IκB激酶α调控巨噬细胞极化在心肌缺血再灌注损伤中的作用及机制研究

With the development of percutaneous coronary intervention (PCI) and coronary artery bypass grafting (CABG) therapeutic technology, reperfusion therapy is effective for reducing overall mortality after myocardial infarction, but restoration of the blood flow through the previously ischemic myocardium can yield additional myocardial ischemic-reperfusion (I/R) injury, which limits the beneficial effects of reperfusion and may hinder from the treatment of cardiovascular disease. The mechanism of immune mediated myocardial I/R injury is taken seriously, recent studies have demonstrated that macrophage plays a pivotal role in I/R injury, and IKKα highly expresses in macrophages, which could regulate macrophages to involve in inflammatory and immune response, however, to date, there is no report about the role of IKKα regulating macrophages in myocardial I/R injury. Hereby, our preliminary experimental results demonstrated that IKKα has a protective effect in myocardial I/R injury, and recruited macrophages were involved in phenotypic transformation also namely polarization, so we hypothesize that IKKα could play a vital role in myocardial I/R injury through regulating macrophage polarization. The present study will adopt macrophage-specific knockout of IKKα and transgenic mice with macrophage-specific overexpression of IKKα to study the role of IKKα and its molecular mechanism regulating macrophages in myocardial I/R injury, and using primary cultures of macrophages, further investigate the effect of IKKα on macrophage polarization. This project will reveal the new molecular mechanisms in the onset and development of myocardial I/R injury, and provide a new therapeutic target for myocardial I/R injury.

随着经皮冠脉介入及冠脉搭桥术等技术的发展, 再灌注治疗成功降低心梗死亡率，但在改善心肌供血同时也会引起损伤，成为阻碍心血管病治疗的重要难题。目前心肌缺血再灌注（I/R）损伤免疫机制正受到重视，最新研究发现巨噬细胞在I/R损伤中扮演重要角色，IκB激酶α（IKKα）高表达于巨噬细胞，其能够调控巨噬细胞参与机体炎症及免疫等过程，但IKKα调控巨噬细胞在心肌I/R损伤中机制尚不清楚。我们预实验结果显示IKKα对心肌I/R损伤具有保护作用，且募集的巨噬细胞发生表型转变即极化，据此我们推测IKKα可能通过调控巨噬细胞极化在心肌I/R损伤发挥重要作用。本项目采用巨噬细胞特异性IKKα敲除和转基因小鼠研究IKKα调控巨噬细胞在心肌I/R损伤中的影响及机理，并利用原代培养巨噬细胞，细胞水平阐明IKKα对巨噬细胞极化的影响及机制。本项目将揭示心肌I/R损伤的新机制，为临床干预心肌I/R损伤提供新的治疗靶点。

背景：IKK复合物在肿瘤及免疫系统中扮演了重要的角色，这一功能主要归因于其在NF-κB通路中主导的调节作用。值得注意的是，IKKα作为IKK复合物的一员，已被报道在多种炎症疾病中发挥重要的多样作用，而在心肌缺血再灌注损伤后巨噬细胞中IKKα的作用和机制尚未研究。目的：探究IKKɑ在心肌缺血再灌注损伤后巨噬细胞中的作用和机制。方法：将巨噬细胞IKKɑ敲除小鼠（mIKKɑ-/-）和野生型小鼠（WT）分为假手术组（Sham）和心肌缺血再灌注损伤组 （IR），并将IR组小鼠结扎左前降支缺血30分钟，之后分别再灌注3天和7天，并且用慢病毒感染RAW264.7干扰或过表达IKKɑ，再用脂多糖（LPS）刺激。结果：mIKKɑ-/-小鼠心脏的心室重塑更为缓慢，炎症反应更为剧烈且巨噬细胞的聚集要明显多于WT小鼠，此外mIKKɑ-/-小鼠在IR后其巨噬细胞更易向M1型极化。缺少IKKɑ的RAW264.7在LPS刺激后比正常RAW264.7表达更多的促炎因子。相反，过表达IKKɑ则会抑制促炎因子。机制研究发现在LPS刺激下IKKɑ能与RelB结合并调节MEK1/2-ERK1/2及下游的p65通路。结论：IKKɑ作为一个新的调节因子，通过负向调节巨噬细胞向M1型极化来保护心肌缺血再灌注损伤后炎症的发展。因此，IKKɑ可以作为治疗心肌缺血再灌注损伤的有效靶点，在缺血性心脏病中发挥重要作用。