薯蓣皂苷对结肠癌奥沙利铂耐药细胞的作用及其机制  

Effect and mechanism of diosgenin on oxaliplatin-resistant colon cancer cells

作　　者：周超熙[1] 李保坤 马洪庆 张建锋[1] 牛健 于滨[1] 王贵英 赵阳[3] Zhou Chaoxi;Li Baokun;Ma Hongqing;Zhang Jianfeng;Niu Jian;Yu Bin;Wang Guiying;Zhao Yang(The Second Department of General Surgery,Fourth Hospital of Hebei Medical University,Shijiazhuang 050019,China;Department of Gastrointestinal Surgery,the Third Hospital of Hebei Medical University,Shijiazhuang 050051,China;Department of Research Management,the Fourth Hospital of Hebei Medical University,Shijiazhuang 050019,China)

机构地区：[1]河北医科大学第四医院外二科,石家庄050019 [2]河北医科大学第三医院胃肠外科,石家庄050051 [3]河北医科大学第四医院科研处,石家庄050019

出　　处：《中华实验外科杂志》2020年第9期1658-1661,共4页Chinese Journal of Experimental Surgery

摘　　要：目的探讨薯蓣皂苷对耐奥沙利铂(L-OHP)的结肠癌细胞株HT-29/L-OHP影响及其机制。方法常规培养结肠癌细胞株HT-29、耐药细胞株HT-29/L-OHP。噻唑蓝(MTT)法检测0~72 h细胞活性。以5μmol/L的薯蓣皂苷处理细胞后流式细胞术(FCM)检测细胞凋亡;划痕实验及Transwell小室实验检测细胞侵袭和迁移能力;实时定量反转录聚合酶链反应(RT-qPCR)和蛋白质印迹法(Western blot)检测薯蓣皂苷干预48 h对HT-29/L-OHP细胞多药耐药基因1(MDR1)、肺耐药蛋白(LRP)、B细胞淋巴瘤/白血病-2(bcl-2)、bcl-2相关X蛋白(bax)、CXC趋化因子受体7(CXCR7)、基质金属蛋白酶(MMP)-2、Livin及半胱氨酰天冬氨酸特异性蛋白酶(Caspase)-3活性的影响。应用SPSS 25.0软件对数据进行ANOVA分析和t检验。结果MTT结果显示,薯蓣皂苷对HT-29、HT-29/L-OHP细胞有抑制作用,呈时间-剂量依赖特征(P<0.05)。5μmol/L的薯蓣皂苷处理后HT-29/L-OHP细胞凋亡率处理组为(19.05±2.99)%,对照组为(9.07±2.84)%(t=5.928,P<0.01);处理组HT-29/L-OHP细胞迁移率为(50.45±10.77)%,对照组(78.25±10.16)%(t=-4.599,P<0.01);处理组HT-29/L-OHP细胞穿膜数[(93.50±16.12)个],对照组[(150.67±20.85)个,t=-5.313,P<0.01]。薯蓣皂苷处理后HT-29/L-OHP细胞中MDR1、bcl-2、CXCR7、MMP-2明显降低,bax增高(RT-qPCR结果:t=6.052、4.699、6.082、2.882、-5.215,P<0.05;Western blot结果:t=5.569、4.162、5.284、3.438、-4.299,P<0.05);LRP、Livin无明显变化(RT-qPCR结果:t=0.188、0.706,P>0.05;Western blot结果:t=0.184、0.188,P>0.05);HT-29/L-OHP细胞Caspase-3活性在处理组(1.63±0.33),对照组(0.77±0.13)(t=5.939,P<0.01)。薯蓣皂苷与L-OHP联合应用对HT-29/L-OHP的抑制率为(71.42±7.83)%,L-OHP组为(35.90±7.51)%,薯蓣皂苷组为(44.03±6.37)%(F=39.361,P<0.01)。结论薯蓣皂苷可能通过调节多种基因表达而减轻结肠癌细胞对L-OHP的耐药性。Objective To investigate the effect and mechanism of diosgenin on resistance to oxaliplatin(L-OHP)in colon cancer cell line HT-29/L-OHP.Methods Human colon cancer cell line HT-29 and drug-resistant cell line HT-29/L-OHP were cultured routinely.Thiazolyl blue methyl thiazolyl tetrazolium(MTT)assay was applied to detect the effect of Dioscin on the activity of 2 cell lines from 0-72 h.5μmol/L Dioscin was used to detect variation of cells.Flow cytometry(FCM)was used to detect the apoptotic changes of HT-29/L-OHP cells after Dioscin intervention.Wound assay and Transwell chamber assay were used to detect the changes of invasion and migration ability of HT-29/L-OHP cells after intervention.The expression of multidrug resistance 1 gene(MDR1),LRP,B cell lymphoma/leukemia-2(bcl-2),bcl-2 associated X protein(bax),CXC chemokine receptor 7(CXCR7),matrix metalloproteinase(MMP)-2,Livin and cysteinyl aspartate-specific protease(Caspase)-3 activity in HT-29/L-OHP cells were detected by real-time quantitative reverse transcriptase-polymerase chain reaction(RT-qPCR)and Western blotting after Dioscin intervention for 48 h.SPSS 25.0 software was used to analyze the data by ANOVA and t test.Results MTT results showed that Dioscin inhibited HT-29 and HT-29/L-OHP cells in a time-dose-dependent manner(P<0.05).FCM results showed that the apoptosis rate of HT-29/L-OHP cells in Dioscin(5μmol/L)treatment group was(19.05±2.99)%,and(9.07±2.84)%in control group(t=5.928,P<0.01);migration rate in treatment group was(50.45±10.77)%,and(78.25±10.16)%in control group(t=-4.599,P<0.01);cells through the membrane in treatment group was(93.50±16.12),and(150.67±20.85)in control group(t=-5.313,P<0.01).After Dioscin treatment,the expression of MDR1,bcl-2,CXCR7 and MMP-2 in HT-29/L-OHP cells decreased significantly,while the expression of bax increased(RT-qPCR:t=6.052,4.699,6.082,2.882,-5.215,P<0.05;Western blotting:t=5.569,4.162,5.284,3.438,-4.299,P<0.05);the expression of LRP and Livin did not change significantly(RT-qPCR:t=0.188,0.706,P>0.05;Wes

关 键 词：结肠癌 薯蓣皂苷 奥沙利铂 药物抵抗 分子机制 

分 类 号：R285[医药卫生—中药学]