骨形态发生蛋白2-MAPK/Smads信号通路调控房颤心房纤维化的实验研究

Atrial fibrosis is one of the most important factors in the development of atrial fibrillation, but its mechanism is not yet fully understood. The assumptions based on the issues raised in the preliminary studies：In atrial fibrillation,the increased expression of bone morphogenetic protein -2 (BMP-2) in atrial could promot the atrium fibroblasts differentiation and collagen secretion through MAPK/Smads signaling pathway. While administration of exogenous BMP-2-siRNA could be able to control the development of atrial fibrosis and reduce occurrence and persistence of atrial fibrillation. To test the hypothesis, the following items will be observed: ① the expression of the related proteins, such as BMP-2,MAPK and Smads in rapid-pacing induced atrial fibrillation canies;②the effect of BMP-2-MAPK/Smads signaling pathway on the atrium fibroblasts in vitro; whether exogenous BMP-2 inhibitors (noggin) could inhibit fibroblast activation and collagen secretion induced by a variety of stimuli (AngⅡ, LPS, and H2O2)in vitro; ③the exogenous BMP-2-siRNA could inhibit the effect of BMP-2-MAPK/Smads signaling pathway ,furthermore，lessen the atrium fibrosis and reduce the inducibility and persistence of atrial fibrillation .The study will help to elucidate the mechanism of fibrosis in atrial fibrillation.It will provide a new direction and targets for atrial fibrillation prevention.

心房纤维化是促进房颤发生发展的重要因素之一，但发生机制仍不完全清楚。本课题组在前期研究基础上提出假设：房颤时心房骨形态发生蛋白2（BMP-2）表达增加，调控MAPK/Smads信号通路，促进心房成纤维细胞分化及胶原分泌增加，是心房纤维化的主要机制；给予外源性BMP-2-siRNA能够抑制心房纤维化，减少房颤发生和持续。为验证该假设，本研究将观察：①房颤犬心房肌BMP-2表达是否上调，进而激活MAPK和Smads表达；②细胞水平BMP-2－MAPK/Smads信号通路在心房成纤维细胞激活中的作用，并观察外源BMP-2抑制剂（noggin）能否通过下调MAPK、Smads的mRNA和蛋白表达,抑制多种刺激因素（AngⅡ、LPS、和H2O2）诱导的成纤维细胞激活，抑制胶原分泌；③在体抑制BMP-2表达能否调控MAPK/Smads信号通路，进而抑制房颤心房纤维化。为房颤防治提供新的方向及靶点。

心房颤动（AF）是临床最常见的心律失常疾病，其病因复杂且发病机制仍不完全清楚。心房间质纤维化是心房结构重构的重要特征，主要表现为间质内胶原大量沉积，从而引起心房传导不均一性增加，最终引起房颤发生。骨形态发生蛋白（BMPs）是转化生长因子超家族的一类具有促进细胞生长和分化的细胞因子，可参与胚胎发育、组织发生和修复等生理过程，但BMP2是否参与AF的发生发展及其分子机制尚无报道。. 本课题历时三年，建立慢性房颤犬模型及应用AngⅡ刺激原代大鼠心房成纤维细胞模型，通过在体及离体实验，从细胞、动物水平探讨了BMP2-MAPK-Smad1/5/8信号通路调控心房成纤维细胞增殖及活化的作用机制。现本项目已按预期计划顺利完成了实验内容的研究，得到如下结论：一、在体实验：1.慢性房颤犬模型房颤发生率显著增加、房颤持续时间明显延长；2.心房组织炎症及纤维化程度显著增加；3.房颤犬心房组织中BMP2、P-ERK、Smad1/5/8、α-SMA及І、Ⅲ型胶原蛋白含量显著增加；4.给予AngⅡ受体拮抗剂可减轻BMP2诱导的心房纤维化程度及房颤的发生；二、离体实验：1.外源性给予BMP2使原代心房成纤维细胞增殖并活化，而BMP2抑制剂可抑制上述改变；2. 外源性给予BMP2可诱导心房成纤维细胞P-ERK、Smad1/5/8、α-SMA及І、Ⅲ型胶原蛋白合成增加；3. BMP2 siRNA则能够抑制AngⅡ引起的心房成纤维细胞增殖及活化，并通过调控BMP2-MAPK-Smad1/5/8信号通路减少胶原的合成。本项目的顺利完成将为AF纤维化发病机制的理解和治疗提供新的思路和途径，并为提供AF的临床疗效提供新的理论基础和基础研究依据。