摘要：
目的:研究纳米镍对大鼠睾丸生精-支持共培养细胞的作用及其相关机制.方法:以体外共培养的大鼠睾丸生精-支持共培养细胞为实验对象,分别加入不同浓度的纳米镍(0、25、50、100、200、400和800 μg/mL)染毒24 h后,采用MTT法检测细胞存活率,Hoechst33258染色法检测细胞凋亡形态及Annexin V/PI双染法检测细胞凋亡率;lncRNA基因芯片检测细胞凋亡相关lncRNA的差异表达.结果:与对照组相比,不同浓度纳米镍染毒后,细胞存活率均降低(P＜0.01),并呈剂量效应关系(P＜0.01).Hoechst33258染色结果显示,与对照组比较,纳米镍染毒组细胞核或细胞质内显示出浓染致密的颗粒块状荧光及明显核形态变化.Annexin V/PI双染法流式检测结果显示,染毒浓度为25、50、100、200、400 μg/mL时,细胞凋亡率分别为8.80％±0.50％、11.00％±0.70％、12.53％±0.71％、15.95％±0.54％和17.80％±0.76％,各浓度组细胞凋亡率与对照组比较均明显增加(P＜0.01),且呈浓度效应关系(P＜0.01).LncRNA基因芯片检测结果显示,与凋亡相关的lncRNA共169个,其中,100 μg/mL纳米镍染毒组与对照组比较,上调的有117个,下调的有52个.结论:纳米镍能引起大鼠睾丸生精-支持共培养细胞凋亡,lncRNA可能在其中起重要作用.%OBJECTIVE:Study the effects of nickel nanoparticles (Ni NPs) on apoptosis in SD rat Sertoli-germ cell culture and research on related mechanisms.METHODS:We established a rat Sertoli-germ cell co-culture system in vitro and the cells were treated with Ni NPs.Survive rates were determined by using the MTT assay.Apoptosis morphology were determined by Hoechst33258.Apoptosis index of Sertoli cells and germ cells were determined by Annexin V/PI staining;lncRNA expression profiles were detected to screen out the apoptosis-related genes.RESULTS:The survival rates in the Ni NPs exposure groups (25,50,100,200,400 and 800 μg/mL) were significantly lower than that in the control group (0 μg/mL) after 24 h (P＜0.01).The survival rates decreased as the concentrations of Ni NPs increased.As the Hoehest33258 shows,the treatment groups showed the dense granular luminescence and obvious nuclear morphological changes in the nucleus or cytoplasm.The apoptosis indexes were 8.80％ ± 0.50％,11.00％ ± 0.70％,12.53％ ± 0.71％,15.95％ ± 0.54％ and 17.80％ ± 0.76％ as the doses were 25,50,100,200,400 μg/mL.Apoptosis indexes increased as the concentrations of Ni NPs increased.The apoptosis indexes in the Ni NPs exposure groups were significantly higher than that in the control group (P＜0.01).Compared with the control group,there were 169 lncRNA (117 up and 52 down) associated with apoptosis in the exposure group of 100 μg/mL.CONCLUSION:The study shows that Ni NPs can cause apoptosis of SD rat Sertoli-germ cell co-culture with significant cytotoxicity.LncRNA were closely related to the apoptosis of SD rat co-culture Sertoli-germ cell.
