基质细胞衍生因子-1及其受体在A2aR基因敲除小鼠低氧性肺动脉高压形成中的作用研究

The relationship between adenosine and pulmonary hypertension is always highly concerned. It has been reported that A2aR could reduce pulmonary hypertension and proliferation of PASMCs induced by hypoxia. Our previous study shows that in chronic hypoxic conditions, proliferation of pulmonary artery smooth muscle cells and collagen deposition in A2aR knockout mice were more significantly than those in wild mice. However, the mechanism is unclear. It is reported that activation of A2aR could inhibit inflammative reaction. As we know, inflammation is closely related to pulmonary hypertension. Stromal cell derived factor -1 (SDF-1) plays an important role in the inflammatory response (CXCR4 is its receptor). Another report is that activating A2aR could inhibit the functions of CXCR. Based on the findings above, this project intends to study the role of SDF-1/CXCR4 axis in pulmonary hypertension induced by hypoxia in A2aR knockout mice through in vitro and in vivo study. It has been reported that baicalin might inhibit SDF-1/CXCR4, prevent pulmonary hypertension induced by endotoxin shock, and affect the function of A2aR. Therefore, the project intends to make sure the effects of baicalin on hypoxic pulmonary hypertension and its mechanism, expecting to find a drug with good efficacy and little side effect to treat pulmonary hypertension.

腺苷与肺动脉高压的关系一直受到高度关注。有报道腺苷通过A2a受体抑制肺动脉高压形成。本课题组近期发现A2aR基因敲除小鼠出现肺动脉平滑肌、成纤维细胞显著增殖，胶原沉积；低氧下，更为明显。但机制尚不清楚。据报道，激活A2a受体可抑制炎症表达，而炎症与肺动脉高压形成密切相关，有研究认为在炎症反应中起重要作用的基质细胞衍生因子-1（SDF-1，受体为CXCR4）与肺动脉高压形成有关。另有报道激活A2aR可抑制CXCR功能。基于上述发现，本项目拟在整体、细胞和分子水平系统研究SDF-1/CXCR4轴在A2aR基因敲除小鼠肺动脉高压形成中的作用，以探明其作用机制。另有报道，黄芩苷可抑制SDF-1/CXCR4；能预防内毒素休克所致肺动脉高压；可能影响A2aR功能。因此，本项目拟系统研究黄芩苷对低氧性肺动脉高压、SDF-1/CXCR4、A2aR的影响，以期找到一种疗效好而副作用小的治疗肺动脉高压药物。

本课题按照标书研究设计方案，完成了预定研究内容，圆满完成课题计划任务。本项目建立了野生及A2aR基因敲除低氧性肺动脉高压小鼠模型，应用Powerlab生理信号记录系统、HE染色、免疫组化、免疫荧光、激光共聚焦显微镜、western blot等技术，在整体水平研究了A2aR基因在低氧肺动脉高压中的作用，并在分子水平系统研究了SDF-1/CXCR4轴在A2aR基因敲除小鼠肺动脉高压形成中的作用以及黄芩苷的影响。离体研究方面，构建了A2aR基因敲减大鼠肺动脉平滑肌细胞株，应用激光共聚焦显微镜、免疫组化、免疫荧光、western blot、CCK-8、transwell等技术深入研究了SDF-1/CXCR4轴在A2aR基因敲除小鼠肺动脉高压形成中的机制以及黄芩苷的作用通路。.项目组首次研究发现在A2aR基因敲除小鼠中，SDF-1/CXCR4通路功能增强可能是缺氧性肺动脉高压形成的重要原因，揭示了A2aR下调、SDF-1/CXCR4通路上调在缺氧性肺动脉高压形成中的作用,进一步丰富了缺氧性肺动脉高压的发病机制理论。同时，在低氧诱导的肺动脉高压模型中，发现黄芩苷可减轻缺氧性肺动脉高压，逆转肺血管结构重建，黄芩苷促进A2aR表达进而抑制SDF-1/CXCR4通路可能是其重要作用机制。离体实验研究发现，腺苷受体A2a能够抑制细胞增殖和迁移，并抑制SDF-1/CXCR4轴介导的细胞趋化作用；黄芩苷能上调腺苷受体A2a，从而抑制低氧诱导的平滑肌细胞的过度增殖和迁移。上述结果表明，A2aR下调、SDF-1/CXCR4通路上调与肺动脉高压、肺血管重塑的形成有关，黄芩苷通过上调腺苷受体A2a、阻断SDF-1/CXCR4轴而发挥抑制肺动脉高压作用，有望成为一个防治肺动脉高压肺血管重塑的新选择。在本课题的赞助下，在国内外共发表与本项目相关的学术论文9篇，其中4篇被SCI收录，授权专利1项。完成所有计划任务。.综上所述，该项研究达到了预期的研究目的，研究成果对于进一步阐明低氧性肺动脉高压的形成机制具有重要的意义，同时也为黄芩苷在低氧性肺动脉高压患者上的临床推广应用提供了更为确切的实验研究依据。