高迁移率族蛋白1通过调控自噬因子P62调节心肌细胞纤维化
HMGB1 induces myocardial fibrosis by regulating autophagy factor P62
聂雅琴 1乌日娜 2金慧 2张广平 2李永明 2智利 2薛仕煊 2娜日格乐2
作者信息
- 1. 014040 包头医学院中心临床医学院
- 2. 014040 包头市中心医院心血管内科
- 折叠
摘要
目的:探究高迁移率族蛋白 1(HMGB1)通过调控自噬活性对心肌纤维化的影响及作用机制.方法:将人心肌细胞AC16 培养于DMEM培养基中,用不同浓度的HMGB1(0、4、20、100 μg/L)干预AC16 细胞 6 h,免疫荧光染色检测心脏成纤维细胞活化的标志性蛋白α平滑肌肌动蛋白(α-SMA)的表达水平,Western blot法检测心肌自噬相关蛋白哺乳动物雷帕霉素靶蛋白(mTOR)、磷酸化mTOR(p-mTOR)、微管相关蛋白 1轻链 3(LC3)、磷酸肌醇-3-激酶 3(PIK3C3)、可溶性及不可溶性P62、α-SMA、Ⅰ型胶原蛋白(CollagenⅠ)在AC16 中的表达水平.免疫共沉淀检测α-SMA与P62 之间的互相作用.结果:不同浓度的HMGB1 刺激AC16 细胞,AC16 细胞中心肌纤维化标志蛋白CollagenⅠ与α-SMA的表达水平呈浓度依赖性升高,自噬蛋白PIK3C3、不可溶性P62 的蛋白表达水平,p-mTOR与mTOR的比值,LC3-Ⅱ与LC3-Ⅰ的比值呈浓度依赖性升高,可溶性P62 的蛋白表达水平呈浓度依赖性降低.各组间的差异均有统计学意义(P均＜0.01).免疫共沉淀实验显示P62 与α-SMA存在相互作用.结论:外源性HMGB1 可抑制AC16细胞中的自噬流,诱导AC16细胞纤维化.
Abstract
Objective:To explore the effect of high mobility group box protein 1(HMGB1)on myocardial fibrosis by regulating autophagy activity of human cardiomyocytes(AC16)and its possible mechanism.Methods:AC16 cells were cultured in DMEM medium and treated with different concentrations of HMGB1(0、4、20、100 μg/L)for 6 h.The expression of α-smooth muscle actin(α-SMA),a marker protein of cardiac fibroblast activation,was detected by immunofluorescence staining.Western blot was used to determine the expression levels of myocardial autophagy-related proteins mammalian target of rapamycin(mTOR),phosphorylated mTOR(p-mTOR),light chain 3(LC3),phosphoinositide-3-kinase 3(PIK3C3),soluble and insoluble p62,α-SMA,and collagenⅠ(CollagenⅠ)in AC16.The interaction between α-SMA and p62 was assessed by co-immunoprecipitation.Results:The expression levels of myocardial fibrosis marker protein(collagenⅠand α-SMA),autophagy protein PIK3C3,insoluble p62,P-mTOR/mTOR ratio,LC3-Ⅱ/LC3-Ⅰratio were increased in a dose-dependent manner.In contrast,the expression level of soluble p62 was decreased in a dose-dependent manner.The differences between groups were statistically significant(all P＜0.01).There was an interaction between p62 and α-SMA.Conclusion:Exogenous HMGB1 inhibits autophagic flux in AC16 cells and induces myocardial fibrosis.
关键词
高迁移率族蛋白1/AC16细胞/自噬/心肌纤维化Key words
High mobility group protein/AC16 cells/Autophagy/Myocardial fibrosis引用本文复制引用
基金项目
内蒙古医科大学科技百万工程联合项目(YKD2020KJBWLH065)
出版年
2024
NETL
NSTL
万方数据