Chromogranin A的关键降解肽段抗动脉粥样硬化机制研究

Exploring and pursuing endogenous proteins and peptides with vascular protective effects is of translational significance. In our study, we have demonstrated that chromogranin A-derived peptide vasostatin-2 and catestatin possess vascular protective effects. The protein levels of vaostatin-2 and catestatin are significantly decreased in severe atherosclerotic coronary artery samples as compared with non-significant atherosclerotic samples. Such alteration of these two peptide levels is also noted in patients with coronary artery disease. Intraperitoneal injection of vasostatin-2 recombinant protein and synthetic catestatin peptide inhibits atherogenesis in apoE-KO mice fed high-fat chow. These two proteins exhibit anti-inflammatory effects in endothelial cells and macrophages. Moreover, microarray analysis of vasostatin-2 and catestatin-induced endothelial cells and macrophages show and is verified that vasostatin-2 produces anti-inflammatory effects through inhibiting RhoGTPase and its downstream elements, and that catestatin creates biological effects via upregulation of ACE2-ang(1-7)-Mas pathway in renin-angiotensin system. Next, we will further investigate and clarify the mechanisms of vasostatin-2- and catestatin- created anti-inflammatory effects and anti-atherosclerosis using cells and genetically modified mice. Interaction site of vasostatin-2 and catestatin in cell will be studied and the mechanism of vasostatin-2 and catestatin alteration in vivo will be explored.

研究内源性血管保护性蛋白和生物活性肽具转化意义。我们发现Chromogranin A降解肽段Vasostatin-2和Catestatin具血管保护效应。与对照相比，严重粥样硬化冠脉组织中Vasostatin-2和Catestatin 水平降低，冠心病患者的血清也有一致的变化。Vasostatin-2和Catestatin重组蛋白腹腔注射均能抑制apoE-KO小鼠主动脉粥样硬化，并在内皮和巨噬细胞中有抗炎症作用。用这两肽段处理内皮和巨噬细胞后进行芯片分析发现并证实：Vasostatin-2经抑制RhoGTPase及其下游通路发挥作用，Catestatin经上调肾素-血管紧张素系统中ACE2-Ang(1-7)-Mas途径拮抗炎症。后续将用分子、细胞和基因修饰小鼠，阐明Vasostatin-2和Catestatin抗炎症抗动脉粥样硬化作用及机制，揭示它们作用的关键位点和它们量异常的病因和机制。

脂肪因子Chromogranin A（CgA）在体内被降解形成约10个肽段，包括vasostatin-2。本研究主要探讨vasostatin-2的功能。.1..血清vasostatin-2水平降低与冠心病发生及冠脉病变严重程度相关。我们发现，严重冠心病患者冠脉剥脱内膜组织、外周血单个核细胞和血清vasostatin-2水平较对照动脉组织、细胞和血清中的CgA显著增高，vasostatin-2显著下降（均P<0.05），与冠脉病变支数及病变严重程度相关。.2..体外和体内实验中，vasostatin-2具有抑制炎症作用和抗动脉粥样硬化功能。我们发现，vasostatin-2能剂量依赖性抑制内皮细胞黏附分子表达，抑制体外巨噬细胞-内皮细胞粘附。体内实验中，与PBS对照相比，高脂喂养的ApoE-KO小鼠腹腔内注射vasostatin-2能显著抑制动脉粥样硬化的发生。vasostatin -2抑制白细胞对肠系膜血管壁的粘附，以及主动脉的黏附分子和炎症因子表达。在机制研究中，vasostatin-2能够通过抑制单核巨噬细胞的迁移，能干预荧光标记的炎性细胞Ly-6Chi对血管壁的滞留和向泡沫细胞的转变，使MOMA-2阳性区域显著降低， vasostatin-2通过抑制Rac1-Pak1途经机制而产生对炎性细胞迁移的干预。.3..血清vasostatin-2水平降低与心梗患者3年随访的MACE事件相关，对大鼠心梗后心肌损伤有抑制作用。我们的研究发现，心梗后心肌组织中vasostatin-2的表达下降，与梗死面积负相关。血清vasostatin-2水平与心梗后3年的MACE事件相关。在心梗大鼠腹腔中注射vasostatin-2能显著改善心功能，抑制心肌组织重构和纤维化，这些作用与vasostatin-2抑制了Rho、Wnt和TLR-4通路，并调控了RAS系统有关。.4..血清CgA水平升高和vasostatin-2水平降低与冠脉支架术后再狭窄有关，对小鼠股动脉内膜损伤后内膜增生有抑制作用 。我们发现，冠脉支架术后再狭窄患者CgA水平显著升高（P<0.05），而vasostatin-2水平明显降低（P<0.05）。体内实验中，vasostatin-2能显著抑制股动脉内膜损伤后内膜增生，而CgA则促进内膜增殖。这一效应主要来源于其对病变血管段组织ACE2的上调。