TLR4/NF-κB-miR 27b反馈环路在心肌梗死后不良心室重塑中的作用及机制研究

Mounting evidence suggests that persistent activation of toll-like receptor 4 (TLR4)/nuclear factor-kappa B (NF-κB) signaling pathway which is triggered by the endogenous ligands contributed to the myocardial inflammation, adverse ventricular remodeling and the pathogenesis of heart failure. The mechanism which is responsible for the persistant activation of TLR4/NF-κB signaling pathway remains unclear. It has been recently demonstrated that microRNA (miRNA) play an essential role in the regulation of signaling pathways. Our pre-experimental study revealed that the activation of TLR4/NF-κB was accompanied by up-regualtion of miR-27b in the infarcted myocardium and LPS-stimulated cardiomyocytes. Bioinformatic analyses indicated that there is binding site for NF-κB in the promoter region of miR-27b and peroxisome proliferator-activated receptor gamma(PPAR-γ) which inhibited the TLR4/NF-κB pathway is the predicted target of miR-27b. Therefore, we hypothesized that a TLR4/NF-κB-miR-27b positive feedback loop is involved in the ventricular remodeling after myocardial infarction. To test the hypothesis, we plan to study the dynamics of TLR4/NF-κB/miR-27b/PPAR-γ after myocardial infarction and the effects of up-regulating or down-regulating miR-27b on TLR4/NF-κB/PPAR-γ and ventricular remodeling.Our study will clarify the mechanism for activation and regulation of TLR4/NF-κB signaling pathway in the ventricular remodeling and provide new target for intervention.

研究表明，TLR4/NF-κB信号通路在心肌梗死（MI）后心室重塑中呈现持续活化状态，参与心室重塑的发生发展，然其持续活化的机制尚未明确。miRNA在信号通路调控中扮演重要角色。我们的前期研究提示心室重塑时TLR4/NF-κB通路活化伴miR-27b表达上调。生物信息学分析提示，miR-27b的启动子区域具备NF-κB的结合位点，TLR4/NF-κB通路的负性调控因子PPAR-γ则很可能是miR-27b的作用靶点。因此，我们推测miR-27b是TLR4/NF-κB通路的"开关"，能启动正反馈效应导致其持续活化参与不良心室重塑。本课题拟动态观察MI后TLR4/NF-κB/miR-27b/PPAR-γ的变化及上调/下调miR-27b对TLR4/NF-κB /PPAR-γ和心室重塑的影响，以期验证以上假说。结果有助于阐明心室重塑中TLR4/NF-κB的激活及调控机制，为靶向干预提供新靶点。

急性心肌梗死严重危害人类健康。由于冠心病治疗学的长足进展，心肌梗死后的急性期存活率明显改善，但梗死后慢性心力衰竭的发病率和死亡率却逐年上升。心室重塑是慢性心力衰竭的病理生理基础。miRNA参与心脏各种病理生理过程中,但其在心肌梗死后心室重塑中的作用仍未完全明确。本研究发现，建立小鼠心肌梗死模型后，在缺血缺氧的刺激下，心脏局部的TLR4/NF-κB信号通路活化，同时上调miR-27b的表达。离体细胞实验发现，LPS能够激活心肌细胞的TLR4/NF-κB信号通路miR-27b的表达上调，NF-κB抑制剂PDTC能够完全逆转LPS刺激所导致的miR-27b的上调，NF-κB能够调节miR-27b的启动子活性。给予模型动物miR-27b的模拟物miR-27b mimics能够加剧心室重塑，促进心功能恶化，心室扩张，加重局部炎症，促进心肌细胞凋亡。与此相反，给予模型动物miR-27b的抑制物anti-miR-27b能够减轻心室重塑，改善心功能，减轻心室扩张和局部炎症，抑制心肌细胞凋亡。本研究提示了TLR4/NF-κB-miR-27b通路在心肌梗死后心室重塑发挥重要作用，一方面丰富了心肌梗死后心室重塑的机制研究内容，另一方面为其干预提供新的干预靶点。此外，我们还扩展了研究，发现慢性心力衰竭患者CD4+T细胞中miR-451表达下调，CD25，CD69和MHC-II活化分子上调，miR-451可能通过myc调节CD25，CD69和MHC-II活化分子的表达。