表没食子儿茶素没食子酸酯在逆转胃癌细胞耐药中的作用及机制探讨  

Role and mechanism research on reversal of 5-fluorouracil resistance by epigallocatechin gallate in gastric cancer drug-resistance cells lines SGC-7901/5-FU

作　　者：唐鸿生[1] 张相良[1] 崔书中[1] 王进[1] 阮强[1] 黄永红[1] 杨定华[2] 

机构地区：[1]广州医科大学附属肿瘤医院腹部肿瘤外科,510095 [2]南方医科大学南方医院肝胆外科,广州510515

出　　处：《中华胃肠外科杂志》2016年第10期1170-1175,共6页Chinese Journal of Gastrointestinal Surgery

基　　金：广州医科大学青年项目（2013A44）;广州市医药卫生科技一般引导项目（20151A011087）;国家自然科学基金青年项目（81201932）;国家自然科学基金面上项目（81372493）

摘　　要：目的探讨表没食子儿茶素没食子酸酯（EGCG）在逆转胃癌氟尿嘧啶（5-Fu）耐药中的作用及分子机制。方法采用大剂量反复冲击联合药物浓度递增法建立5-Fu耐药细胞株（SGC-7901／5-FU）。应用MTT法检测药物处理后亲本细胞和耐药细胞的增殖活性，计算细胞存活率[（实验组A490平均值／空白组4490平均值）×100％]，并使用Graphpadprime6．0软件计算半数抑制浓度（IC50值）和耐药指数（RI，RI=耐药株IC50值／亲本株IC50值）；采用碘化丙啶和膜联蛋白-V双染流式细胞术检测细胞凋亡率；采用Western blot法检测的耐药相关蛋白（ABCG2、P—gP、MDR-1和GST-π）和凋亡相关蛋白（PARP、Survivin、Bax和Bcl-2）的表达水平。结果耐药细胞株较亲本细胞株的IC50值明显升高[（64．7±3．9）μg／ml比（4．1±0．3）μg／ml；t=26．46，P=0．000]，RI为15．6；不同5-FU浓度条件下，耐药细胞株增殖活性均高于亲本细胞株（均P〈0．05）。与单纯5-FU处理相比，经5-FU加EGCG处理后耐药细胞株的IC50值明显降低[（7.3±0．1）μg／ml比（63．1±1．4）μg／ml；t=40．84，P=0．000]，RI为0．12；不同5-Fu浓度条件下，加入EGCG后耐药细胞株的增殖活性均明显降低（均P〈0．05）。流式细胞法检测空白组、5-Fu组、EGCG组和EGCG联合5-Fu组耐药细胞株的细胞凋亡率分别为（3．0±1．0）％、（7．0±1．3）％、（6．0±1．2）％和（18．0±1．4）％，其中EGCG联合5-Fu组的细胞凋亡率明显高于其他3组（F=129．5，P=0．000）。Western blot检测结果显示，经EGCG处理后．耐药细胞中耐药相关蛋白ABCG2、P—gp、MDR-1和GST-π的表达水平明显下降，凋亡标志蛋白PARP和促凋亡蛋白Bax的表达水平明显升高，抗凋亡蛋白Survivin和Bcl-2的表达水平明显下降（均P〈0．05）。结论EGCG可以降低胃癌耐药细胞株SGC-7901对5-Fu的耐药性，其作用可能是通过抑制耐药相Objective To study the role and molecular mechanism of epigallocatechin gallate （EGCG） in reversing drug-resistance to 5-fluorouracil （5-FU） in gastric cancer drug-resistant cell line SGC-7901/5-FU. Methods Drug-resistance gastric cancer cell line （SGC-7901/5-FU） was established by high doses of repeated impact joint drug concentration increment methods. The cell viability of the parent cell line and the drug-resistance cell line were determined by standard MTT assay. Cell survival rate of drug-resistance was calculated by the formula [ （A490 of the treatment group / A490 of the control group） × 100%]. Cell half inhibitory concentration （IC50） and resistance index （RI） were calculated by the Graphpad prime 6.0 software （ RI = IC50 value of drug-resistance cells / IC50 value of parent cells）. The apoptosis rate of SGC-7901/5-FU cells was quantified by flow cytometry after staining with annexin- V and PI. Western blot was used to detect the protein expression of drug-resistance-related proteins （ABCG2, P-gp, MDR-I and GST-π） and apoptosis-related proteins （PARP, Survivin, Bax and bcl-2）. Results IC50 value was significantly increased in drug-resistant ceils compared with parental cells [ （64.7 ± 3.9） rng/L and （4.1±0.3） rag/L, respectively, t = 26.46, P= 0.000], and the RI was 15.6. Proliferation activity in the drug-resistant cells was higher than that in parental cells at different 5-FU concentrations （all P 〈0.05）. In drug-resistant cells, the IC50 value of 5-FU combined with EGCG group obviously decreased compared with 5-FU group [（7.3 ± 0.1） mg/L and （63.1 ± 1.4） mg/L respectively, t = 40.84, P = 0.000], and the RI was 0.12. Proliferation activity in drug-resistant cells was significantly decreased after EGCG treatment at different 5-FU concentrations （all P 〈 0.05）. Cell apoptosis rates in control group, 5-FU group, EGCG group and 5-FU combined with EGCG group were （3.0 ± 1.0）%, （7.0 ± 1.3）%, （6.0 ± 1.2）% and （18.0 ± 1.4�

关 键 词：胃肿瘤 表没食子儿茶素没食子酸醋 氟尿嘧啶 耐药 

分 类 号：R735.2[医药卫生—肿瘤]