p300/p53/Smad3通路参与人心房成纤维细胞衰老相关心房纤维化

p300/p53/Smad3 pathway involved in aging-related atrial fibrosis in hu⁃man atrial fibroblasts

目的:探究p300在人心房成纤维细胞(HAFs)衰老相关心房纤维化中的作用及其可能机制。方法:采用组织贴块法从人左心耳组织中分离HAFs并进行传代培养建立细胞衰老模型,检测衰老相关β-半乳糖苷酶(SA-β-Gal)活性以比较P3和P7代细胞衰老程度的不同。Western blot实验比较不同代数(P3、P7和P11)细胞的p300、p53和Smad3等衰老和纤维化相关因子的蛋白表达水平。高代数(P7)细胞给予不同浓度(6、9和12μmol/L)的p300抑制剂姜黄素处理或转染p300 shRNA质粒敲减p300的表达,低代数(P3)细胞转染p300过表达质粒,分别观察p53和Smad3等衰老和纤维化相关因子蛋白水平的变化。结果:随着HAFs传代,衰老细胞增加,伴随p300、p53、p21、p16、p-Smad3、基质金属蛋白酶2/9(MMP-2/9)和纤溶酶原激活物抑制因子1(PAI-1)水平的升高(P<0.05)。姜黄素处理或转染p300 shRNA质粒后,高代数(P7)细胞的p53和Smad3等衰老和纤维化相关因子的蛋白水平减少(P<0.05);而过表达p300后,低代数(P3)细胞的p53和Smad3的蛋白水平明显增加(P<0.05)。结论:p300可以促进HAFs衰老相关纤维化,其机制可能是通过激活p53/Smad3通路实现的。

AIM:To investigate the role of p300 in aging-related atrial fibrosis in human atrial fibroblasts(HAFs)and its potential mechanism.METHODS:HAFs were obtained from human left atrial tissue,and the senescence model was established by cell passage.Senescence-associatedβ-galactosidase(SA-β-Gal)staining was used to detect the cell senescence,and Western blot was used to determine the protein levels of p300,p53,Smad3 and other senescence and fibrosis associated proteins in HAFs.RESULTS:Compared to passage 3 HAFs,the proportion of senescent cells,and the protein levels of p300,p53,p-Smad3 and other senescence and fibrosis associated proteins were increased in HAFs at passage 7 and 11(P<0.05).After treated with curcumin(a p300 inhibitor)or transfection with p300 smallhairpin(sh)RNA plasmid,the protein levels of p300,and the senescence and fibrosis associated proteins were decreased in HAFs at passage 7(P<0.05).Up-regulation of p300 by transfection with p300 over-expression plasmid increased the protein levels of p53,Smad3 and MMP-2 in HAFs at passage 3(P<0.05).CONCLUSION:p300/p53/Smad3 signaling pathway plays an important role in aging-related atrial fibrosis in HAFs.