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目的:探讨补肾益气方对正常早孕期人滋养层细胞凋亡的调节作用及机制。方法:体外培养正常早孕期人细胞滋养层细胞,分别加入空白对照组、5%含药血清组、10%含药血清组、20%含药血清组,用噻唑蓝(MTT)法、流式细胞仪检测含药血清作用24、48、72h后细胞的增殖活性,Caspase-3活性检测试剂盒检测细胞Caspase-3酶活性,Western blot检测Fas、FasL、Bcl-2和XIAP蛋白表达。结果:含药血清培养24、48、72h后,细胞在490nm波长的吸光度值增加(P<0.01);sub-G1期细胞减少(P<0.01),S期细胞增加(P<0.01),G2/M期细胞明显减少(P<0.01),Caspase-3酶活性降低;含药血清对Fas和FasL表达均有上调作用,呈剂量依赖式诱导(P<0.05);Bcl-2蛋白表达水平上升,随着浓度增加,表达量也增加,同时伴随XIAP水平升高(P<0.05)。结论:补肾益气方通过提高凋亡抑制蛋白Bcl-2水平,从而增强XIAP表达,抑制Caspase-3活性,抑制细胞死亡。FasL表达水平的上调有利于母胎免疫耐受的维持。
Objective: To investigate the effect and mechanism of Bushen Yiqi Recipe on human trophoblast apoptosis in normal early pregnancy. METHODS: Human trophoblastic cells were cultured in vitro during normal early pregnancy and added into blank control group, 5% drug-containing serum group, 10% drug-containing serum group, and 20% drug-containing serum group, respectively, using MTT method and flow method. The cell proliferation activity of the drug-containing serum after 24, 48, and 72 hours was measured by cytometry. The Caspase-3 activity assay kit was used to detect the activity of Caspase-3, and the expression of Fas, FasL, Bcl-2, and XIAP protein was detected by Western blot. RESULTS: After 24, 48 and 72 hours of culture with drug-containing serum, the absorbance at 490 nm increased (P<0.01); the number of sub-G1 phase decreased (P<0.01), and the number of cells in S phase increased (P<0.01). G2 The number of cells in /M phase was significantly decreased (P<0.01), and the activity of Caspase-3 enzyme was decreased. The drug-containing serum increased the expression of Fas and FasL in a dose-dependent manner (P<0.05). The expression of Bcl-2 protein was increased. As the concentration increased, the expression level also increased, accompanied by elevated XIAP levels (P<0.05). Conclusion: Bushen Yiqi Recipe enhances the expression of apoptosis inhibitory protein Bcl-2, thereby enhancing XIAP expression, inhibiting Caspase-3 activity and inhibiting cell death. The up-regulation of FasL expression levels is beneficial for the maintenance of maternal-fetal immune tolerance.