基于PI3K/AKT信号通路探讨苦参碱对人皮肤鳞状细胞癌A431细胞的作用研究

Study on the effect of matrine on human skin squamous cell carcinoma A431 cells based on PI3K/AKT signaling pathway

目的基于PI3K/AKT信号通路探讨苦参碱对人皮肤鳞状细胞癌A431细胞的影响。方法体外培养人皮肤鳞状细胞癌A431细胞株,将处于对数生长期的人皮肤鳞状细胞癌A431细胞分为空白对照组和10、20、40、80 nmol/L苦参碱组。采用CCK-8法检测人皮肤鳞状细胞癌A431细胞增殖抑制率;ELISA法检测人皮肤鳞状细胞癌A431细胞培养上清TNF-α和IL-1α的浓度;real-time qPCR法检测人皮肤鳞状细胞癌A431细胞Bax和Bcl-2基因表达;Western blot法检测人皮肤鳞状细胞癌A431细胞AKT蛋白的表达水平;分子对接验证苦参碱与AKT的结合活性。结果苦参碱给药组对人皮肤鳞状细胞癌A431细胞的增殖具有明显的抑制作用。与空白对照组比较,苦参碱给药组显著降低炎症因子TNF-α、IL-1α浓度(P<0.05),显著降低人皮肤鳞状细胞癌A431细胞中Bcl-2 mRNA和AKT蛋白的表达水平(P<0.05),升高Bax mRNA表达水平(P<0.05)。苦参碱与AKT具有较好的结合活性。结论苦参碱可抑制人皮肤鳞状细胞癌A431细胞的增殖并诱导其凋亡,其机制可能是通过抑制PI3K/AKT信号通路中AKT蛋白的表达,调节凋亡因子Bax和Bcl-2的表达水平,以及下调细胞炎症相关因子TNF-α和IL-1α的浓度有关。

Objective To investigate the effect of matrine on human skin squamous cell carcinoma A431 cells based on PI3K/AKT signaling pathway.Methods A431 cells cultured with logarithmic growth stage were divided into the control group and different dosages of matrine groups.The inhibition rate of cell proliferation was detected by CCK-8 method.Levels of TNF-αand IL-1αin culture supernatant of A431 cells were detected by ELISA.The expressions of Bax and Bcl-2 mRNA in A431 cells were detected by real-time qPCR,and the expression of AKT in A431 cells was detected by western blot.Molecular docking was used to verify the binding activity of matrine with AKT.Results Matrine significantly inhibited the proliferation of A431 cells.Compared with the control group,matrine significantly reduced the levels of TNF-αand IL-1α,suppressed the expression of Bcl-2 mRNA and AKT in A431 cells,but increased the expression of Bax mRNA(P<0.05).Matrine had good binding activity with AKT.Conclusion Matrine can inhibit the proliferation and induce the apoptosis of A431 cells.The mechanism may be related to the inhibition of AKT expression in PI3K/AKT pathway,the regulation of the expression of Bax and Bcl-2,and the downregulation of TNF-αand IL-1αlevels.