부산대학교 도서관

Introduction: Extracellular vesicles (EVs) and their MicroRNA (miR) content are involved in several physiological processes, including the response to ischemia and angiogenesis. Hypercholesterolemia (HC) is an important cardiovascular risk factor associated with impaired angiogenesis and ischemia-induced neovascularization. However, the effect of HC on the modulation of miR content in EVs in the setting of tissue ischemia is unknown.Hypothesis: We tested the hypothesis that HC causes alterations in the miR content of EVs related to ischemia/neovascularization, and that engineered EVs can be harnessed as angiogenic vectors to promote angiogenesis and therapeutic neovascularization in this situation.Methods: We used a mouse model of peripheral artery disease to induce severe ischemia in the hindlimbs of hypercholesterolemic ApoE and control C57Bl/6J mice. Microvesicles (MVs) and exosomes (exo) were isolated from ischemic skeletal muscles by differential centrifugation. Total RNA was extracted from these 2 types of EVs, and the miR content was analyzed using next generation sequencing. Selected miRs were transfected in EVs and the angiogenic capacity of the engineered EVs was assessed in vitro using a Matrigel assay.Results:: Bioinformatic analysis of the 100 most expressed miRs in EVs showed an enrichment of several miRs in EVs compared to ischemic skeletal muscles. Interestingly the highly expressed miR-21a-5p, which has previously been shown to have angiogenic properties, is specifically enriched in exo. Moreover, we found that miR-21a-5p expression is reduced in exo isolated from ischemic skeletal muscles in hypercholesterolemic conditions. We next forced miR-21a-5p expression in exo, and found that these engineered exo significantly increase tube formation in HUVECs.Conclusions: We show that several angiogenic miRs are enriched in EVs in the setting of tissue ischemia, and that HC can alter EV miR profile in this situation. miR-21a-5p, which expression is reduced by HC, was identified as an important modulator of exo angiogenic activity. Exo engineered to overexpress miR-21a-5p could eventually constitute a novel therapeutic strategy to promote neovascularization and reduce ischemic damages in severe vascular diseases.