常春藤皂苷元与5-氟尿嘧啶或奥沙利铂联用对人结肠癌HT-29细胞增殖的影响

Effect of Hederagenin and 5-fluorouracil/oxaliplatin on the Proliferation of Human Colon Adenocarcinoma Cell Line HT-29

目的观察常春藤皂苷元(HG)与5-氟尿嘧啶(5-FU)或奥沙利铂(Oxa)联用对人结肠癌HT-29细胞增殖的抑制作用及可能作用机制。方法采用四甲基偶氮唑蓝(MTT)法测定HG、5-FU、Oxa单药及联合作用于指数增殖期HT-29细胞的效应;应用中效原理进行统计分析,绘制单药及联合用药对肿瘤细胞生长的影响曲线,确定联合用药时的药物效应-合用指数的关系,判定药物间的相互作用;集落形成实验检测HG、5-FU、Oxa单药及联合用药对集落形成的影响;采用RT-q PCR方法检测HG对5-FU疗效相关基因乳清酸磷酸核糖转移酶(OPRT)、胸苷酸合成酶(TS)mRNA表达及Oxa疗效相关基因切除修复交叉互补基因(ERCC1)和乳腺癌易感基因(BRCA1)mRNA表达的影响。结果随着药物浓度增加,HG、5-FU、Oxa单药或联合用药对HT-29生长的抑制效应也增加,高浓度联合应用效应增强。HG联合5-FU或Oxa作用于HT-29细胞株可产生较好的协同效应,HG、5-FU、Oxa、HG+5-FU、HG+Oxa各组的克隆抑制率分别为39.34%、45.37%、51.01%、70.43%、70.96%,与对照组比较差异有统计学意义(P<0.01)。HG作用后TS mRNA下调至0.554,ERCC1 mRNA下调至0.182,BRCA1mRNA下调至0.044,OPRT mRNA上调至1.655,与对照组比较差异均有统计学意义(P<0.05或P<0.01)。结论 HG联合5-FU或Oxa用药对人结肠癌HT-29细胞有显著降低细胞增殖能力,增强化疗药物的疗效作用。下调HT-29细胞的TS、ERCC1、BRCA1基因表达水平,上调OPRT基因表达水平,可能是其协同作用机制之一。

Objective To observe the effect of concomitant administration of hederagenin（ HG） and 5-fluorouracil（ 5-FU） / oxaliplatin（ Oxa） on the proliferation of human colon adenocarcinoma cell line HT19 and its possible mechanism. Methods Using MTT,the single and concomitant effect of HG,5-FU and Oxa on HT-29 during exponential phase of proliferation were assayed. By performing statistical analysis such as Median-effect Equation,the response curves are plotted and the drug-drug interaction by establishing the drug combination-response model predicted. By colony formation assay,we have measured the effect of single and concomitant usage of the 3 drugs on the colony formation. Using RT-qRCP,the effects on mRNA expressions of OPRT,TS,ERCC1 and BRCA1 were tested.Result As the dose increased,the inhibiting effect of HG,5-FU and Oxa on HT-29 proliferation,singly or concomitantly administered,also increased with the synergistic effect in high concentration enhanced. The effect of concomitant administration of HG and 5-FU / Oxa produced favorable synergistic effect. The inhibitory rate for HG,5-FU,Oxa,HG ＋ 5-FU and HG ＋ Oxa groups were 39. 34%,45. 37%,51. 01%,70. 43% and 70. 96%,respectively,and their differences to control group showed statical significance（ P 〈0. 01）. After the administration of HG,TS mRNA was down-regulated to 1. 655,ERCC1 mRNA down-regulated to 0. 182,BRCA1 mRNA down-regulated to0. 44 and OPPRT mRNA up-regulated to 1. 655 with statistical significances to control group（ P 〈0. 05 or P 〈0. 01）.Conclusion the concomitant administration of hederagenin（ HG） and 5-fluorouracil（ 5-FU） / oxaliplatin（ Oxa）showed significant inhibitory effect on the proliferation of human colon adenocarcinoma cell line HT19 with the efficacy concomitantly enhanced. The one of the synergistic mechanisms is possibly the down-regulation of HT-29 in TS,ERCC1 and BRCA1 expressions and up-regulation of OPRT expression.