MSCs修复糖尿病血管内皮细胞线粒体损伤的作用及机制

Diabetic artery disease is one of the important diabetes complications, characterized by poor therapeutic outcome and high mortality. Development of new therapeutic strategies for diabetic artery disease is essential for the clinical treatment. In diabetic condition, glucolipotoxicity, caused by high levels of glucose and lipids in circulation, profoundly increases the accumulation of superoxide anion and lipid peroxide in endothelial cells. The excessive ROS is capable to impair mitochondrial biogenesis, reduce NO production and damage the NO homeostasis, leading to endothelial dysfunction and subsequent various cardiovascular diseases. Mesenchymal Stem Cells (MSCs) are multipotent non-hematopoietic stem cells that have the capacity for self-renewal and terminal differentiation into a multitude of different cell types. Due to their paracrine, transdifferentiation, and immunosuppresive effects, MSCs hold great promise as therapy for cardiovascular disease, such as myocardium regeneration and ischemic vascular disease. Recent studies discovered that intercellular communication using tunnneling nanotubes (TNT) can transfer mitochondria between MSCs and target cells, which is considered as a new machinary for the therapeutic function of MSCs. Nevertheless, the precise mechanism involved in the rescue of endothelia cells by MSCs is still to be elucidate. . Our previous study showed that high glucose level induced endothelial damage, and MSCs protected the endothelial cells from it by rescuing mitochondrial function and cell viability, and reducing ROS production. Additionally, we found that the machinery was relevant to the SIRT1 signaling pathway. Based on these results, we will establish human umbilical vein endothelial cell (HUVEC) model and rat diabetic nephropathy model to in depth investigate the regulation of energetic metabolism of diabetic endothelium by MSCs; discover the role of mitochondrial transferring in MSCs mediated endothelial rescue; and elucidate the molecular mechanism in regulation of mitochondrial function by MSCs focusing on SIRT1 signaling pathway. Our project will shed light to the clinical treatment of MSCs on diabetic vascular disease.

糖尿病血管病是糖尿病主要并发症之一，其新型治疗手段的研究是目前临床研究的热点。糖脂毒性引起的血管内皮细胞ROS堆积，线粒体损伤，NO生物利用度降低是导致该细胞失功，引起心血管病变的重要原因。大量实验已证明间充质干细胞（MSCs）对心肌梗死、缺血性血管病等心血管疾病有良好治疗作用。近年来研究发现MSCs细胞通过在细胞间形成隧道纳米管结构，介导线粒体向损伤靶细胞转移是MSCs修复靶细胞损伤的新机制。但其对血管内皮细胞的修复机制还有待深入研究。本课题将通过建立人脐静脉内皮细胞模型和糖尿病肾病大鼠模型，系统研究MSCs调节糖尿病血管内皮细胞能量代谢紊乱的分子机制；通过分子影像定位技术，揭示线粒体转移在MSCs修复糖尿病内皮细胞损伤中的机制；以前期SIRT1分子相关信号通路研究发现为线索，深入研究并阐明MSCs调节靶细胞线粒体功能的分子机制。为MSCs在糖尿病血管病中的应用提供科学依据。

本项目旨在研究间充质干细胞（MSCs）对糖尿病内皮细胞损伤的干预作用及机制。主要回答了以下问题：.1. MSCs移植的体内效果.建立大鼠糖尿病或肥胖模型，通过体内移植大鼠骨髓MSCs，证明了MSCs能改善高血脂，缓解大鼠血管炎症反应，降低血管内皮细胞糖脂毒性损伤，保护血管功能的作用，缓解糖尿病并发症进程。.2. MSCs对内皮细胞糖脂毒性保护作用的分子机制.1）MSCs条件培养基（MSCs-CM）干预能够抑制高糖诱导的内皮细胞线粒体ROS生成，提高细胞抗氧化能力,维持线粒体的稳态，改善线粒体生物功能。MSCs能激活PI3K/AKT信号通路，促进Sirt1蛋白的表达和胞浆的富集，再通过激活Sirt1/AMPK/PGC-1α信号通路，提高内皮细胞线粒体生物合成，恢复线粒体功能，缓解糖尿病血管病变的进程。.2）MSCs干预能通过Pink1/Parkin介导的线粒体自噬，促进受损线粒体的自噬清除，缓解高糖刺激引起的线粒体形态及功能损伤，降低线粒体ROS的累积，从而抑制线粒体依赖的内皮凋亡，改善内皮失功。.3）MSCs可以调节饱和脂肪酸棕榈酸引起的内皮细胞脂代谢紊乱，并可通过抑制内质网应激（ER stress）及内皮间质转分化（EndMT）途径减轻脂毒性损伤，恢复内皮细胞活性及功能。糖脂联合毒性反应可以加重内皮细胞的损伤，特别表现在炎性因子表达升高和NF-B活化。MSCs通过分泌STG-6因子可以缓解糖脂毒性引起的炎性反应和内皮损伤。.4）MSCs可以调节TNF-α诱导的血管内皮细胞慢性炎症反应，降低炎性因子的表达和NF-B的活化。MSCs分泌的STC-1能通过上调p-AMPK促进UCP-2的表达，进而降低细胞内ROS积累，其从而达到缓解内皮炎性损伤。.3. MSCs对内皮细胞炎性微环境的调节作用。.糖尿病状态下，巨噬细胞易于向M1促炎表型分化。MSCs能调节高糖诱导下巨噬细胞的极性分化，促进抗炎型M2巨噬细胞分化，减轻局部炎症反应并且MSCs可以通过转运线粒体到高糖诱导巨噬细胞中，改善巨噬细胞线粒体功能，调节炎性反应。