线粒体钙离子摄入蛋白1在血管紧张素Ⅱ诱导小鼠心肌肥大病理变化中的保护作用

Protection of MICU1 against myocardial hypertrophy induced by angiotensin Ⅱ

目的探讨线粒体钙离子摄入蛋白1(MICU1)在血管紧张素Ⅱ(AngⅡ)诱导小鼠心肌肥大病理变化中的作用及其相关机制。方法采用300nmol/L AngⅡ干预小鼠心肌细胞(MCM)48h以构建心肌肥大体外模型,然后以特异性siRNA转染技术敲低MCM中线粒体MICU1水平;另采用腺病毒增加MCM中线粒体MICU1水平。通过qRT-PCR法检测心肌细胞中心钠肽(ANP)、脑钠肽(BNP)mRNA的表达;JC-1及ATP试剂盒分别检测心肌细胞线粒体膜电位及ATP水平;qRT-PCR和Western blotting检测心肌细胞中的MICU1的表达;原子火焰吸收法检测心肌细胞线粒体中Ca^(2+)水平;细胞免疫荧光染色(α-actinin)检测心肌细胞大小。结果 AngⅡ所诱导的肥大心肌细胞中线粒体膜电位水平降低、ATP生成减少(P<0.05);心肌肥大导致MICU1表达下降,同时明显增加了细胞线粒体内Ca^(2+)含量(P<0.05)。通过基因沉默发现,MICU1的下调明显加重了线粒体Ca^(2+)超载(P<0.05),并导致心肌细胞横截面积增大和心肌细胞中BNP的表达增加(P<0.05)。相反,过表达MICU1则能明显减轻线粒体Ca^(2+)超载,使心肌细胞横截面积减小,ANP、BNP的表达降低。结论MICU1通过抵抗线粒体Ca^(2+)超载减轻了AngⅡ所诱导的心肌肥大。

Objective To investigate the role of mitochondrial calcium uptake 1 （MICU1） in myocardial hypertrophy of mice and underlying mechanism. Methods The model of myocardial hypertrophy was established via incubation of mouse cardiac myocytes （MCM） with 300nmol/L angiotensin Ⅱ （Ang Ⅱ ） for 48 hours in vitro. After that, MICU1 specific small interfering RNA （siRNA） was delivered to knockdown MICU1 levels in MCM. On the other hand, adenovirus-mediated over-expression of MICU1 was transfected into MCM. Accordingly, the expressions of ANP and BNP in myocardial cells were measured by qRT- PCR. Mitochondrial membrane potential and ATP contents were detected byJC-1 assay kit and ATP assay kit, respectively. Then, Western blotting and qRT-PCR were used to detect the levels of MICU1 in myocardial cells. The mitochondrial Ca2＋ contents were measured via atomic absorption flame spectroscopy. The size of myocardial cells was determined by α-actinin staining. Results Mitochondrial membrane potential and ATP contents in hypertrophic cardiomyocytes induced by Ang Ⅱ were both decreased. Meanwhile, myocardial hypertrophy significantly increased mitochondrial Ca2＋ contents but decreased MICU1 levels. With the method of genetic intervention, we found that MICU1 deficiency exacerbated mitochondrial Ca2＋ overload, increased cell surface and elevated the expression of BNP. Conversely, the overexpression of MICU1 obviously decreased mitochondrial Ca2＋ overload,cell surface of MCM and expressions of ANP and BNP. Conclusion MICU1 alleviates Ang Ⅱ -induced myocardial hypertrophy via 2＋ mhlbmngmltochondmal Ca overload.