ALC对缺氧精原细胞线粒体呼吸功能及自噬的影响  

Effects of ALC on mitochondrial respiratory function and autophagy in spermatogonia cells under hypoxia

作　　者：蓝贤娜 李敏瑜 乔娜[1] 陈汉明 曾绮雯[1] 刘冰贤 庞聪颖 唐兆新[1] 李英[1] LAN Xianna;LI Minyu;QIAO Na;CHEN Hanming;ZENG Qiwen;LIU Bingxian;PANG Congying;TANG Zhaoxin;LI Ying(College of Veterinary Medicine,South China Agricultural University,Guangzhou 510642,China)

机构地区：[1]华南农业大学兽医学院,广州510642

出　　处：《黑龙江畜牧兽医》2021年第11期6-11,16,共7页Heilongjiang Animal Science And veterinary Medicine

基　　金：国家重点研发计划项目(2017YFD0502203)。

摘　　要：为了探讨乙酰左旋肉碱(ALC)对缺氧条件下小鼠精原细胞线粒体呼吸功能与自噬的影响,试验以GC-1精原细胞系为研究对象,将其分为3组,分别为对照组(5%CO2和95%空气)、缺氧组(5%CO2和3%O2)和缺氧+ALC组(5%CO2和3%O2,培养基中添加150 mmol/L的ALC),各组细胞经相应条件处理24 h后,以高分辨率呼吸仪检测线粒体呼吸功能相关指标的变化,使用实时荧光定量PCR方法检测自噬相关基因的相对表达量,通过Western-blot法检测自噬相关蛋白的表达情况。结果表明:与对照组相比,缺氧组GC-1精原细胞线粒体基础呼吸、质子漏、ATP相关呼吸、线粒体最大呼吸、备用呼吸能力和呼吸控制比率均极显著降低(P<0.01),剩余氧通量升高但差异不显著(P>0.05),而添加ALC后对缺氧诱导的上述指标的变化无明显改善作用(P>0.05)。与对照组相比,缺氧组ATG5、Beclin1、p62基因相对表达量极显著下调(P<0.01),LC3b基因相对表达量轻微上调但差异不显著(P>0.05),Beclin1、ATG5、LC3b/LC3a蛋白相对表达量显著下调(P<0.05)。与缺氧组相比,添加ALC显著或极显著逆转缺氧诱导的自噬相关基因ATG5、Beclin1、p62相对表达量的下调(P<0.05或P<0.01),同时添加ALC显著上调LC3b基因相对表达量(P<0.05);LC3b/LC3a蛋白相对表达量极显著上调(P<0.01),而ATG5、Beclin1蛋白相对表达量呈上调趋势但差异不显著(P>0.05)。说明ALC可能激活缺氧环境下诱导的GC-1精原细胞自噬抑制,但不能逆转线粒体呼吸功能障碍。To investigate the effects of acetyl-L-carnitine(ALC) on mitochondrial respiratory function and autophagy in mouse spermatogonia cell line under hypoxia, GC-1 spermatogonia cell line was used as the research object, which was devided into 3 groups: control group(5%CO2 and 95%AIR), hypoxia group(5%CO2 and 3%O2) and hypoxia+ALC group(5%CO2 and 3%O2, the medium contained with 150 mmol/L ALC). After treatment for 24 hours, the changes of related indexes of mitochondrial respiratory function were detected by high-resolution respirator. The mRNA relative expression levels of autophagy related-genes were detected by qRT-PCR, and the expression levels of autophagy related proteins were detected by Western-blot. The results showed that compared with control group, the basal respiration, proton leak, ATP-linked respiration, maximal respiration, spare respiratory capacity and respiratory control rate were extremely significantly decreased(P<0.01), while the residual oxygen consumption increased slightly in hypoxia group(P>0.05). However, ALC supplementation did not significantly improve the changes of above indexes induced by hypoxia(P>0.05). Compared with control group, the mRNA relative expression levels of ATG5, Beclin1, p62 genes in hypoxia group were extremely significantly down-regulated(P<0.01), the mRNA relative expression levels of LC3 b were slightly up-regulated but the difference was not significant(P>0.05), and the expression of autophagy related protein Beclin1, ATG5, LC3 b/LC3 a down-regulated significantly(P<0.05). Compared with hypoxia group, the supplementation of ALC significantly or very significantly reversed the down-regulation of mRNA expression of autophagy related genes ATG5, Beclin1, p62(P<0.05 or P<0.01), while the supplementation of ALC significantly up-regulated the relative expression levels of LC3 b(P<0.05). The relative expression levels of LC3 b/LC3 a protein were extremely significantly up-regulated(P<0.01), while the relative expression levels of ATG5 and Beclin1 protein were up-regulated

关 键 词：乙酰左旋肉碱(ALC) 缺氧 线粒体功能 呼吸指标 自噬 精原细胞 

分 类 号：Q731[生物学—分子生物学]