Regulatory effects of miR-15a,miR-466b-3p,miR-328-3p,and miR-144-5p on neodymium nitrate induced proliferation and apoptosis of chorionic villus in small for gestational age infants
Regulatory effects of miR-15a,miR-466b-3p,miR-328-3p,and miR-144-5p on neodymium nitrate induced proliferation and apoptosis of chorionic villus in small for gestational age infants
张宏波 1郭婧 2张琦 1霍宇婷 3程霞霞 3邰大鹏 1乔瑞1
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作者信息
1. 包头医学院,内蒙古 包头 014040
2. 包头市国药北方医院;包头市昆都仑区医院
3. 包头市国药北方医院
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摘要
目的:探究硝酸钕[Nd(NO3)3]对小于胎龄儿(small for gestational age,SGA)绒毛膜细胞凋亡及标志性microRNA表达变化的影响,揭示钕致SGA的潜在分子作用机制.方法:选取人绒毛膜细胞系(JEG-3),通过CCK-8确定Nd(NO3)3最佳染毒剂量、时间;流式细胞术检测细胞凋亡情况;提取对照组(0 μg/mL)、实验组[低、中、高剂量组(0.2、2、20 μg/mL)]细胞的miRNA进行qPCR,检测 miR-15a、miR-466b-3p、miR-328-3p、miR-144-5p的表达水平.转染 miR-15a、miR-144-5p抑制剂后检测细胞转染情况,再次进行CCK-8、流式细胞术验证细胞凋亡情况.结果:CCK-8结果显示,中剂量组细胞存活率下降至75.53％,流式细胞检测同样发现在中剂量组出现了更多凋亡.对JEG-3细胞染毒24 h提取细胞miRNA检测其miR-15a、miR-466b-3p、miR-328-3p、miR-144-5p表达水平,与对照组相比均出现不同程度上调.JEG-3细胞系转染上述microRNA抑制剂后,CCK-8中剂量组miR-144-5p、miR-15a抑制剂转染组其存活率由原75.53％回升至76.71％、83.03％,而凋亡水平则由原18.70％下降至18.44％、15.34％.结论:硝酸钕对JEG-3毒性影响呈现剂量依赖性,miR-15a、miR-466b-3p、miR-328-3p、miR-144-5p表达水平出现上调,激活下游信号通路调节细胞凋亡,对硝酸钕引起的细胞应激反应具有重要影响,为探究SGA的分子作用机制提供了新的研究方向.
Abstract
Objective:To investigate the effects of neodymium nitrate[Nd(NO3)3]on chorionic cell apoptosis and the changes of microRNA expression in small for gestational age(SGA)infants,aiming to explore the molecular mecha-nisms of neodymium inducing SGA.Methods:The human trophoblast cell line(JEG-3)was selected.The optimal dose and time of Nd(NO3)3 exposure were determined by CCK-8 assay.Apoptosis was measured using flow cytometry.MicroRNA(miRNA)was extracted from the control group(0 p.g/mL)and experimental group treated with low,medi-um,and high doses(0.2,2,20 μg/mL)of Nd(NO3)3.The expression levels of miR-15a,miR-466b-3p,miR-328-3p,and miR-144-5p were measured by qPCR.After transfection with miR-15a and miR-144-5p inhibitors,transfection efficiency was evaluated,and CCK-8 and flow cytometry assays were repeated to confirm their effects on apoptosis.Results:The CCK-8 assay showed that cell viability in the medium-dose group decreased to 75.53％,and Flow cytometry also indicated increased apoptosis in this group.After 24 hours of Nd(NO3)3 exposure,qPCR analysis showed that miR-15a,miR-466b-3p,miR-328-3p,and miR-144-5p were upregulated com-pared to the control group.After JEG-3 cell line transfected with the above microRNA inhibitors,the survival rate of miR-144-5p and miR-15a inhibitor transfected group in CCK-8 medium-dose group increased from 75.53％to 76.71％and 83.03％,respectively,while the apoptosis level decreased from 18.70％to 18.44％and 15.34％respec-tively.Conclusion:The toxic effect of[Nd(NO3)3]on cells is dose-dependent.With increased expression levels of miR-15a,miR-466b-3p,miR-328-3p,and miR-144-5p,downstream signaling pathway is activated and ap-optosis mechanism is regulated,which plays an important role in cell stress response induced by(Nd(NO3)3).This study provides a new research direction for exploring the molecular mechanism of SGA.
关键词
稀土元素钕/小于胎龄儿/胎盘/miRNA表达/凋亡
Key words
Neodymium nitrate/Small for gestational age/Placenta/miRNA expression/Apoptosis
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