摘要
目的探究普罗布考对过氧化氢(H_(2)O_(2))介导的小鼠睾丸间质细胞系(TM3)损伤的影响。方法体外培养TM3细胞,分为3组:对照组、H_(2)O_(2)损伤模型组和普罗布考处理组。CCK8检测细胞存活率,流式细胞术检测细胞凋亡率,活性氧(ROS)检测试剂盒检测细胞内ROS水平,ELISA测定细胞上清睾酮水平,实时荧光定量PCR检测细胞内睾酮合成酶类固醇激素合成急性调节蛋白(StAR)和17β-羟基类固醇脱氢酶(HSD-17β)、细胞凋亡指标Bax、Caspase3和氧化应激相关指标核因子E2相关因子2(NRF2)、血红素加氧酶-1(HO-1)的mRNA水平,Western blot检测细胞内Bax和Caspase3蛋白表达改变。结果与对照组比较,H_(2)O_(2)损伤模型组细胞存活率显著降低、凋亡率显著升高(P<0.05),Bax和Caspase3表达显著上调(P<0.05),细胞内ROS水平显著升高、NRF2和HO-1水平显著降低(P<0.05),睾酮分泌量和睾酮合成酶表达显著降低(P<0.05);与H_(2)O_(2)损伤模型组比较,普罗布考处理组细胞存活率显著升高、凋亡率显著下降(P<0.05),Bax和Caspase3表达显著下调(P<0.05),细胞内ROS水平显著下降、NRF2和HO-1水平显著升高(P<0.05),睾酮分泌量和睾酮合成酶表达增高(P<0.05)。结论普罗布考可以减轻H_(2)O_(2)诱导的TM3细胞凋亡和睾酮合成异常,这种保护作用可能与普罗布考的抗氧化和抗凋亡效应有关。
Objective:To investigate the effects of probucol on H_(2)O_(2)-mediated Leydig cells(TM3)injury.Methods:TM3 cells were cultured in vitro and divided into three groups:control group,H_(2)O_(2)injury group and probucol treatment group.The cell survival rate was detected by CCK8,the cell apoptosis rate was detected by flow cytometry,and intracellular reactive oxygen species(ROS)level was determined by ROS test kit.The testosterone level in cell supernatant was detected by ELISA,and the mRNAs of intracellular testosterone synthase steroidogenic acute regulatory protein(StAR)&17β-hydroxysteroid dehydrogenase(HSD-17β),apoptosis indicator Bax,Caspase3 and oxidative stress-related indicator nuclear factor erythroid 2-related factor 2(NRF2)and heineoxygenase-l(HO-1)were detected by real-time PCR.The protein expression of Bax and Caspase3 was detected by Western blot.Results:Compared with the control group,the cell survival rate was significantly decreased and the apoptosis rate was significantly increased(P<0.05),the expression of Bax and Caspase3 was significantly up-regulated(P<0.05),the intracellular ROS levels were significantly increased(P<0.05),the levels of NRF2 and HO-1 were significantly decreased(P<0.05),and the secretion of testosterone and the expression of testosterone synthase were significantly decreased(P<0.05)in the H_(2)O_(2)injury group.Compared with the H_(2)O_(2)injury group,the cell survival rate was significantly increased(P<0.05),the apoptosis rate was significantly decreased(P<0.05),the expression of Bax and Caspase3 was significantly down-regulated(P<0.05),intracellular ROS level was significantly decreased(P<0.05),NRF2 and HO-1 levels were significantly increased(P<0.05),and testosterone secretion and testosterone synthase expression were significantly increased(P<0.05)after probucol treatment.Conclusions:Probucol can alleviate H_(2)O_(2)induced apoptosis and abnormal testosterone synthesis of TM3 cells,and this protective effect may be related to the effect of antioxidant and anti-apoptotic effects of probucol.
作者
罗凌波
穆杨
杨菁
LUO Ling-bo;MU Yang;YANG Jing(Reproductive Medical Center,Renmin Hospital of Wuhan University,Wuhan430060)
出处
《生殖医学杂志》
CAS
2023年第3期420-426,共7页
Journal of Reproductive Medicine
基金
国家自然科学基金(81971356)
国家自然科学基金青年项目(82001609)。
