姜黄素阻断ERK1/2信号通路抑制前列腺癌PC-3细胞增殖的研究

Inhibitory effect of curcumin on PC-3 cell proliferation of prostatic carcinoma by blocking ERK1/2 signaling pathway

目的探讨姜黄素对前列腺癌PC-3细胞株细胞增殖的抑制作用。方法不同浓度(5μmol/L、10μmol/L、20μmol/L、40μmol/L 4个浓度及空白对照组)姜黄素作用于PC-3细胞1 d、2 d、4 d、6 d后,采用噻唑蓝(MTT)法检测细胞增殖抑制情况;实时荧光定量聚合酶链式反应(PCR)检测PC-3细胞细胞外信号调节激酶(ERK)1/2 m RNA表达水平;蛋白质印迹法检测磷酸化ERK1/2(p-ERK1/2)蛋白表达水平。结果 MTT法检测:姜黄素明显抑制前列腺癌PC-3细胞的体外增殖,40μmol/L、20μmol/L、10μmol/L、5μmol/L组在细胞生长1 d、2 d、4 d和6 d后细胞抑制率明显高于对照组,差异均有统计学意义(χ2分别=8.36、7.89、5.89、5.35;8.03、7.69、6.17、5.57;8.84、8.53、7.39、6.76;11.58、9.86、8.20、7.26,P均<0.05),且不同浓度姜黄素实验组在细胞生长1 d、2 d、4 d和6 d,细胞抑制率随药物浓度的增加而上升(F分别=5.65、4.63、4.64、5.76,P均<0.05)。实时荧光定量PCR检测:姜黄素浓度与ERK1/2 m RNA的表达水平成反比,40μmol/L、20μmol/L、10μmol/L、5μmol/L组在细胞生长2 d、4 d、6 d的ERK1/2 m RNA表达均明显低于对照组,差异均有统计学意义(t分别=8.76、8.13、7.69、6.86;8.56、8.16、7.76、6.85;9.03、8.68、8.11、7.27,P均<0.05),且随着作用时间的增加,5μmol/L、10μmol/L、20μmol/L、40μmol/L浓度条件下的ERK1/2 m RNA水平逐渐降低(F分别=4.35、4.99、4.05、4.28,P均<0.05)。蛋白质印迹法:40μmol/L姜黄素抑制ERK1/2的磷酸化呈时间依赖性,即随时间的延长下调pERK1/2表达的活性。结论姜黄素对人前列腺癌PC-3细胞株增殖具有明显的抑制作用,其机制可能与抑制ERK1/2m RNA及p-ERK1/2蛋白的表达有关。

Objective To investigate the inhibitory effect of curcumin on PC-3 cell proliferation of prostatic carcinoma.Methods Different concentrations of curcumin including 5μmol/L,10μmol/L,20μmol/L,40μmol/L and blank acted on PC-3 cells for 1 day,2 days,4 days and 6 days.The inhibition of proliferation was detected by MTT assay.The ERK1/2 mRNA expression level in PC-3 cells was detected by real-time quantitative PCR.The phosphorylated p-ERK1/2 protein expression level was detected by Western blot.Results MTT assay showed that curcumin significantly inhibited the proliferation of PC-3 cells in vitro,and the inhibitory rates of 40μmol/L,20μmol/L,10μmol/L and 5μmol/L curcumin groups were significantly higher than that of the blank group on 1st day,2nd day,4th day and 6th day(χ2=8.36,7.89,5.89,5.35;8.03,7.69,6.17,5.57;8.84,8.53,7.39,6.76;11.58,9.86,8.20,7.26,P<0.05).In the experimental group,with the increasing curcumin concentration,the cell inhibition rate was gradually increased at 1st day,2nd day,4th day and 6th day(F=5.65,4.63,4.64,5.76,P<0.05).The PCR indicated that the concentration of curcumin ester was inversely proportional to the expression level of ERK1/2 mRNA.The expressions of ERK1/2 mRNA in 40μmol/L,20μmol/L,10μmol/L and 5μmol/L curcumin groups were significantly lower than that of the blank group on 1 day,2 days,4 days and 6 days(t=8.76,8.13,7.69,6.86;8.56,8.16,7.76,6.85;9.03,8.68,8.11,7.27,P<0.05).Withtheactiontimegoing,theERK1/2mRNAlevelsof5μmol/L,10μmol/L,20μmol/L,and 40μmol/L curcumin decreased gradually(F=4.35,4.99,4.05,4.28,P<0.05).Western blot showed that 40μmol/L curcumin could down regulated the activity of p-ERK1/2 expression,and displayed the time dependent.Conclusion Curcumin has a significant inhibitory effect on the proliferation of human prostate cancer PC-3 cell line,which maybe related to the inhibition of ERK1/2 mRNA and p-ERK1/2 proteins expression.