二十二碳六烯酸联合5氟尿嘧啶对胃癌细胞生长及bcl-2、bcl 2112和bax表达的影响

The effect of docosahexaenoic acid and 5-fluorouracil on human gastric cancer cell lines and the expression of bcl-2, bcl 2112 and bax

目的评估二十二碳六烯酸（DHA）联合5氟尿嘧啶（5-Fu）对胃癌细胞SGC 7901生长以及bcl-2、bcl 2112和bax基因表达的影响。方法采用台盼蓝拒染方法检测两药物单用和联用对细胞活力的影响，用联合系数判断两药合用效果，倒置显微镜下观察细胞生长状况，PI染色流式细胞术检测亚二倍体峰比并拟合细胞周期曲线，Annexin-V／PI双标记方法检测早期细胞凋亡，RT—PCR方法检测bcl-2、bcl 2112和bax基因的表达。结果DHA可抑制胃癌SGC 7901细胞生长，且呈剂量和时间依赖性（P〈0．05），24h、48h的半数抑制浓度分别为67．81μg／ml、45．76μg／ml；DHA联合5-Fu对细胞生长的抑制具有协同作用（CI〈1，P〈0．01），倒置显微镜下可见两药联合处理后细胞稀疏；亚二倍体峰比、Annexin—V早期凋亡率示DHA、5-FU均能诱导细胞凋亡且联合用药后细胞凋亡更明显（DHA、5、FU、联合组的亚二倍体峰比分别为5．2％、6．2％、13．9％；早期细胞凋亡率分别为4．00％、5．37％、13．11％）；细胞周期曲线在联合组停滞于G0／G1和S期；RT—PCR显示DHA、5-FU可下调bcl-2和bcl 2112表达，两药联合表达时下调更显著，bax表达则无明显改变。结论DHA能抑制胃癌SGC7901细胞增殖，联合5-FU后对细胞生长抑制和周期阻滞具有协同作用，可能通过下调bcl-2和bcl 2112基因诱导胃癌SGC 7901细胞发生凋亡。

Objective To evaluate the growth inhibition of human gastric carcinoma cell lines SGC 7901 in vitro and the expression of bcl-2, bcl 2112 and bax with docosahexaenoic acid （DHA） and 5-fluorouracil （5-FU）. Methods The effect of DHA and 5-FU was measured by trypan blue, and the interaction between two agents was judged by combination index （ CI ）. Cells were observed by inverted microscope. Flow cytometry was used for analysis of apoptosis by PI staining and Annexin-V/PI. RT-PCR was used to analyze the levels of bcl-2, bcl 2112 and bax mRNA. Results DHA significantly inhibited the growth of SGC 7901 cells in a dose- and time-dependent way （ P 〈 0. 05 ）, the IC50 of 24 h and 48 h was 67.81μg/ml and 45.76 μg/ml, and a strong synergism was found in the combination of DHA and 5-FU （CI 〈 1 ,P 〈0. 01 ）. Treated by DHA and 5-FU for 48 h, cells became sparse under inverted microscope. DHA or 5-FU was able to induce apoptosis and the effect became even more significant by the combination of DHA and 5-FU. Cells were holted in phase of G0/G1 and S. RT-PCR showed that DHA or 5-FU downregulated the expression of bcl-2 and bcl 2112 mRNA, while bax mRNA expression was not downregulated. Conclusions DHA could inhibit the growth of gastric carcinoma cells, DHA and 5-FU had synergetic effect in the inhibition of the cells growth and blockage of the cell cycles possibly by down-regulating the expression of bcl-2 and bcl 2112.