扶正解毒祛瘀方联合奥沙利铂对人结肠癌HT-29细胞增殖与凋亡的影响及机制研究

摘要

目的:研究扶正解毒祛瘀方(简称"扶正方")联合奥沙利铂(L-OHP)对人结肠癌HT-29细胞增殖与凋亡的影响及机制.方法:将HT-29细胞分为空白对照组(不加药物)、扶正方组(1000 mg/L)、L-OHP组(31.25 mg/L)和联合用药组(1000 mg/L扶正方+31.25 mg/L L-OHP).加入相应药物作用48 h后,采用MTT法检测细胞增殖情况,倒置显微镜下观察细胞形态的改变,流式细胞术检测细胞周期和凋亡率,实时荧光定量聚合酶链式反应(qRT-PCR)法检测细胞中促凋亡基因Bax、抑凋亡基因Bcl-2 mRNA的表达,Western blot法检测细胞中Bax、Bcl-2蛋白的表达.结果:与空白对照组比较,L-OHP组和联合用药组细胞增殖均受到抑制、S期和G2/M期细胞比例升高、G0/G1期细胞比例降低(P<0.05),L-OHP组、扶正方组和联合用药组细胞凋亡率升高、细胞中Bax mRNA及蛋白的表达上调、细胞中Bcl-2 mRNA的表达下调(P<0.05),且联合用组变化较两药单用组更明显(P<0.05).结论:扶正方与L-OHP联合应用可抑制HT-29细胞的增殖、促进细胞凋亡,且作用优于两药单用;其机制可能与上调细胞中Bax基因与蛋白表达、下调细胞中Bcl-2基因表达有关.%OBJECTIVE:To study the effect and its mechanism of Fuzheng Jiedu Quyu formula(short for"Fuzheng formula") combined with oxaliplatin (L-OHP) on human colon cancer HT-29 cell proliferation and apoptosis. METHODS:HT-29 cells were divided into blank control group(without drugs),Fuzheng formula group(1000 mg/L),L-OHP group(31.25 mg/L)and combi-nation group (1000 mg/L Fuzheng formula+31.25 mg/L L-OHP). After cultured with corresponding drug for 48 h,MTT method was used to detect the cell proliferation;the changes of cellular morphology were observed by invert microscope;flow cytometry was used to detect the cell cycle and apoptosis rate. Proapoptotic gene Bax,apoptotic gene Bcl-2 mRNA expressions were deter-mined by real-time fluorescence quantitative polymerase chain reaction method;Bax,Bcl-2 protein expressions were assayed by Western blot. RESULTS:Compared with blank control group,cell proliferation was inhibited in L-OHP group and combination group;cell proportion was increased in S stage,G2/M stage and decreased in G0/G1 stage(P<0.05). Cell apoptosis rate in L-OHP group,Fuzheng formula group and combination group was increased;Bax mRNA and protein expression were up-regulated,Bcl-2 mRNA expression was downregulated(P<0.05);and combination group changed more obviously than the single drug groups(P<0.05). CONCLUSIONS:Fuzheng formula combined with L-OHP can inhibit HT-29 cell proliferation and promote its apoptosis, showing better effects than either of the two drugs alone. The mechanism may be associated with up-regulation of Bax gene and pro-tein expressions and down-regulation of Bcl-2 gene expressions in cells.