双酚A致雄性生殖损害毒性通路的富集分析及关键凋亡信号通路的实验研究  

Enrichment analysis and experimental study of BPA-induced male reproductive damage toxicity pathways and key apoptosis signaling pathways

作　　者：袁文博 刘文斌[2] 陈洪强[2] 柴子力 张光辉[1] 曹佳[2] YUAN Wenbo;LIU Wenbin;CHEN Hongqiang;CHAI Zili;ZHANG Guanghui;CAO Jia(School of Public Health,Xinxiang Medical University,Xinxiang,Henan Province,453001;Institute of Toxicology,Faculty of Military Preventive Medicine,Army Medical University(Third Military Medical University),Chongqing,400038,China)

机构地区：[1]新乡医学院公共卫生学院,河南新乡453001 [2]陆军军医大学(第三军医大学)军事预防医学系毒理学研究所,重庆400038

出　　处：《第三军医大学学报》2021年第1期38-44,共7页Journal of Third Military Medical University

基　　金：国家重点研发计划(2017YFC1002001);国家自然科学基金重点项目(81630087)。

摘　　要：目的探讨低剂量双酚A(bisphenol A,BPA)暴露导致的雄性生殖损伤中可能涉及的关键毒性通路。方法利用比较毒理基因组学数据库(Comparative Toxicogenomics Database,CTD)筛选BPA毒性相关基因,对其进行功能富集,探索可能的作用通路,确认相关通路中的关键基因。采用终浓度为0(DMSO对照组)、20、40、80μmol/L BPA处理小鼠精原细胞株GC-2细胞72 h,观察细胞形态并检测凋亡发生率,采用RT-qPCR和Western blot检测关键基因mRNA和蛋白表达水平。结果通过对CTD数据库中BPA相关的疾病进行筛选,得到1460个相关基因,GO富集分析发现这些基因在生殖系统发育、生殖结构发育等分子生物过程中有明显富集,KEGG富集分析发现其在HIF-1和PI3K-Akt等信号通路中有明显富集(P<0.01)。可视化分析后得到通路上的关键基因分别为HIF-1α、PKC、P300/CBP以及PI3K、JAK、P53、BCL-2。BPA染毒GC-2细胞,发现凋亡率上升,且存在剂量-效应关系(P<0.05)。与对照组相比,染毒组PI3K和JAK的mRNA表达下调,P53和BCL-2上调(P<0.05);PI3K和BCL-2的蛋白水平下调,P53则上调(P<0.05)。结论PI3K-Akt信号通路在低剂量BPA暴露致GC-2细胞凋亡过程中可能发挥重要作用。Objective To explore the possible key pathways in the male reproductive toxicity induced by exposure to low-dose bisphenol A(BPA).Methods BPA-related genes were screened based on Comparative Toxicogenomics Database(CTD).Functional enrichment analysis was used for these genes to explore the possible pathways and identify the key genes in related pathways.Mouse spermatogonia cell line GC-2 cells were treated with BPA at a final concentration of 0(DMSO control group),20,40 and 80μmol/L for 72 h.Then the changes of cell morphology and apoptotic rate were observed.The mRNA and protein expression levels of the obtained key genes were detected by RT-qPCR and Western blotting,respectively.Results A total of 1460 BPA-related genes were screened out from the CTD database.Gene Ontology(GO)enrichment analysis showed that these genes were significantly enriched in molecular biological processes,such as reproductive system development and reproductive structure development,and Kyoto Encyclopedia of Genes and Genomes(KEGG)analysis found that they were significantly enriched in HIF-1 and PI3K-Akt signaling pathways(P<0.01).Visual analysis showed that the key genes in the pathway were HIF-1α,PKC and P300/CBP,and PI3K,JAK,P53 and BCL-2.The apoptotic rate of GC-2 cells exposed to BPA was significantly increased in a dose-dependent manner(P<0.05).Compared with the control cells,the mRNA expression levels of PI3K and JAK were significantly down-regulated,while those of P53 and BCL-2 were up-regulated(P<0.05).The protein levels of PI3K and BCL-2 were obviously down-regulated,while that of P53 up-regulated(P<0.05).Conclusion The PI3K-Akt signaling pathway may play an important role in the process of GC-2 cell apoptosis induced by low-dose BPA exposure.

关 键 词：双酚A 比较毒理基因组学数据库 GC-2细胞 细胞凋亡 PI3K-AKT信号通路 男性泌尿生殖系统疾病 

分 类 号：R394.1[医药卫生—医学遗传学] R698[医药卫生—基础医学]