机械力敏感离子通道Piezo1参与高血压诱导心房纤维化的机制研究

Involvement of mechanically activated Piezo1 channels in atrial fibrosis induced by elevated hydrostatic pressure

目的:利用小鼠心房成纤维细胞探讨机械敏感离子通道Piezo1在调控高血压所致心房纤维化,进而导致房颤中的作用及可能机制。方法:采用酶消化法分离培养6~8周龄雄性C57BL/6小鼠的原代心房成纤维细胞,并采用课题组自制的加压装置(专利号201420109263.1)建立高血压模型,Western blot比较不同静水压(0、20和40 mmHg)干预下细胞Piezo1、Src/p-Src及纤维化指标Ⅰ/Ⅲ型胶原蛋白α1链(Col1A1/3A1)和基质金属蛋白酶2/9(MMP-2/9)蛋白表达水平。高静水压(40 mmHg)干预下的心房成纤维细胞,分别给予不同浓度(1、3和10μmol/L)的Piezo1抑制剂GsMTx4,或转染Piezo1 siRNA质粒降低Piezo1的表达,以及不同浓度(5和10μmol/L)的Src抑制剂PP1,观察细胞中Src/p-Src和纤维化相关因子蛋白水平的变化。常压下给予不同浓度(1、3和10μmol/L)的Piezo1特异性激动剂Yoda1,观察心房成纤维细胞中Src/p-Src和纤维化相关因子蛋白水平的变化。结果:随着压力升高,小鼠心房成纤维细胞中Piezo1、Src/p-Src及纤维化指标Col1A1/3A1和MMP-2/9蛋白表达水平显著升高(P<0.05);GsMTx4/Piezo1 siRNA或PP1处理后,可使高静水压导致的Src/p-Src及纤维化相关因子水平下降(P<0.05)。而细胞给予Piezo1特异性激动剂Yoda1可模拟出高静水压干预的结果(P<0.05)。结论:在小鼠心房成纤维细胞中,机械敏感通道蛋白Piezo1可能通过调控Src/p-Src参与高血压所致的心房纤维化。

AIM:To investigate the role and possible mechanism of mechanosensitive ion channel Piezo1 in hypertension-induced atrial fibrosis and even atrial fibrillation.METHODS:Primary atrial fibroblasts from 6-to-8-weekold male C57BL/6 mice were isolated and cultured by enzyme digestion,and the cell hypertension model was established using self-made pressure device.Western blot analysis was conducted to compare the expression levels of Piezo1,Src/pSrc,and fibrosis indicators collagen type Ⅰ/Ⅲα1 chain(Col1A1/3A1)and matrix metalloproteinase-2/9(MMP-2/9)in the cells under different pressure interventions(0,20 and 40 mmHg).At 40 mmHg,the cells were given different concentrations(1,3 and 10μmol/L)of ion channel inhibitor GsMTx4 or Piezo1 siRNA,or Src inhibitor PP1(5 and 10μmol/L),and the changes of Src/p-src and fibrosis-related factor protein levels were observed.The changes of Src/p-Src and fibrosis-related factor protein levels were also observed when the cells were given different concentrations of Piezo1-specific agonist Yoda1(1,3 and 10μmol/L).RESULTS:Higher hydrostatic pressure(20 and 40 mmHg)increased the expression of Piezo1 and Src in mouse atrial fibroblasts(P<0.05),accompanied by increases in Src/p-Src,and fibrosis indicators Col1A1/3A1 and MMP-2/9.GsMTx4,Piezo1 siRNA or PP1 significantly reversed the above changes(P<0.05).In addition,Piezo1 agonist Yoda1 simulated the changes of electrical remodeling and related signal molecules in atrial myocytes induced by high hydrostatic pressure(P<0.05).CONCLUSION:Mechanosensitive ion channel Piezo1/Src kinase signaling pathway is activated during hypertension,leading to the decreases in Col1A1/3A1 and MMP-2/9 and the electrical remodeling of atrial myocytes.