在低氧环境下沉默EPAS1对JEG-3细胞生物学行为的影响

Effects of silencing EPAS1 on biological behavior of JEG-3 cells in hypoxic environment

目的在常氧及低氧条件下培养人滋养细胞株JEG-3细胞,探讨在低氧环境下沉默内皮PAS蛋白1(EPAS1)对人滋养细胞株JEG-3细胞增殖活性、侵袭行为、凋亡程度及细胞周期分布的影响。方法通过转染小干扰RNA(siRNA)抑制JEG-3细胞中EPAS1的表达,将转染后的JEG-3细胞分为4组(常氧对照组、常氧抑制组、低氧对照组及低氧抑制组)。在此基础上,根据处理时间分为48、72 h组。用CCK8实验检测4组细胞活力;用体外侵袭实验(transwell)检测4组细胞的侵袭能力;用流式细胞术检测不同处理条件对4组细胞凋亡和细胞周期的影响。结果(1)低氧及敲低HIF-2α的表达均可使JEG-3细胞的增殖活性显著降低,差异有统计学意义(P<0.05);经低氧处理72 h的JEG-3细胞的增殖活性较48 h时的降低,差异有统计学意义(P<0.05)。(2)低氧条件及敲低HIF-2α的表达均可使JEG-3细胞的侵袭能力降低,差异有统计学意义(P<0.05)。(3)低氧条件及敲低HIF-2α的表达均可使JEG-3细胞的凋亡率显著增高,差异有统计学意义(P<0.05)。(4)低氧条件及敲低HIF-2α的表达均可使JEG-3细胞的G1相细胞数量增加,表示细胞被阻滞在G1期,差异有统计学意义(P<0.05)。结论在低氧环境下沉默EPAS1使人滋养细胞株JEG-3细胞增殖活性及侵袭能力降低、凋亡率增高且使JEG-3细胞被阻滞在G1期。由此推测,在低氧环境下沉默EPAS1可影响滋养细胞生物学行为,以此参与子痫前期的发生。

Objective The effects of downregulation of EPAS1 on the proliferation,invasion ability,apoptosis,and cell cycle distribution of human trophoblast cell line JEG-3 were studied by culturing JEG-3 cells under normoxia and hypoxic.Methods The expression of EPAS1 in JEG-3 cells was inhibited by transfection of small interfering RNA(siRNA).The JEG-3 cells after transfection were divided into four groups:normoxic control group,normoxic inhibition group,hypoxic control group,and hypoxic inhibition group.Moreover,it was divided into 48 h and 72 h according to the treatment time.CCK8 test was used to detect the proliferation of the four groups of cells;Transwell test was used to detect the invasion ability of the four groups of cells;The effects of four groups of apoptosis and cell cycle were detected by flow cytometry.Results(1)Hypoxic conditions and downregulation of HIF-2αexpression all can significantly reduce the proliferation activity of JEG-3 cells,and the difference was statistically significant(P<0.05);The proliferative activity of JEG-3 cells treated with hypoxic for 72 h was lower than that at 48 h,and the difference was statistically significant(P<0.05).(2)Hypoxic conditions and downregulation of HIF-2αexpression all can significantly reduce the invasion activity of JEG-3 cells,and the difference was statistically significant(P<0.05).(3)Hypoxic conditions and downregulation of HIF-2αexpression all can significantly increase the apoptosis rate of JEG-3 cells,and the diference was statistically significant(P<0.05).(4)Hypoxic conditions and downregulation of HIF-2αexpression all can increase the number of G1 phase cells of JEG-3 cells,indicating that cells are blocked in Gl phase,and the difference was statistically significant(P<0.05).Conclusions Downregulation of EPASl expression under a hypoxic environment reduced the proliferation activity and invasion ability of JEG-3 cells,increased the apoptosis rate,and made JEG-3 cells blocked in Gl phase.It is speculated that downregulation of EPAS1 expression under a hypoxic environment can affect the biological behaviors of trophoblast cells and thus be involved in the occurrence of preeclampsia.