CD137-CD137L信号调控自噬转归介导内膜微钙化形成的机制研究

Atherosclerotic microcalcification is calcium deposition (< 60μm in diameter) which is located in the intimal and embedded in the plaque. It is an important cause of acute cardio- and cerebro-vascular events as it easily cause plaque rupture. Our previous studies showed that the spot mircrocalcification formation is closely related to autophagy in the atherosclerotic plaque of ApoE-/- mice induced by CD137-CD137L signal. In vitro, activation of CD137-CD137L signal can mediate autophagosome exocytosis and promote the osteogenic phenotype transformation of smooth muscle cells and extracellular matrix calcium deposition. Therefore, we hypothesize that CD137-CD137L signal mediated the formation of atherosclerotic microcalcification by modulating authophagic outcome. In this study, we focus on how CD137-CD137L signal initiates the formation of microcalcification through modulating autophagy. ApoE-/- mice and VSMCs were used as models to verify the effect of CD137-CD137L signal on autophagy and microcalcification. We aim to investigate how CD137-CD137L signal modulates autophagy, and reveal the mechanism of CD137-CD137L in the formation of microcalcification through modulating autophagic outcome. The study is expected to bring new opportunities for prevention and treatment of atherosclerotic intimal microcalcifuation.

动脉粥样硬化（AS）微钙化是位于内膜镶嵌于斑块内<60μm的钙沉积灶，极易引起斑块破裂造成急性心脑血管事件的发生。CD137-CD137L信号是参与AS进展的关键分子。我们前期研究显示：CD137-CD137L信号诱发ApoE-/-小鼠斑块内点灶状微钙化的形成与自噬密切相关；体外激动CD137-CD137L能介导自噬小体出胞并促进平滑肌细胞外基质钙沉积及成骨表型转变。因此我们提出如下假说：CD137-CD137L信号通过调控自噬转归介导AS内膜微钙化形成。本研究在前期基础上以CD137-CD137L信号调控自噬启动微钙化形成为主线，在ApoE-/-小鼠和平滑肌细胞模型上，分别激动和抑制CD137-CD137L信号，观察该信号对自噬转归及微钙化形成的影响，明确CD137-CD137L信号如何调控自噬转归，阐明该信号调控自噬转归介导内膜微钙化形成机制。该研究有望为AS内膜微钙化防治带来新契机。

本项目在前期基础上，探讨CD137L-CD137信号是否通过调控自噬介导AS微钙化形成及其机制。.研究发现：在ApoE-/-小鼠及体外培养VSMC中均证实激动CD137-CD137L信号能够促进AS斑块钙化的形成，其机制是由于激动该信号激活了阻断了自噬小体与溶酶体的结合，而未被结合的自噬小体出胞，促进斑块内钙化的形成。而CD137-CD137信号阻断自噬小体与溶酶体结合是由于激动该信号抑制了Rab7的活性及表达，使得Rab7与溶酶体上FYCO1蛋白结合受阻，导致自噬过程受阻，导致了AS斑块的形成。同时，我们发现CD137-CD137信号可通过JNK和p38通路分别调控小鼠血管平滑肌自噬及钙化。激动CD137-CD137L信号还可通过Rab7影响小鼠平滑肌细胞外泌体分泌，并且分泌的外泌体可能通过传递NFATc1信号介导小鼠血管平滑肌细胞钙化。.利用3%巯基乙酸盐肉汤诱导法高效提取小鼠腹腔巨噬细胞发现CD137-CD137L还可调控巨噬细胞MMP-9表达影响VSMC钙化形成；而且，CD137-CD137L可调控巨噬细胞M1/M2极性转变影响血管新生；此外，CD137-CD137L通过内皮细胞TET2蛋白表达及转运至血管平滑肌细胞促进新生内膜形成。 .临床研究：运用ELISA分析患者血清发现急性心肌梗死患者抑凋亡蛋白明显减少，促炎因子明显升高；动物和细胞实验表明CD137-CD137L通过抑制抗氧化Nrf2的核转位促氧化和通过激活NF-kB促炎导致ECs凋亡影响AS斑块稳定性。.本研究阐明了CD137-CD137L信号调控自噬转归介导内膜微钙化形成机制，发表SCI论文22篇，中华8篇。培养研究生15名。获得江苏省医学技术新引进一等奖。