褪黑素缓解玉米赤霉烯酮引发的卵巢颗粒细胞氧化应激与雌二醇合成异常

Melatonin alleviates zearalenone⁃induced oxidative stress and abnormal synthesis of estradiol in ovarian granulosa cells

旨在研究玉米赤霉烯酮(Zearalenone,ZEN)对卵巢颗粒细胞的毒性作用以及对雌二醇(Estradiol,E2)合成的影响,为筛选缓解ZEN毒性的药物提供参考.试验利用人颗粒细胞系(KGN)建立ZEN细胞攻毒模型,采用CCK⁃8法检测细胞活力,使用试剂盒检测细胞活性氧及线粒体膜电位水平,利用RT⁃qPCR检测E2合成基因的mRNA表达水平;同时,筛选缓解ZEN毒性的药物并验证其挽救作用.结果显示,添加ZEN(0、30、90μM)后,细胞活力随着ZEN浓度的升高而显著降低(P<0.05),细胞内ROS水平增加并且线粒体膜电位水平显著下降(P<0.05),E2合成基因CYP19A1、CYP11A1、STAR及HSD3B的mRNA表达水平显著下调(P<0.05).与ZEN处理组相比,联合添加褪黑素(Melatonin,MT)可以增加活细胞数量并显著提高细胞活力(P<0.05),同时,E2合成基因CYP19A1、CYP11A1、STAR及HSD3B的mRNA表达水平显著上调(P<0.05),细胞内ROS水平及细胞膜电位水平显著恢复.综上,ZEN导致KGN细胞氧化应激及E2合成紊乱,而MT可部分缓解上述中毒症状,提示MT可用于缓解ZEN毒性.

The aim of this experiment was to study the toxic effects of zearalenone(ZEN)on ovarian granulosa cells and its effect on estradiol(E2)synthesis,and to provide a reference for the screening of drugs to alleviate the toxicity of ZEN.In the present study,human granulosa cell line(KGN)was utilized to establish a ZEN cell model.Cell viability was assessed using the CCK⁃8 method,while levels of reactive oxygen species(ROS)and mitochondrial membrane potential were measured using commercial kits.Additionally,mRNA expression levels of genes involved in E2 synthesis were determined through RT⁃qPCR a⁃nalysis.Simultaneously,compounds with potential alleviating effects on ZEN toxicity were screened and their rescue efficacy was validated.The results showed that the cell viability was significantly decreased with the increase of ZEN concentration(P<0.05).Meanwhile,the intracellular ROS level increased and the mitochondrial membrane potential level decreased significantly(P<0.05).Additionally,the mRNA expression levels of E2 synthesis genes CYP19A1,CYP11A1,STAR and HSD3B were sig⁃nificantly downregulated(P<0.05).Compared with the ZEN treatment group,the combined addition of melatonin(MT)could increase the number of living cells and significantly improve the cell viability(P<0.05).The mRNA expression levels of E2 synthesis genes CYP19A1,CYP11A1,STAR and HSD3B were significantly upregulated(P<0.05),and the intracellular ROS level and cell membrane potential level were significantly restored.In summary,ZEN caused oxidative stress and disordered E2 synthesis in KGN cells,while MT partially alleviated the above toxic symptoms,suggesting that MT could be used to alleviate the toxicity of ZEN.