HLA-G表达对滋养细胞系JEG-3细胞侵袭和增殖能力的影响

Effects of human leucocyte antigen-G expression on invasion and proliferation of chorionic trophoblastic cell line JEG-3

目的探讨在常氧和低氧条件下沉默滋养细胞系JEG-3细胞的人类白细胞抗原G(HLA-G)表达,对JEG-3细胞侵袭和增殖能力的影响。方法通过转染小分子干扰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、低氧对照组、常氧抑制组、低氧抑制组。通过实时荧光定量PCR技术及蛋白印迹法检测4组细胞中HLA-G mRNA和蛋白的表达水平,采用四甲基偶氮唑蓝法及体外侵袭实验检测4组细胞的增殖及侵袭能力。结果(1)低氧抑制组JEG-3细胞HLA-GmRNA的表达水平为(0.220±0.050),分别与低氧对照组(0.630±0.030)、常氧抑制组(0.400±0.020)比较,差异均有统计学意义(P均<0.05)。(2)低氧抑制组JEG-3细胞HLA-G蛋白的表达水平为(0.260±0.010),分别与低氧对照组(0.850±0.100)、常氧抑制组(0.560±0.020)比较,差异均有统计学意义(P均<0.05)。(3)常氧抑制组JEG-3细胞的增殖能力(0.490±0.070)与常氧对照组(0.850±0.050)比较,差异有统计学意义(P<0.05);低氧抑制组JEG-3细胞的增殖能力(0.330±0.070)与常氧抑制组(0.490±0.070)比较,差异有统计学意义(P<0.05)。(4)常氧抑制组JEG-3细胞的穿膜细胞数为(73±7)个,与常氧对照组[(98±7)个]比较,差异有统计学意义(P<0.05);低氧抑制组JEG-3细胞的穿膜细胞数为(52±11)个与低氧对照组[(72±7)个]比较,差异有统计学意义(P<0.05);低氧抑制组细胞的穿膜细胞数与常氧抑制组比较,差异有统计学意义(P<0.05)。结论低氧条件下通过siRNA抑制滋养细胞系JEG-3细胞HLA-G的表达,可影响滋养细胞的增殖及侵袭能力,可能参与妊娠期高血压疾病的发生。

Objective 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. Methods 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. Results (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 (P<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;P<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 (P<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 (P<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 (P<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 (P<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 (P<0.05). Conclusion 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.