摘要
目的:探讨芒柄花黄素(Form)对人胃癌细胞株SGC7901的作用及机制.方法:用不同浓度(0、20、40、80 μ mol/L)Form处理SGC7901后,采用Hoechst、MTT、流式细胞术等检测细胞抑制率、细胞周期、细胞核形态及凋亡.采用蛋白质印迹检测β-Catenin、p-β-Catenin、CyclinD1、CDK4及p-AKT、AKT、BCL-2、BAX、Caspase3 等蛋白的表达,RT-PCR 检测 BCL-2、CDK4、CDK6、AKT、CyclinD1 及BAX、Caspase3等mRNA表达.结果:Form抑制SGC7901细胞的增殖,呈浓度和时间依赖性.Hoechst结果表明,相比于对照组(0 μ mol/L),实验组Form(20、40、80 u mol/L)作用48 h后,细胞核固缩、溶解,破裂后呈颗粒状聚集.流式细胞术结果显示,不同浓度Form处理48 h,细胞凋亡率分别为17.0％、21.5％、32.5％,呈浓度依赖性.RT-PCR结果显示,SGC7901中Caspase3、BAX mRNA表达量随Form 浓度增加而上升(P＜0.01);BCL-2、CDK4、CDK6、AKT、CyclinD1 的 mRNA 表达量逐渐降低(P＜0.01).WB 结果表明,SGC7901 细胞中 β-Catenin、p-β-Catenin、CyclinD1、CDK4 以及 p-AKT、AKT、BCL-2等蛋白的相对表达量随Form浓度增加而降低(P＜0.01);Caspase3、BAX等蛋白表达逐渐上升(P＜0.01).结论:Form具有促进SGC7901细胞凋亡的作用;其机制可能是通过抑制Wnt/β-catenin信号,活化Caspase家族实现的.
Abstract
Objective:To investigate the effect of anthoxanthin on the proliferation and apopto-sis of human gastric cancer cell line SGC7901 and its possible mechanism.Methods:SGC7901 was cultured in the medium containing different concentrations(0,20,40,80 μ mol/L)of antho-cyanin.Hoechst staining,MTT colorimetry and flow cytometry were used to verify the effect of Form on the inhibition rate,cell cycle,nuclear morphology and apoptosis.Western Blotting was used to test the level of β-Catenin、p-β-Catenin,CyclinD1,CDK4,p-AKT,AKT,BCL-2,BAX,caspase3 and other proteins.The mRNA expression of BCL-2,CDK4,CDK6,AKT,CyclinD1,BAX,Caspase3 was detected by RT-PCR.Results:MTT results showed that the inhibitory effect of Form on SGC7901 was concentration dependent and time dependent.Hoechst staining showed that the cells in the experimental group(20,40,80 μ mol/L),the phenomenon of pyknosis and dissolution of the nucleus,even granular aggregation after rupture.Flow cytometry showed that the apoptosis rate of the experimental group(20 μ mol/L,40 μ mol/L,80 μ mol/L)was 17.0％,21.5％,32.5％respectively after treated with different concentrations of Form for 48 hours,which was positively correlated with the drug concentration.RT-PCR showed that caspase3 and BAX mRNA expression increased with the increase of Form intervention concentration(P＜0.01);the mRNA expression of BCL-2,CDK4,CDK6,AKT,CyclinD1 decreased(P＜0.01).WB results showed that β-Catenin、p-β-Catenin,Cy-clinD1,CDK4 and p-AKT,AKT,BCL-2 decreased with the increase of Form intervention concentra-tion(P＜0.01);The relative expression of caspase3,BAX and other proteins increased(P＜0.01).Con-clusion:Form can promote apoptosis of SGC7901,and the possible mechanism is to inhibit Wnt/β-Catenin signal;it also mediates the activation of Caspase family.
维普
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