摘要
目的研究miR-221/222对人滋养细胞HTR-8/SVneo(简称HTR-8细胞)增殖、凋亡和凋亡相关蛋白的影响,了解其促人滋养细胞凋亡作用,探讨miR-221/222通过促进人滋养细胞凋亡损害胎盘胆汁酸转运功能,参与妊娠期肝内胆汁淤积症(ICP)发生与发展的机制。方法实验分为转染组和阴性对照组,转染组用瞬时转染方法将miR-221/222mimic转入HTR-8细胞中,阴性对照组用相同方法将mimicNC转入HTR-8细胞。转染48h后用RT-qPCR检测转染效率,CCK-8检测HTR-8细胞增殖情况,流式细胞术分析HTR-8细胞凋亡变化,Westernb10t检测HTR-8细胞内凋亡抑制蛋白B-celllymphoma2(简称bcl-2)的表达。数据比较用f检验。结果与阴性对照组(1.14±0.14、1.58±0.14)相比,miR-221/222转染组(25.43±0.80、22.70±0.95)在HTR-8细胞内表达量明显升高,P值均〈0.01,差异具有统计学意义;miR-221/222转染组bcl-2蛋白表达量分别为(0.56±0.03、0.53±0.03),与各自阴性对照组(0.72±0.003、0.76±0.04)相比,蛋白表达量下降,P值均〈0.05,差异具有统计学意义;与miR-221/222阴性对照组(8.827±0.48、11.80±0.45)相比,miR-221/222转染组(42.53±4.47、24.09±2.53)细胞凋亡明显增加,尸值均〈0.01,差异具有统计学意义;m|R-221/222转染组增殖率分别为(0.82±0.02、0.74±0.01),与对照组(1.15±0.08、1.06±0.08)相比增殖被抑制,P值均〈0.05,差异具有统计学意义。结论miR-221/222可能通过下调凋亡抑制蛋白bcl-2的表达促进人滋养细胞凋亡,导致胎盘功能障碍,损害胎盘正常胆汁酸转运功能,其机制可能参与了ICP的发生与发展。
Objective MicroRNA-221/222 is involved in the pathogenesis ofintrahepatic cholestasis of pregnancy (ICP) to promote the apoptosis of placental bile acids through human trophoblastic cells. This study investigates the effects of miR-221/222 on proliferation, apoptosis and apoptosis-related proteins of human trophoblast HTR-8/SVneo (HTR-8 cells) to understand its role in promoting trophoblastic apoptosis. Methods The experiment was divided into transfection group and negative control group. Transient transfection method was used in both groups. The transfection efficiency was detected by RT-QPCR after 48 h transfection. CCK-8 was used to detect the proliferation of HTR-8 ceils and the apoptosis of HTR-8 ceils were analyzed by flow cytometry. Western blot was used to detect the expression of B-cell Lymphoma 2 (Bcl-2) in HTR-8 cells. Data were compared with t-test Results The expression of miR-221/222 transfected group (25.43 ± 0.80, 22.70 ± 0.95) was increased significantly in the HTR-8 cells than that to negative control group ( 1.14 ± 0.14, 1.58 ± 0.14), and P value was 〈 0.01, the difference was statistically significant. The expression of Bcl-2 protein in mir-221/222 transfection group was (0.56 ± 0.03, 0.53 ± 0.03), and the protein expression was decreased compared with negative control group (0.72 ± 0.003, 0.76 4- 0.04). P value was 〈 0.05, the difference was statistically significant, and compared with the mir-221/222 negative control group (8.827 ± 0.48, 11.80 ± 0.45), cell apoptosis of mir-221/222 transfection group (42.53± 4.47, 24.09 ± 2.53) increased significantly, P value was 〈 0.01, and the difference was statistically significant. Proliferation rate in mir- 221/222 transfection group was (0.82 ± 0.02, 0.74±0.01), and proliferation was inhibited, when compared with control group (1.15 ± 0.08, 1.06 ± 0.08), P value was 〈 0.05, and the difference was statistically significant. Conclusion miR-221/222 may promote the apoptosis of human trophoblastic cells by down regulating the expression of apoptosis inhibitory protein bcl-2, leading to placental dysfunction and impairing the normal bile acid transport function of placenta. This mechanism may be involved in the occurrence and development of ICR .
作者
戢兰馨
刘建
Ji Lanxin;Liu Jian(Department of Obstetrics and Gynecology,The Second Affiliated Hospital of Chongqing Medical University,Chongqing 400010,China)
出处
《中华肝脏病杂志》
CAS
CSCD
北大核心
2018年第8期607-611,共5页
Chinese Journal of Hepatology
基金
重庆市渝中区科委科技计划重点项目(20110502)
重庆市卫生计生委医学科研计划重点项目(08-1108-002)