Xie Y Y, Qu X X, Zhao H N, Ma M, Xu M T, He C Q
Department of Obstetric, Qinghai University Affiliated Hospital, Xining 810001, China.
Department of Oncology Surgery, Qinghai University Affiliated Hospital, Xining 810001, China.
Zhonghua Fu Chan Ke Za Zhi. 2019 Mar 25;54(3):179-183. doi: 10.3760/cma.j.issn.0529-567x.2019.03.007.
To investigate the effects of human leukocyte-associated antigen-G (HLA-G) expression in silencing trophoblast cell line JEG-3 under normal and hypoxic conditions on invasion and proliferation of JEG-3 cells. Inhibition of HLA-G expression in JEG-3 cells by transfection of small interfering RNA (siRNA),the transfected JEG-3 cells were divided into 4 groups: normoxia control group, hypoxia control group, normoxia inhibition group and hypoxia inhibition group. The levels of HLA-G mRNA and protein in 4 groups of cells were detected by real-time quantitive PCR and western blot. The proliferation activity and invasion ability of 4 groups of cells were determined by methylthiazolyl tetrazolium (MTT) assay and invasion assay. (1) Real-time quantitive PCR technology showed: the level of HLA-G mRNA in the hypoxic inhibition group (0.220±0.050) was significantly different (0.05), when compared with that in the hypoxic control group (0.630±0.030) and normoxic inhibition group (0.400±0.020). (2) Western blot analysis showed: the expression level of HLA-G protein in the hypoxic inhibition group was 0.260±0.010, statistically different from that in the hypoxic control group (0.850±0.100) and the normoxic inhibition group (0.560±0.020; 0.05).(3) MTT showed: proliferative activity of JEG-3 cells in the normoxic inhibition group was 0.490±0.070, the ability of cell proliferation was reduced. When compared with that in the normoxic control group (0.850±0.050), the differences was statistically significant (0.05). The proliferative activity of JEG-3 cells in the hypoxic inhibition group (0.330±0.070) was lower than that in the normoxic inhibition group (0.490±0.070), and there was a significant difference (0.05). (4) Invasion assay showed: compared with the normoxic control group (98±7), the invasive ability of JEG-3 cells in the normoxic inhibition group (73±7) was weakened, and the difference was statistically significant (0.05). The number of transmembrane cells (52±11) of JEG-3 cells in the hypoxic inhibition group was lower than that in the hypoxic control group (72±7), and the difference was statistically significant (0.05). Compared with the normoxic inhibition group, the invasion ability of JEG-3 cells in the hypoxic inhibition group decreased, and the difference was statistically significant (0.05). Under hypoxia, using siRNA technology to down-regulate the expression of HLA-G may affect the proliferation and invasion ability of trophoblast cells, which may be involved in the occurrence of hypertensive disorder of pregnancy.
探讨人白细胞相关抗原G（HLA-G）在正常及缺氧条件下沉默滋养层细胞系JEG-3中的表达对JEG-3细胞侵袭和增殖的影响。通过转染小干扰RNA（siRNA）抑制JEG-3细胞中HLA-G的表达，将转染后的JEG-3细胞分为4组：常氧对照组、缺氧对照组、常氧抑制组和缺氧抑制组。采用实时定量PCR和蛋白质印迹法检测4组细胞中HLA-G mRNA和蛋白水平。采用甲基噻唑基四氮唑（MTT）法和侵袭试验测定4组细胞的增殖活性和侵袭能力。（1）实时定量PCR技术显示：缺氧抑制组HLA-G mRNA水平（0.220±0.050）与缺氧对照组（0.630±0.030）和常氧抑制组（0.400±0.020）相比，差异有统计学意义（P<0.05）。（2）蛋白质印迹分析显示：缺氧抑制组HLA-G蛋白表达水平为0.260±0.010，与缺氧对照组（0.850±0.100）和常氧抑制组（0.560±0.020；P<0.05）相比，差异有统计学意义。（3）MTT法显示：常氧抑制组JEG-3细胞增殖活性为0.490±0.070，细胞增殖能力降低。与常氧对照组（0.850±0.050）相比，差异有统计学意义（P<0.05）。缺氧抑制组JEG-3细胞增殖活性（0.330±0.070）低于常氧抑制组（0.490±0.070），差异有统计学意义（P<0.05）。（4）侵袭试验显示：与常氧对照组（98±7）相比，常氧抑制组JEG-3细胞侵袭能力（73±7）减弱，差异有统计学意义（P<0.05）。缺氧抑制组JEG-3细胞穿膜细胞数（52±11）低于缺氧对照组（72±7），差异有统计学意义（P<0.05）。与常氧抑制组相比，缺氧抑制组JEG-3细胞侵袭能力降低，差异有统计学意义（P<0.05）。在缺氧条件下，利用siRNA技术下调HLA-G的表达可能影响滋养层细胞的增殖和侵袭能力，这可能与妊娠期高血压疾病的发生有关。
