黄连素逆转人结肠癌细胞奥沙利铂耐药性的作用及机制

The role and mechanism of berberine in reversing oxaliplatin resistance in human colon cancer cells

目的探究黄连素(Ber)逆转人结肠癌细胞奥沙利铂(OXA)耐药性的作用及机制。方法选用人结肠癌耐OXA细胞THC-8307/OXA进行实验,分为对照组、OXA组和Ber(5、10、20μg/ml)+OXA组。采用倒置相差显微镜观察细胞形态,MTT法检测细胞存活率,流式细胞术检测细胞凋亡率,RT-qPCR检测MDR-1 mRNA的表达,Western blot检测P-gp、PI3K、Akt和p-Akt蛋白的表达。结果与OXA组比较,Ber(5、10、20μg/ml)+OXA组的细胞存活率显著降低(P<0.05),凋亡率显著升高(P<0.05);MDR-1 mRNA、P-gp蛋白、p-Akt蛋白、PI3K蛋白的表达水平及p-Akt/Akt值均显著降低(P<0.05)。其中Ber(10、20μg/ml)+OXA组的效应显著高于Ber(5μg/ml)+OXA组,Ber(20μg/ml)+OXA组的效应显著高于Ber(10μg/ml)+OXA组。OXA组与对照组的检测结果比较差异无统计学意义(P>0.05)。结论Ber可显著逆转THC-8307/OXA细胞对OXA的耐药性,其作用机制可能与PI3K/Akt信号通路抑制,P-gp蛋白表达下调有关。

Objective To explore the effect and mechanism of berberine(Ber)in reversing the resistance of human colon cancer cells to oxaliplatin(OXA).Methods Human colon cancer OXA-resistant cells THC-8307/OXA were selected for experiment,and they were divided into control group,OXA group and Ber(5,10,20μg/ml)+OXA groups.Cell morphology was observed by inverted phase contrast microscope.Cell survival rate was detected by methyl thiazolyl tetrazolium[JP4](MTT)assay method,and cell apoptosis rate was detected by flow cytometry.Multidrug resistanc-1(MDR-1)mRNA expression was detected by real-time quantitative reverse transcription-polymerase chain reaction(RT-qPCR),and the expressions of P-glycoprotein(P-gp),phosphatidylinositol 3-kinase(PI3K),protein kinase B(Akt)and phosphorylated protein kinase B(p-Akt)proteins were detected by Western blot.Results Compared with those of the OXA group,the cell survival rates of Ber(5,10,20μg/ml)+OXA groups were significantly reduced(P<0.05),and the apoptosis rates were significantly increased(P<0.05);the expression levels of MDR-1 mRNA,P-gp protein,p-Akt protein,PI3K protein,and p-Akt/Akt value were significantly reduced(P<0.05).Among them,the effects of Ber(10,20μg/ml)+OXA groups were significantly higher than those of Ber(5μg/ml)+OXA group,and the effect of Ber(20μg/ml)+OXA group was significantly higher than that of Ber(10μg/ml)+OXA group.There were no significant differences in the detection results between the OXA group and the control group(P>0.05).Conclusion Ber can significantly reverse the resistance of THC-8307/OXA cells to OXA,and its mechanism of action may be related to the inhibition of PI3K/Akt signaling pathway and the down-regulation of P-gp protein expression.