组蛋白去甲基化酶KDM5B在心肌梗死后炎症和心肌纤维化中的调控作用及其表观机制研究

The inflammatory responses and myocardial fibrosis after myocardial infarction (MI) play critical roles in the later cardiac remodeling, but the uncontrolled inflammation and fibrosis will lead to the generation of adverse cardiac remodeling and heart failure. So, the full understand of the precise molecular regulation mechanism underlying post-MI inflammation and fibrosis is of great significance in the clinical treatment of myocardial infarction and its complication heart failure. During the development of cardiovascular diseases, the levels of histone methylation on the promoters of many genes are alerted to regulate and control gene expression, thus the histone methyltransferase and demethylase are involved in the pathological process. However, the roles of histone methyltransferase and demethylase in post-MI inflammation and fibrosis remain unclear. In the preliminary experiments of this case, we found that the expression of histone H3K4 demethylase KDM5B was markedly increased in the infarct and border zone of myocardium in mice 3-7 days after MI. Furthermore, the left ventricular function of KDM5B knockout mice was better than that in wild type mice after MI, while the expression of alpha-SMA and myocardial fibrosis was significantly decreased in the infarct and border zone of KDM5B deficient heart, which suggest that KDM5B affects the post-MI myocardial fibrosis. Since the inflammation strength is involved in the generation of post-MI myocardial fibrosis, thus in this case, we aim to further investigate the regulatory roles of KDM5B in the inflammation, myocardial fibrosis and cardiac remodeling after MI, and also explore the target genes and the underlying molecular mechanisms through epigenetic approaches, which will uncover the mechanisms behind adverse cardiac remodeling and shed new light on the treatment of post-MI heart failure.

心肌梗死后炎症反应和心肌纤维化的失调会导致不良心脏重构及心力衰竭。组蛋白甲基修饰酶通过控制基因的组蛋白甲基化状态以改变基因的表达水平，参与心血管疾病发生发展，然而组蛋白甲基修饰酶在心梗后心脏重构中的作用尚不清楚。我们预实验研究发现组蛋白H3K4去甲基化酶KDM5B的表达在心梗后3-7天显著上升，提示其可能参与心梗后炎症和修复过程的调控。KDM5B基因敲除小鼠心梗后的心功能较野生型小鼠明显改善，其梗死/边缘区心肌组织中a-SMA表达和胶原纤维沉积较野生型小鼠显著降低，表明KDM5B影响心梗后心肌纤维化。鉴于心梗后心肌纤维化的发生与炎症反应强度密切相关，在本项目中，我们将深入研究KDM5B在心梗后炎症损伤、心肌纤维化及心脏重构中的调控作用，并利用表观组学的方法寻找其作用的靶基因，探索分子调控机制，为心梗后的不良心脏重构及心衰的发生机制提供新的表观遗传学解说，为心梗后心衰的干预发现潜在的新靶点。

本项目研究了组蛋白去甲基化酶KDM5B对心肌梗死后心肌纤维化及心功能的调节作用及机制。研究发现KDM5B在梗死心肌组织及增殖分化的肌成纤维细胞中的表达显著升高；利用KDM5B基因敲除小鼠构建心肌梗死模型，发现KDM5B缺陷可显著降低小鼠心肌梗死后的心肌纤维化，改善心功能，抑制不良心脏重构；在体外原代心肌成纤维细胞中，TGF-β诱导的纤维化水平在KDM5B敲除小鼠心脏成纤维细胞中显著降低，利用KDM5B siRNA干扰野生型原代心肌成纤维细胞KDM5B的表达也得到类似结果；机制研究发现KDM5B可以直接结合纤维化抑制因子ATF3的启动子区域，并能下调ATF3的转录表达，从而增强TGF-beta/Smad依赖的经典以及非经典纤维化通路的活化，促进成纤维细胞中纤维化分子的表达。我们的研究结果表明组蛋白去甲基化酶KDM5B是心肌梗死后心肌纤维化的正向促进因子，可能参与了临床心梗后期不良心脏重构及心衰的发生，为心肌纤维化的控制和治疗药物开发提供了潜在的方向。另外，在本项目资助下，我们同期研究发现一系列相关成果，包括研究发现组蛋白去甲基化酶KDM2B通过招募染色质重塑复合物的核心催化亚基BRG1而介导IL-6启动子区染色质开放，选择性促进IL-6的转录表达，加重组织炎症损伤，诠释了KDM2B在炎症反应中发挥非甲基化酶活性依赖的新的调控功能（Cell Mol Immunol.2019.doi: 10.1038/s41423-019-0251-z，通讯作者）；研究发现磷酸酶PPM1L结合并抑制IKKb的磷酸化和活化，从而降低心梗后心肌组织炎症，防止不良心脏重构发生（The Journal of Immunology. 2019;203:1338-1347，通讯作者）；研究发现miR-21通过靶向KBTBD7及其下游的p38和NF-kB通路，防止心梗后过度炎症反应导致的不良心脏重构及心功能障碍（Cell Death and Disease.2018;9:769，通讯作者）；研究发现肺缺血再灌注后肺泡巨噬细胞发生的自噬能促进肺组织炎性细胞因子的产生，加重肺缺血后炎症损伤（Cell Death and Differentiation.2017;24:683-693，通讯作者）等。综上，本项目发表标注本项目资助的SCI论文6篇，申请国家发明专利2项，已顺利完成所有研究内容和目标。