王祺, 马璐, 张爱华. 亚砷酸钠对人正常肝细胞线粒体功能及SIRT1/PGC-1α通路相关蛋白表达的影响[J]. 环境与职业医学, 2022, 39(12): 1411-1416. DOI: 10.11836/JEOM22152
引用本文: 王祺, 马璐, 张爱华. 亚砷酸钠对人正常肝细胞线粒体功能及SIRT1/PGC-1α通路相关蛋白表达的影响[J]. 环境与职业医学, 2022, 39(12): 1411-1416. DOI: 10.11836/JEOM22152
WANG Qi, MA Lu, ZHANG Aihua. Effects of sodium arsenite on mitochondrial function and expression of SIRT1/PGC-1α pathway-related proteins in human normal liver cell[J]. Journal of Environmental and Occupational Medicine, 2022, 39(12): 1411-1416. DOI: 10.11836/JEOM22152
Citation: WANG Qi, MA Lu, ZHANG Aihua. Effects of sodium arsenite on mitochondrial function and expression of SIRT1/PGC-1α pathway-related proteins in human normal liver cell[J]. Journal of Environmental and Occupational Medicine, 2022, 39(12): 1411-1416. DOI: 10.11836/JEOM22152

亚砷酸钠对人正常肝细胞线粒体功能及SIRT1/PGC-1α通路相关蛋白表达的影响

Effects of sodium arsenite on mitochondrial function and expression of SIRT1/PGC-1α pathway-related proteins in human normal liver cell

  • 摘要: 背景

    长期砷暴露可引起不同程度的肝损伤,线粒体损伤可能是砷致肝损伤的早期关键事件,沉默信息调节因子2(sir2)相关酶1(SIRT1)/过氧化物酶体增殖物受体辅助激活因子-1α(PGC-1α)是调控线粒体质量和功能的重要通路,而砷所致肝损伤是否与SIRT1/PGC-1α通路介导的线粒体功能障碍有关目前尚不清楚。

    目的

    探讨亚砷酸钠(NaAsO2)对人正常肝细胞线粒体功能及SIRT1/PGC-1α通路相关蛋白表达的影响及机制。

    方法

    以人正常肝细胞(MIHA细胞)为研究对象,分别以不同浓度的NaAsO2(0、5、10、20 μmol·L−1)处理MIHA细胞24 h后收集细胞进行研究。采用透射电镜观察线粒体超微结构,荧光法检测三磷酸腺苷(ATP)浓度,流式细胞术检测线粒体膜电位(MMP)水平,免疫印迹法检测SIRT1、PGC-1α及其下游核呼吸因子1(NRF1)抗体、线粒体转录因子A(TFAM)蛋白表达水平。采用单因素方差分析及趋势性检验进行数据统计分析。

    结果

    MIHA细胞活力随NaAsO2浓度的升高逐渐降低(F=6495.47,P<0.001)。透射电镜结果显示10 μmol·L−1 NaAsO2处理组线粒体大小不一、肿胀或伸长呈棒状,20 μmol·L−1 NaAsO2处理组线粒体肿胀呈气球状或空泡状。MIHA细胞ATP浓度及MMP水平均随NaAsO2浓度的升高逐渐降低(FATP趋势=172.28,FMMP趋势=59.91;均P<0.001)。与对照组相比,5 μmol·L−1 NaAsO2处理组SIRT1、PGC-1α、NRF1及TFAM蛋白表达水平均无明显变化,但10 μmol·L−1 NaAsO2处理组SIRT1、PGC-1α及TFAM蛋白表达水平降低(P<0.05),20 μmol·L−1 NaAsO2处理组SIRT1、PGC-1α及NRF1蛋白表达水平降低(P<0.05);趋势性检验结果显示,SIRT1、PGC-1α、NRF1及TFAM蛋白表达水平均随NaAsO2浓度的升高逐渐降低(FSIRT1趋势=47.07,P<0.001;FPGC-1α趋势=15.17,P<0.01;FNRF1趋势=13.54,P<0.01;FTFAM趋势=4.20,P<0.05)。

    结论

    SIRT1/PGC-1α及其下游NRF1和TFAM的表达下调可能参与了NaAsO2诱导肝细胞线粒体功能障碍。

     

    Abstract: Background

    Long-term exposure to arsenic can cause liver injury of varying degrees. Mitochondrial damage may be an early key event of arsenic-induced liver injury. Silent mating type information regulation 2 homolog 1 (SIRT1)/ recombinant peroxisome proliferators-activated receptor gamma coactivator 1 alpha (PGC-1α) is an important pathway regulating mitochondrial mass and function. However, whether arsenic-induced liver injury is related to mitochondrial dysfunction mediated by SIRT1/PGC-1α pathway remains unclear.

    Objective

    To investigate potential effects of sodium arsenite (NaAsO2) on mitochondrial function and expressions of SIRT1/PGC-1α pathway-related proteins in human normal liver cell.

    Methods

    Human normal liver cells (MIHA cells) were used as the research object. MIHA cells were treated with different concentrations of NaAsO2 (0, 5, 10 and 20 μmol·L−1) for 24 h, and the cells were collected for study. The ultrastructure of mitochondria was observed by transmission electron microscopy, adenosine triphosphate (ATP) concentration by fluorescence method, mitochondrial membrane potential (MMP) level by flow cytometry, and SIRT1, PGC-1α and their downstream nuclear respiratory factor 1 (NRF1) and mitochondrial transcription factor A (TFAM) protein expression levels by Western blotting. One-way analysis of variance and trend test were used for data statistical analysis.

    Results

    The viability of MIHA cells decreased gradually with the increase of NaAsO2 concentration (F=6495.47, P<0.001). The transmission electron microscope observation showed that the size of mitochondria in the 10 μmol·L−1 NaAsO2 treatment group was different, and the mitochondria were swollen or elongated in a rod-like shape. The mitochondria in the 20 μmol·L−1 NaAsO2 treatment group swelled like air spheres or vacuoles. The ATP concentration and MMP level of MIHA cells gradually decreased with the increase of NaAsO2 concentration (Ftrend of ATP=172.28, Ftrend of MMP=59.91, both Ps<0.001). Compared with the control group, the protein expression levels of SIRT1, PGC-1α, NRF1, and TFAM were not significantly changed in the 5 μmol·L−1 NaAsO2 treatment group, while the protein expression levels of SIRT1, PGC-1α, and TFAM were decreased in the 10 μmol·L−1 NaAsO2 treatment group, and the protein expression levels of SIRT1, PGC-1α, and NRF1 were decreased in the 20 μmol·L−1 NaAsO2 treatment group. The results of trend test showed that the protein expression levels of SIRT1, PGC-1α, NRF1, and TFAM decreased gradually with the increase of NaAsO2 concentration (Ftrend of SIRT1=47.07, P<0.001; Ftrend of PGC-1α=15.17, P<0.01; Ftrend of NRF1=13.54, P<0.01; F trend of TFAM=4.20, P<0.05).

    Conclusion

    The down-regulation of SIRT1/PGC-1α and its downstream NRF1 and TFAM may be involved in NaAsO2-induced mitochondrial dysfunction in liver cells.

     

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