Abstract 19049: MiR-126 Containing Exosomes From Human CD34+ Stem Cells Activate Endothelial Cells to Initiate Angiogenesis and Ischemic Tissue Repair

Abstract

Introduction: Locally transplanted human CD34+ stem cells have been shown to improve exercise tolerance in patients with myocardial ischemia and promote angiogenesis in animal models. Recently we have demonstrated that CD34+ cells secrete membrane bound nano-vesicles called exosomes as a major component of their pro-angiogenic paracrine secretion. We hypothesize that cell-free exosomes from CD34+ cells (CD34 Exo) mimic the beneficial effects of the cells and promote angiogenesis and ischemic tissue repair via transfer of pro-angiogenic microRNAs, possibly to endothelial cells. Methods and Results: Therapeutic potential of CD34 Exo isolated from equal number of adult human peripheral blood-derived CD34+ cells was evaluated in an immunodeficient murine model of hind limb ischemia. Similar to cells, treatment with CD34 Exo resulted in significant improvement in ischemia compared to treatment with PBS (perfusion ratio: 1.01±0.04 v 0.57±0.1; capillary density: 1.8±0.3 v 0.9±0.1/HPF; limb amputation: 16% v 100%; p<0.05; n=10-21). MicroRNA expression profiling and confirmatory tests indicated that CD34 Exo are significantly enriched with pro-angiogenic miRNAs such as miR126. Treatment with CD34 Exo in mouse hind limb ischemic tissue increased mature miR126 expression, but not endogenous mouse pri-miR126, and activated pro-angiogenic miR126 target-gene expression. Treatment of CD34 Exo lacking miR126 abolished this increase and diminished the pro-angiogenic function of CD34 Exo-treated endothelial cells. This data suggests that miR126 is important for CD34 Exo function. Interestingly, confocal imaging and flow cytometry analyses revealed that CD34 Exo were selectively internalized by endothelial cells when injected to the ischemic tissue. Live imaging using fluorescent miR126 confirmed that CD34 Exo directly transfers miR126 and possibly other yet to be identified moieties from its repertoire specifically to endothelial cells in the ischemic tissue. Conclusion: CD34+ exosomes transfer miR126 to endothelial cells to induce angiogenesis and ischemic tissue repair. The functional benefits associated with CD34+ cell therapy may be mediated by exosomes-mediated targeted transfer of angiogenic microRNAs to endothelial cells.