furthermore, expression of VEGF-A, VEGF-C, and VEGF receptor messenger ribonucleic acid and protein was observed in CD11b+ cells.ConclusionsThe VEGF-C derived from CD11b+ cells play a critical role in angiogenesis and lymphangiogenesis in a murine model of hind limb ischemia. Consequently, treatment with self-CD11b+ cells accelerated recovery from ischemia and may be a promising therapeutic strategy for peripheral arterial disease patients.

Clinical RelevanceThe incidence of peripheral arterial disease is rapidly increasing commensurate with a rapid rise in the elderly population. Conventional therapy has a 5-year prognosis and survival rates <30%; thus, an alternative approach, termed therapeutic angiogenesis, has been developed, consisting of local injection of bone marrow cells (BMCs). However, the precise therapeutic mechanism, including paracrine effects, is unclear. We focused on CD11b+ cells, which promote regulation of vascular endothelial growth factor-A and especially vascular endothelial growth factor-C. CD11b+ treatment could reduce the total number of cell injections and improve the early operative points of blood flow compared with conventional therapy with BMCs. Consequently, CD11b+ treatment accelerates recovery from ischemia and is helpful in reducing the need for repeated aspiration of BMCs.