活性氧对门静脉高压症内脏高动力循环影响机制的研究

Splanchnic vasodilation and the development of portosystemic collaterals are the two main factors leading to splanchnic hyperdynamic circulation. The increasement of reactive oxygen species (ROS) has been found in cirrhosis and portal hypertension(PHT), however it is unknown that whether it plays an important role in the regulation of splanchnic hyperdynamic circulation, especially the splanchnic vasodilation. This research exploring the mechanism of ROS affecting splanchnic hyperdynamic circulation in the cirrhosis and portal hypertension rat model. Then it studies the mechanism how ROS regulates G protein-coupled receptors desensitization through β-arrestin-2 by adopting the technology of western blot, co-immunoprecipitation, immunofluorescence histochemistry, so as to explore the influence of G protein-coupled receptors desensitization, regulated by ROS, on vasoconstriction signaling pathway. Simutaneously, this research works over how high level of ROS activates ERK's and JAK2/STAT3's signaling pathway, adjusts and controls vascular endothelial growth factor(VEGF) from different levels of transcriptional and post-transcriptional, and promotes portosystemic collaterals angiogenesis. Further, this study will search for the possibility and mechanism of interdicting ROS and treating PHT hyperdynamic circulation through clinical trial. This study will systematically explore the mechanism of ROS playing in PHT splanchnic hyperdynamic circulation for the first time and contribute new though for the treatment of PHT esophageal and gastric variceal bleeding.

内脏动脉扩张和门体侧支循环的建立是肝硬化门静脉高压症（PHT）内脏高动力循环的两大原因。研究发现肝硬化PHT活性氧增高，其在调控内脏高动力循环尤其内脏动脉扩张中的作用和机制，目前尚不清楚。本课题采用肝硬化PHT大鼠模型研究活性氧对PHT内脏高动力循环的影响，应用免疫印迹、免疫共沉淀和免疫荧光定位等技术探索活性氧通过β-arrestin-2调控G蛋白偶联受体脱敏的机制，进而探讨活性氧调控后的受体脱敏对血管收缩性信号通路的影响；同时探索β-arrestin-2脱敏条件下高ROS水平活化ERK和JAK2/STAT3信号通路，从转录及转录后水平不同层面调控血管内皮生长因子的高表达，促进门体侧支血管增生的新机制，并通过临床研究，进一步探求阻断ROS治疗PHT高动力循环的可行性及其机制。本课题首次系统地研究活性氧在PHT内脏高动力循环中的作用机制，为临床治疗PHT食管胃底静脉曲张破裂出血提供新思路。

内脏动脉扩张和门体侧支循环的建立是肝硬化门静脉高压症（PHT）内脏高动力循环的两大原因。研究发现肝硬化PHT活性氧增高，其在调控内脏高动力循环尤其内脏动脉扩张中的作用和机制，目前尚不清楚。本课题采用肝硬化PHT大鼠模型研究活性氧对PHT内脏高动力循环的影响，应用体外微血管、免疫印迹和免疫共沉淀等技术探索活性氧对血管收缩性信号通路的影响；同时探索高ROS水平活调控血管内皮生长因子的高表达，促进门体侧支血管增生的新机制，并通过临床研究，进一步探求阻断ROS治疗PHT高动力循环的可行性及其机制。我们的研究结果如下：（1）应用PEG-catalase降低肠系膜组织H2O2水平能抑制门体侧支血管的形成， 此外我们也发现，PHT大鼠肠系膜组织VEGF和VEGFR-2蛋白水平显著高于假手术组。PHT大鼠经PEG-catalase处理后，肠系膜组织VEGF及VEGFR-2蛋白表达显著下调。这说明改善PEG-catalase内脏高动力循环状态，与VEGF介导的内脏血管增生受抑制有关。（2）CCl4致肝硬化门静脉高压症大鼠肝外环境sEH表达增高，与活性氧含量相关。使用NAPDH特异性抑制剂去除活性氧后，sEH表达减低，血管肌源性反应恢复，提示在肝硬化门静脉高压症中限制sEH作用可作为缓解氧化应激外的另一治疗方向。（3）研究探讨了GTP对门静脉高压大鼠肠系膜小动脉低反应性的保护作用，并揭示了GTP显着改善肠系膜小动脉低反应的机制。 GTP通过抑制Akt / eNOS信号通路激活来调节氧化应激和NO， 因此，GTPs治疗增强肠系膜小动脉的收缩性并降低门静脉压力。 这些新的发现支持GTP具有预防和治疗PHT血管并发症的巨大潜力。总之，通过本项目，我们基本可以确定抗氧化剂降低ROS，有助于缓解门静脉高压症内脏高动力循环，为将来治疗门静脉高压症提供新的治疗方案。