健脾和胃法通过IL-6/STAT3信号通路对食管癌细胞株EC9706生长的干预作用

Intervention effect of invigorating the spleen and regulating stomach method on growth of EC9706 via IL-6/STAT3 signaling pathway

目的:通过观察健脾和胃法代表方六君子汤对EC9706细胞增殖以及细胞中IL-6/STAT3通路相关因子和蛋白表达的影响,探讨健脾和胃法及其代表方六君子汤治疗食管癌的分子机制。方法:用MTT法检测六君子汤乙酸乙酯提取物、Stattic、IL-6对EC9706细胞生长的影响;实验设空白组、六君子汤组、Stattic抑制剂组、六君子汤+Stattic抑制剂组、IL-6组、六君子汤+IL-6组、Stattic抑制剂组+IL-6组、六君子汤+Stattic抑制剂+IL-6组,空白组用RPMI 1640培养基,六君子汤组用六君子汤乙酸乙酯提取物和Stattic浓度均为其IC35,所有含IL-6组IL-6加入时间为两药孵育6h后加入,浓度为20ng/mL。利用Western Blot检测六君子汤乙酸乙酯提取物对细胞中STAT3和p-STAT3蛋白表达的影响。结果:六君子汤乙酸乙酯提取物、Stattic可以抑制EC9706细胞的增殖;IL-6可以促进细胞增殖,但无明显量效关系。六君子汤乙酸乙酯/Stattic孵育6h然后加入IL-6培养基,各实验组OD值与空白组比较均显著降低(P<0.05),表现为抑制细胞增殖。六君子汤乙酸乙酯提取物和Stattic及其联合用药可降低p-STAT3蛋白表达(P<0.05)及p-STAT3蛋白占STAT3蛋白比例(P<0.05)。结论:六君子汤乙酸乙酯部位可以干预IL-6/STAT3信号通路抑制EC9706细胞的增殖。

Objective: To explore the molecular mechanism of invigorating the spleen and regulating stomach method and its representative formula Liujunzi Decoction in the treatment of esophageal carcinoma, through the proliferation of EC9706 cells, and STAT3 pathway related factors and proteins of EC9706 cells. Methods: The effects of ethyl acetate extract of Liujunzi Decoction, Stattic, and IL-6 on the growth of EC9706 cells was detected by MTT method. The cells were divided into control group, Liujunzi Decoction group, Stattic inhibitor group, Liujunzi Decoction+Stattic inhibitor group, IL-6 group, Liujunzi Decoction group+IL-6, Stattic inhibitor+IL-6 group, Liujunzi Decoction+Stattic inhibitor+IL-6 group. The blank control group was cultured in RMPI 1640 medium, and the concentration of ethyl acetate extract of Liujunzi Decoction and Stattic were their IC35, and the two drugs incubated for 6 hours and then added IL-6(the concentration was 20 ng/mL). The effects on the expression of STAT3 and p-STAT3 in EC9706 cells of ethyl acetate extract of Liujunzi Decoction was detected with Western Blot. Results:The ethyl acetate extract of the Liujunzi Decoction and Stattic could inhibit the proliferation of EC9706;IL-6 could promote the proliferation of EC9706 cells, which had no significantly dose-effect relationship. Compared with control group, the OD value of experimental groups was significantly lower(P<0.05), when adding IL-6 after the ethyl acetate extract of Liujunzi Decoction and Stattic were incubated for 6 hours, which was shown to inhibit of cell proliferation. The ethyl acetate extract of the Liujunzi Decoction and Stattic and their combined use could down-regulate the expression of the p-STAT3(P<0.05) and the ratio of p-STAT3 in STAT3 in EC9706 cells(P<0.05). Conclusion: The ethyl acetate extract of the Liujunzi Decoction can inhibit the proliferation of EC9706 cells by interfering the IL-6/STAT3 signaling pathway.