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目的体外研究选择性环氧合酶-2(COX-2)抑制剂尼美舒利和5-氟尿嘧啶(5-FU)对胃癌细胞的抑制作用及机制。方法以胃癌细胞株MKN45、MKN28为研究对象,观察尼美舒利和5-FU单独或联合应用对细胞增殖、凋亡和细胞周期的影响。应用MTT法检测细胞增殖,流式细胞仪检测细胞凋亡(FITC-Annexin-V/PI双标记)和细胞周期,RT-PCR观察用药前后COX-2mRNA在两株细胞中的表达,Western免疫印迹法观察经两种药物单独和联合作用48h后细胞内凋亡相关蛋白Bax和Bcl-2的表达。结果在MKN45和MKN28细胞中均可观察到不同水平的COX-2mRNA表达,尼美舒利和5-FU联合应用可明显抑制COX-2mRNA表达。尼美舒利可抑制两株细胞的增殖并诱导凋亡。尼美舒利和5-FU具有协同抑制细胞增殖及诱导凋亡的作用,该作用与两种药物作用顺序无关,但在联用时作用最强。两药协同抑制增殖的作用主要通过协同杀伤和诱导凋亡而实现。5-FU增强了凋亡诱导蛋白Bax的表达,而尼美舒利则减少凋亡抑制蛋白Bcl-2的表达。两药联用可明显抑制胃癌细胞株生长。结论选择性环氧合酶-2抑制剂尼美舒利和5-FU通过抑制COX-2mRNA的表达及增强Bax/Bcl-2的表达比率诱导胃癌细胞凋亡,从而对胃癌细胞起到协同抑制增殖的作用。
Objective To investigate the inhibitory effect and mechanism of nimesulide and 5-fluorouracil (5-FU), a selective cyclooxygenase-2 (COX-2) inhibitor, on gastric cancer cells in vitro. Methods The gastric cancer cell lines MKN45 and MKN28 were used as research objects to observe the effects of nimesulide and 5-FU alone or in combination on cell proliferation, apoptosis and cell cycle. Cell proliferation was detected by MTT assay. Cell apoptosis was detected by flow cytometry (FITC-Annexin-V / PI double labeling) and cell cycle. The expression of COX-2 mRNA in both cell lines was detected by RT-PCR. Western blotting We observed the expression of Bax and Bcl-2 in the cells after 48h treatment by both drugs alone and in combination. Results Different levels of COX-2 mRNA expression were observed in both MKN45 and MKN28 cells. The combination of nimesulide and 5-FU significantly inhibited the expression of COX-2 mRNA. Nimesulide can inhibit both cell proliferation and induce apoptosis. Nimesulide and 5-FU have a synergistic effect in inhibiting cell proliferation and inducing apoptosis. The effect is independent of the order of action of the two drugs, but is most effective when combined with 5-FU. Synergistic inhibition of proliferation of two drugs mainly through synergistic killing and induction of apoptosis. 5-FU increased the expression of apoptosis-inducing protein Bax, while nimesulide decreased the expression of Bcl-2. Combination of two drugs can significantly inhibit the growth of gastric cancer cell lines. Conclusions Nimesulide and 5-FU, a selective cyclooxygenase-2 inhibitor, induce apoptosis of gastric cancer cells by inhibiting the expression of COX-2 mRNA and enhancing the ratio of Bax / Bcl-2 expression, thereby synergistically inhibiting the proliferation of gastric cancer cells Role.