Department of Internal Medicine, Research Center for Experimental Medicine (CeRMS) and Center for Molecular Biotechnology, and University of Turin, Turin, Italy.
Int J Immunopathol Pharmacol. 2012 Jan-Mar;25(1):75-85. doi: 10.1177/039463201202500110.
Paracrine mediators released from endothelial progenitor cells (EPCs) have been implicated in neoangiogenesis following ischemia. Recently, we demonstrated that microvesicles (MVs) derived from EPCs are able to activate an angiogenic program in quiescent endothelial cells by a horizontal transfer of RNA. In this study we aim to investigate whether EPC-derived MVs are able to induce neoangiogenesis and to enhance recovery in a murine model of hindlimb ischemia. Hindlimb ischemia was induced in severe combined immunodeficient (SCID) mice by ligation and resection of the left femoral artery and mice were treated with EPC-derived MVs (MVs), RNase-inactivated MVs (RnaseMVs), fibroblast-derived MVs or vehicle alone as control (CTL). Since MVs contained the angiogenic miR-126 and miR-296, we evaluated whether microRNAs may account for the angiogenic activities by treating mice with MVs obtained from DICER-knock-down EPC (DICER-MVs). The limb perfusion evaluated by laserdoppler analysis demonstrated that MVs significantly enhanced perfusion in respect to CTL (0.50±0.08 vs 0.39±0.03, p<0.05). After 7 days, immunohistochemical analyses on the gastrocnemius muscle of the ischemic hindlimb showed that MVs but not fibroblast-MVs significantly increased the capillary density in respect to CTL (MVs vs CTL: 24.7±10.3 vs 13.5±6, p<0.0001) and (fibroblast-MVs vs CTL: 10.2±3.4 vs 13.5±6, ns); RNaseMVs and DICER-MVs significantly reduced the effect of MVs (RNaseMVs vs CTL: 15.7±4.1 vs 13.5±6, ns) (MVs vs DICER-MVs 24.7±10.3 vs 18.1±5.8, p <0.05), suggesting a role of RNAs shuttled by MVs. Morphometric analysis confirmed that MVs enhanced limb perfusion and reduced injury. The results of the present study indicate that treatment with EPC-derived MVs improves neovascularization and favors regeneration in severe hindlimb ischemia induced in SCID mice. This suggests a possible use of EPCs-derived MVs for treatment of peripheral arterial disease.
内皮祖细胞 (EPC) 释放的旁分泌介质已被牵涉到缺血后的新血管生成。最近，我们证明 EPC 衍生的微泡 (MVs) 能够通过 RNA 的水平转移激活静止内皮细胞的血管生成程序。在这项研究中，我们旨在研究 EPC 衍生的 MVs 是否能够诱导新血管生成并增强严重免疫缺陷 (SCID) 小鼠后肢缺血模型中的恢复。通过结扎和切除左股动脉诱导 SCID 小鼠后肢缺血，并将 EPC 衍生的 MVs (MVs)、RNA 酶失活的 MVs (RnaseMVs)、成纤维细胞衍生的 MVs 或单独的载体作为对照 (CTL) 用于治疗。由于 MVs 包含了促血管生成的 miR-126 和 miR-296，我们通过用 DICER 敲低的 EPC (DICER-MVs) 获得的 MVs 处理小鼠，评估微 RNA 是否可能是促血管生成活性的原因。通过激光多普勒分析评估的肢体灌注显示，MVs 与 CTL 相比显著增强了灌注 (0.50±0.08 对 0.39±0.03，p<0.05)。7 天后，对缺血后肢的比目鱼肌进行免疫组织化学分析表明，MVs 但不是成纤维细胞-MVs 与 CTL 相比显著增加了毛细血管密度 (MVs 与 CTL：24.7±10.3 对 13.5±6，p<0.0001) 和 (成纤维细胞-MVs 与 CTL：10.2±3.4 对 13.5±6，ns)；RNaseMVs 和 DICER-MVs 显著降低了 MVs 的作用 (RNaseMVs 与 CTL：15.7±4.1 对 13.5±6，ns) (MVs 与 DICER-MVs 24.7±10.3 对 18.1±5.8，p<0.05)，表明 MVs 转运的 RNA 起作用。形态计量学分析证实 MVs 增强了肢体灌注并减少了损伤。本研究结果表明，用 EPC 衍生的 MVs 治疗可改善严重免疫缺陷小鼠后肢缺血引起的新血管生成和促进再生。这表明 EPC 衍生的 MVs 可能用于治疗外周动脉疾病。
