Exosomes from miR-primed endothelial progenitor cells for treating ischemic stroke

项目概要 缺血性中风（IS），即由于脑血管闭塞导致的脑血流丧失，仍然是最常见的疾病之一。 世界上最严重的健康问题。目前治疗中风的方法包括溶栓剂、PTAS、 神经保护药物已被证明是有帮助的；然而，它们也受到治疗时间狭窄的限制 窗口，再中风的风险和低效率。寻找一种理想的治疗策略来保护大脑免受 急性损伤并促进血管生成和神经发生对于神经功能恢复来说势在必行 需要。内皮祖细胞（EPC）具有分化为成熟内皮细胞（EC）的能力， 已知参与血管生成和神经发生。 EPC移植显示出可喜的结果 用于 IS 治疗。新的证据表明干细胞释放的 EX 可用于治疗 IS。 EX 是 内体来源的小膜囊泡通过转移 miRNA (miR) 介导细胞间通讯。 干细胞 EX 比干细胞具有优势，因为它缺乏自我增殖能力，并且更容易储存和使用 比细胞传递。在这个项目中，我们的目标是使用 EPC 释放的 EX（EPC-EX）作为 IS 治疗的新途径 因为它们选择性地携带富集的 miR-126。我们发现 EPC-EX 可以保护 EC 免受 缺氧/复氧（H/R）主要通过其携带的miR-126引起的损伤和功能障碍；输血 EPC-EXs 可以在 IS 后保留脑血流量 (CBF) 并减少梗塞体积。有趣的是，反 已发现中风患者中的细胞凋亡/抗氧化应激 miR-210 降低。我们发现 miR-210 引发的 EPC-EX (EPC-EXsmiR210) 在保护 EC 免受 H/R- 影响方面比 EPC-EX 更有效 诱发的伤害。总而言之，通过结合 EPC（干细胞）、EX（新的细胞因子）的有益作用， 脑微环境）和 miR（多信号通路靶标），我们相信 EPC-EXsmiR210 可以 作为 IS 的一种新的治疗途径。设计了三个目标来检验我们的假设：EPC-EXsmiR210 通过保护脑细胞免受急性缺血性损伤并促进 通过 EPC-EX、miR-126 和 miR-210 促进神经功能恢复的血管生成/神经生成 下游通路：VEGF/VEGFR2/PI3K/Akt、PIK3R2/PI3K/Caspase 和 BDNF/TrkB/PI3K/Akt。目标 1： 确定 EPC-EXsmiR210 是否具有增强保护 EC 和神经元免受 H/R 诱导的作用 通过针对多种途径来治疗损伤/功能障碍。目标 2：确定 EPC-EXsmiR210 是否发挥增强作用 通过针对多种途径保护大脑免受 IS 引起的急性损伤。目标 3：确定 EPC-EXsmiR210是否在促进神经血管生成和功能恢复方面发挥增强作用 IS 通过针对多种途径。 。

Project Summary Ischemic stroke (IS), loss of cerebral blood flow due to occlusion of cerebral blood vessels, remains one of the most severe health problems in the world. Current treatments for stroke, such as thrombolytic agents, PTAS, and neuroprotective drugs have shown to be helpful; however, they are also limited by a narrow therapeutic time window, risk of re-stroke and low effectives. Looking for an ideal therapeutic strategy that protect brain from acute injury and promote angiogenesis and neurogenesis for neurological functional recovery, is thus imperative needed. Endothelial progenitor cells (EPCs) with the ability to differentiate into mature endothelial cells (ECs), are known to participate in angiogenesis and neurogenesis. EPC transplantation has shown promising results for IS therapy. Emerging evidence suggests that stem cell released EXs could be used for treating IS. EXs are endosomal origin small-membrane vesicles mediating cell-cell communication by transferring miRNAs (miRs). Stem cell EXs hold advantages over stem cells because of lacking self-proliferation and are easier to store and deliver than cells. In this project, we aim to use EPC-released EXs (EPC-EXs) as a novel avenue for IS therapy as they selectively carry enriched miR-126. We have found that EPC-EXs protect ECs from hypoxia/reoxygenation (H/R)-induced injury and dysfunction mainly through their carried miR-126; transfusion of EPC-EXs could preserve cerebral blood flow (CBF) and decrease infarct volume after IS. Of interest, the anti- apoptosis/anti-oxidative stress miR-210 has been found to be decreased in the stroke patients. We discovered that miR-210 primed EPC-EXs (EPC-EXsmiR210) are more effective than EPC-EXs in protecting ECs from H/R- induced injury. Taken together, by combining the beneficial effects of EPCs (stem cells), EXs (novel player of brain microenvironment), and miRs (multiple signaling pathway targets), we believe that EPC-EXsmiR210 could serve as a novel therapeutic avenue for IS. Three aims are designed to test our hypothesis that EPC-EXsmiR210 have enhanced therapeutic effects on IS by protecting brain cells from acute ischemic injury and promoting angiogenesis/neurogenesis for neurological functional recovery through EPC-EXs, miR-126 and miR-210 downstream pathways, VEGF/VEGFR2/PI3K/Akt, PIK3R2/PI3K/Caspase and BDNF/TrkB/PI3K/Akt. Aim 1: Determine whether EPC-EXsmiR210 have enhanced effects on protecting ECs and neurons from H/R-induced injury/dysfunction by targeting multiple pathways. Aim 2: Determine whether EPC-EXsmiR210 exert enhanced effects on protecting brain from IS-induced acute damage by targeting multiple pathways. Aim 3: Determine whether EPC-EXsmiR210 exert enhanced effects on promoting neurovascular genesis and functional recovery after IS by targeting multiple pathways. .