循环HMGB1/DNA复合物通过脾免疫细胞RAGE/TLR9受体加剧心肌再灌注损伤的机制研究

循环中HMGB1与DNA以复合物形式存在参与机体免疫调控。申请人发表论文证实HMGB1(心脏 )-RAGE(脾)轴是介导心肌再灌注免疫损伤的核心通路，可显著上调脾免疫细胞IL-1β(NLRP3炎症复合体主要产物)水平。我们预实验提示，缺血心肌释放HMGB1/DNA复合物，激活脾脏介导的免疫炎症反应。文献报道，HMGB1可以通过RAGE受体促进DNA进入免疫细胞作用于胞浆中TLR9受体，上调ROS水平，激活NLRP3炎症复合体。据此，本项目拟在小鼠心肌缺血再灌注模型中，利用RAGE-/-，TLR9KO及NLRP3-/-等基因修饰小鼠，结合骨髓移植/脾细胞重建技术，证实缺血心肌释放HMGB1/DNA复合物，作用于脾免疫细胞RAGE受体促使DNA进入胞内，通过TLR9-ROS通路活化NLRP3炎症复合体，促进全身炎症反应，加剧再灌注心肌损伤。深入探索心肌再灌注免疫损伤机制，寻求新的心肌保护靶点。

Circulating HMGB1/DNA complex plays an important role in immune and inflammatory responses. We have already demonstrated that a HMGB1 (Heart) – RAGE (Spleen) axis is activated following myocardial ischemia and exacerbates infarct size during reperfusion. This activated heart-spleen axis exacerbates inflammatory response in part by enhancing IL-1beta production, a key component of NLRP3 inflammasome. In our preliminary studies, we found that ischemic myocardium released HMGB1/DNA complex during reperfusion, which activated splenic leukocytes. Recently, it is reported that HMGB1 promotes cellular uptake of DNA by activating RAGE. Increased cytosol DNA subsequently binds TLR9 and triggers downstream inflammatory response. One of the downstream pathway following TLR9 activation includes ROS – inflammasome. By using RAGE-/-, TLR9-/- and NLRP3-/- mice as well as techniques of bone marrow transplantation and immune cell adoptive transfer, the current project is to test our hypothesis that ischemic myocardium releases HMGB1/DNA complex into the blood stream, which in turn activates splenic leukocytes via RAGE/TLR9-ROS-NLRP3 inflammasome pathway and ultimately exacerbates infarct size during reperfusion.