丹参酮ⅡA通过NLRP3/Caspase-1信号通路对心肌成纤维细胞的保护作用

郑鹏 王俊帅 占大钱 王敏 夏海发 刘旭东 明晓青 周代星 冯俊

摘要 目的：探討丹参酮ⅡA对心肌成纤维细胞损伤的保护作用及可能机制。方法：分离和培养大鼠心肌成纤维细胞（Cardiac Fibroblast，CFs）。以脂多糖（LPS）刺激CFs建立脓毒症心肌损伤体外模型，以不同浓度丹参酮ⅡA预处理CFs 30 min后，给予LPS刺激，设对照组，LPS模型组，LPS+不同浓度TSA（2 μmol/L、10 μmol/L、50 μmol/L）组;采用蛋白质免疫印迹试验法（Western blotting）检测CFs的NLRP3和Caspase-1蛋白表达水平，酶联免疫吸附试验（ELISA）检测细胞上清中IL-1β和IL-18的含量。结果：LPS刺激24 h后，CFs的NLRP3蛋白的表达水平最高。LPS模型组NLRP3和Caspase-1蛋白的表达较对照组明显升高（P<0.01）。与LPS模型组比较，丹参酮ⅡA（10 μmol/L、50 μmol/L）处理组NLRP3和Caspase-1蛋白的表达均明显降低（P<0.05）;丹参酮ⅡA处理组的IL-1β和IL-18水平均明显低于LPS模型组（P<0.05）。结论：丹参酮ⅡA能抑制LPS诱导的心肌成纤维细胞内NLRP3/Caspase-1炎症反应信号通路分子的表达，这可能是其保护心肌细胞的分子机制之一。

关键词 丹参酮ⅡA;脓毒症;心肌成纤维细胞;NLRP3/Caspase-1信号通路

Protective Effect of Tanshinone ⅡA on Cardiac Fibroblasts through the NLRP3/Caspase-1 Signaling Pathway

ZHENG Peng1，WANG Junshuai1，ZHAN Daqian1，WANG Min1，XIA Haifa2，LIU Xudong1，MING Xiaoqing1，ZHOU Daixing1，FENG Jun1

（1 Department of Emergency，Tongji Hospital，Tongji Medical College，Huazhong University of Science and Technology，Wuhan 430000，China; 2 Department of Anesthesia，Wuhan Union Hospital，Tongji Medical College，Huazhong University of Science and Technology，Wuhan 430000，China）

Abstract Objective：To explore the protecting role and mechanism of tanshinone ⅡA in lipopolysaccharide （LPS）-induced cardiac fibroblast injure.Methods：Rat cardiac fibroblasts （Cardiac Fibroblast，CFs） were Isolated and cultured.Lipopolysaccharide （LPS） was used to stimulate CFs to establish an in vitro model of septic myocardial injury.After CFs were pretreated with different concentrations of Tanshinone ⅡA for 30 min，they were given LPS stimulation.A control group，an LPS model group and LPS+different TSA （2 μmol/L，10 μmol/L，50 μmol/L） were set up.Western blotting was used to detect the NLRP3 and Caspase-1 protein expression levels of CFs，and the enzyme-linked immunosorbent assay （ELISA） was used to detect the content of 1β and IL-18.Results：After 24 h of LPS stimulation，the expression level of NLRP3 protein in CFs was the highest.The expression of NLRP3 and Caspase-1 protein in the LPS model group was significantly higher than that in the control group （P<0.01）.Compared with the LPS model group，the expressions of NLRP3 and Caspase-1 protein in the Tanshinone ⅡA （10 μmol/L，50 μmol/L） treatment group were significantly reduced （P<0.05）; the level of IL-1β and IL-18 in the Tanshinone ⅡA treatment group was significantly lower than that of the LPS model group （P<0.05）.Conclusion：Tanshinone ⅡA may be a potent agent to protect against myocardial dysfunction in sepsis by regulating NLRP3/Caspase-1 signaling pathway.

Keywords Tanshinone ⅡA; Sepsis; Cardiac fibroblast; NLRP3/Caspase-1 signaling pathway

中图分类号：R285.5文献标识码：Adoi：10.3969/j.issn.1673-7202.2021.06.011

脓毒症是宿主对感染的反应异常，产生危及生命的器官功能损害，大约50%的脓毒症患者存在心脏功能损害[1]。脓毒症引起心肌损害的机制尚不明确。目前普遍认为脓毒症患者体内过度的炎症反应可能参与了心脏功能损害的过程[2-3]。在心脏的细胞组成中，心肌细胞占比低于50%，而心脏成纤维细胞（CFs）占比则高达60%。研究显示心肌成纤维细胞发挥着感受损伤和炎症反应的前哨作用[4]。

Nod受体蛋白3（NLRP3）是一类重要的胞质内受体蛋白，它对多种物理和化学刺激做出反应[5-6]，与Caspase-1形成的炎性小体可导致炎症介质IL-1β和IL-18的成熟[7]。NLRP3炎性小体及其下游炎症介质IL-1、IL-18在冠状动脉疾病或心肌梗死患者中的表达升高[8]。NLRP3炎性小体主要表达在心肌成纤维细胞上，心肌细胞上则很少[9]。心肌成纤维细胞上NLRP3炎性小体的过度激活参与了心肌缺血再灌注损伤过程。……