金雀异黄素联合5-氟尿嘧啶抑制人结肠癌细胞株SW48O增殖的分子机制  

Investigation of the molecular mechanism how genistein combined with 5-fluorouracil inhibits proliferation of human colon cancer cell line SW480

作　　者：凌航[1] 陈裕庆[2] 高美钦[2] 张文敏[2] 

机构地区：[1]福建省立医院病理科,福州350001 [2]福建医科大学基础医学院病理学系

出　　处：《中华消化杂志》2015年第6期390-394,共5页Chinese Journal of Digestion

基　　金：福建省新世纪优秀人才项目（NCETFJ-0603）;福建省自然科学基金（H12100564）

摘　　要：目的初步探讨金雀异黄素提高结肠癌细胞对5-氟尿嘧啶（5-Fu）的敏感性，促进结肠癌细胞凋亡的分子机制。方法选择SW480结肠癌细胞株为实验对象，将金雀异黄素80μmol／L和5-Fu30μg／mL单用或联用48h作为药物实验组，另设不加药组为对照组。分别采用实时定量PCR和Western印迹法检测各组细胞的生存素、Bcl2、p21、caspase3和caspase9基因和蛋白表达情况。采用比较阈值法（2^-ΔΔCT法）来确定基因的相对表达量。以β-actin作为内参照，进行半定量分析计算蛋白相对表达量。电泳迁移率改变分析（EMSA）检测各组细胞NF-κB的DNA结合活性。多组间样本均数比较采用单因素方差分析，组间两样本均数比较采用LSD检验。结果联合用药组的caspase3、caspase9和p21基因表达（1．903±0．122、2．726±0．050、2．541土0．393）与蛋白表达（0．534±0．077、1．161±0．172、0．463±0．016）高于对照组（1．001±0．052、1．000±0．014、1．001±0．037和0．080±0．043、0．248±0．059、0．139±0．021）、金雀异黄素组（1．559±0．038、2．394±0．095、1．686士0．061和0．335±0．052、0．478±0．059、0．304±0．018）和5-FU组（1．198±0．063、1．051±0．043、1．399±0．055和0．194±0．015、0．337±0．036、0．231±0．011），联合用药组的生存素基因与蛋白表达（0．165±0．018和0．216±0．014）低于对照组、金雀异黄素组和5-氟尿嘧啶单组（1．001±0．033、0．775±0．044、0．395±0．030和0．594±0．079、0．375±0．014、0．295±0．014），差异均有统计学意义（基因表达，F-802．865、52．760、39．992、187．288、37．435；蛋白表达，F-10．466、44．483、19．490、200．011、45．238；P均〈0．01），caspase3、p21在两单药组的表达与对照组相比，差异也有统计学意义（LSD检验，P〈0．05）。EMSA检测显示，与对照组（1067．97±36．01）和5-FU组（718．Objective To investigate the molecular mechanism how genistein increased the sensitivity of colon cancer cell to 5-fluorouracil（FU） and promoted colon cancer cell apoptosis. Methods SW480 colon cancer cell line was chosen as experimental object. Genistein 80 μmol/L and 5-FU 30 μg/mL used separated or combined for 48 hours were set as drug experiment group. There were four experiment groups in this study： control group （without any drug）, 5-FU group, genistein group, 5-FU and genistein combined group. The expression of survivin, Bcl-2, p21, caspase3 and caspase 9 in the tumor cells of each group at mRNA and protein level was deteced by real-time quantitative polymerase chain reaction （PCR） and Western blotting. The relative expression quantity of genes was determined by comparative threshold method （2^-ΔΔCT） and β-actin was taken as an internal reference. Semi quantitative analysis was performed for protein relative expression quantity. The DNA combination activity of nuclear factor（NF） -κB of each group was measured by electrophoretic mobility shift assay （EMSA）. Single factor analysis of variance was used for mean comparison among multiple groups and LSD test was for mean comparison between two groups. Results The expression of caspase3, caspase 9 and p21 of 5-FU and genistein combination group at mRNA （1. 903±0. 122, 2. 726±0. 050 and 2. 541±0.39a） and protein level （0. 534±0. 077, 1. 161±0. 172 and 0. 463±0. 016） were all higher than those of control group （1. 001±0. 052, 1. 000±0. 014 and 1. 001±0. 037;0. 080±0. 043, 0. 248±0. 059 and 0. 139±0. 021）, genistein group （1. 559 ± 0. 038, 2. 394±0. 095 and 1. 586±0,051; 0. 335±0.052, 0. 478±0. 059 and 0. 304±0. 018） and 5-FU group （1. 198±0.063, 1.051±0.043 and 1. 399±0. 055; 0.194±0.015, 0. 337±0.036 and 0. 231±0. 011）; the expression of survivin of 5-FU and genistein combined group at mRNA and protein level （0. 165 ± 0. 018 and 0. 215±0. 014） were all lower than those of control gr

关 键 词：金雀异黄素 氟尿嘧啶 结肠肿瘤 凋亡 机制 

分 类 号：R735.35[医药卫生—肿瘤]