心肌细胞外泌体相关linc-ROR在通心络"逆向强化"保护微血管内皮减轻心肌缺血再灌注损伤中的核心作用和机制

Myocardial ischemia/reperfusion injury (MIRI) has been the main obstacle to myocardial protection in the times of myocardial reperfusion therapy for acute myocardial infarction (AMI). Based on the series of previous studies from our lab, we consequently further hypothesized that, in addition to protecting the cardiomyocytes directly, Tongxinluo is capable of conferring further protection on endothelial cells (ECs) indirectly by promoting cardiomyocytes to secret exosomes carrying linc-ROR during ischemia/reperfusion and linc-ROR/miR-145/p70s6k1/eNOS pathway is supposed to be the underlying mechanism. To validate such a scientific hypothesis, in-vivo study will be performed in I/R model of SD rats, treated with Tongxinluo, the inhibitor preventing the release of exosomes GW4869, eNOS inhibitor L-NNA, MEK inhibitor PD98059, exosomes from cardiomyocytes after hypoxia/reoxygenation, exosomes from Tongxinluo-pretreated cardiomyocytes after hypoxia/reoxygenation, miR-145 inhibitor miR-145 antagomir, in-vivo conditional knockout of linc-ROR in cardiomyocytes and in-vivo conditional knockout of p70s6k1 in ECs. Pathological, immunohistological, and molecular biological methods will be used to unravel the role and mechanism of linc-ROR/miR-145/p70s6k1/eNOS pathway in protecting ECs further, and thereby mitigating MIRI. The outcomes of this study will offer scientific evidence for the central regulation mechanism and signaling pathway in the mitigation of MIRI injury by Tongxinluo. Additionally, it will be helpful for us to unravel the mechanism of MIRI, and develop and apply cardioprotective drugs in the clinical setting after revascularization of AMI. As our study is both original and innovative, it is of great scientific and clinical value.

在系列研究的基础上，我们进一步假设：“在‘心肌灌注单元’中，通心络在缺血再灌注时除能直接保护心肌细胞外，还可能通过心肌细胞外泌体中的linc-ROR这一核心因子调控miR-145/p70s6k1/eNOS通路‘强化’保护内皮细胞（‘民拥军’），是通心络‘逆向强化’保护心肌微血管内皮的结构和功能，从而减轻心肌再灌注损伤的核心机制。”拟以SD大鼠建立心肌I/R模型，以通心络、外泌体释放抑制剂GW4869、心肌细胞linc-ROR或内皮细胞p70s6k1条件性敲除等干预措施，以分子生物学和病理学等国际公认的方法，验证上述假设。研究结果有助于揭示通心络保护心肌缺血再灌注损伤的核心调控机制和信号通路；阐明心肌缺血再灌注损伤的机制，并为其防治提供全新策略。

急性心肌梗死（AMI）是危害中国人民健康的第一杀手。作为治疗AMI的首选方法，早期冠脉再灌注疗法虽然可使堵塞冠状动脉迅速再通，恢复心肌供血供氧，改善患者预后；但再灌注同时也可引起缺血再灌注损伤，导致患者预后不佳。另外，中国幅员辽阔以及地区医疗水平参差不齐，由于错过早期冠脉开通最佳时间而导致心肌大面积不可逆性损伤也使得患者预后严重恶化。针对上述问题，本课题组主要研究中药通心络减轻急性心肌梗死缺血再灌注损伤核心作用机制和主要信号通路。本项目发现，通心络在缺血再灌注时可通过心肌细胞外泌体中的linc-ROR这一核心因子调控miR-145/p70s6k1/eNOS通路‘强化’保护内皮细胞，是TXL保护心肌微血管内皮的结构和功能，从而减轻心肌再灌注损伤的核心机制。此外，发现TXL预处理MSCs（间充质干细胞）提高外泌体治疗急性心梗的疗效，机制为TXL预处理可提高MSCs来源外泌体中miR-146a-5p的表达水平，且通过miR-146a-5p/IRAK1/NF-κB p65通路保护心肌细胞。还比较了外泌体（Exo）和不同方式预处理的MSC联合移植治疗AMI的疗效，发现Exo联合静脉移植“低氧+TXL”预处理MSC能够显著增强MSC在心肌局部的归巢和存活，显著提高心梗后心功能、缩小梗死面积。相关成果目前共发表3篇SCI，获批2项发明专利，荣获一个国家科学技术进步一等奖。