Objective To observe the effects of total flavonoids of Oxytropis falcata ( FOF) on apoptosis and Toll-like receptor 4 ( TLR4) /MyD88/NF-κB pathway in the gastric cancer cells. Methods The proliferation of gastric cancer cell lines SGC7901, MKN-49 P, AGS, MKN45 and KATOIII was detected by MTT assay. The SGC7901 cells with the lowest IC50 were taken as the experimental cells. The concentration of FOF was 10-40 μg/m L and the duration of action was 48 h.The SGC7901 cells in the logarithmic phase were divided into 6 groups. The cells in the LPS group were treated with 2 μg/m L TLR4 activator LPS. The cells in the ER + LPS group were first treated with 100 nmol/L TLR4 inhibitor Eritoran for 2h and then were added with 2 μg/m L LPS. The cells in the LFOF, MFOF, and HFOF + LPS groups were first treated with10, 20, and 40 μg/m L FOF for 2 h, followed by 2 μg/m L LPS, and the cells in the blank group were added with the same amount of solvent. After 48-hour treatment, the apoptosis rate was measured by flow cytometry. The apoptosis-related proteins ( Bax, Bcl-2, Survivin) and TLR4/MyD88/NF-κB pathway-related proteins ( TLR4, p-TLR4, MyD88, p-MyD88, p65, and p-p65) were detected by Western blotting. The cell immunofluorescence was used to observe the nuclear importation of p65 protein. Results Compared with the blank group, the apoptosis rate of SGC7901 cells in the LPS group decreased, the expression of Bax protein decreased, the expression of Bcl-2, Survivin, p-TLR4, p-MyD88 and p-p65 protein increased, and the amount of p65 increased ( all P < 0. 05) . Compared with LPS group, the apoptosis rate of SGC7901 cells in the ER + LPS group, LFOF + LPS group, MFOF + LPS group, and HFOF + LPS group increased, Bax protein expression increased, the expression of Bcl-2, Survivin, p-TLR4, p-MyD88, and p-p65 proteins decreased, and the amount of p65 decreased ( all P < 0. 05) . Conclusion FOF can induce apoptosis of gastric cancer SGC7901 cells, which is related to the inhibition of TLR4/MyD88/NF-κB pathway activity.%目的 观察镰形棘豆总黄酮 (FOF) 对胃癌细胞凋亡及Toll样受体4 (TLR4) /MyD88/NF-κB通路的影响.方法 MTT法检测不同浓度FOF作用下胃癌细胞株SGC7901、MKN-49P、AGS、MKN45、KATOⅢ的增殖情况, 确定以IC50最小的SGC7901细胞作为受试细胞, FOF作用浓度为10～40μg/m L、作用时间为48 h.将对数生长期的SGC7901细胞分为6组, 脂多糖 (LPS) 组加入2μg/m L的TLR4激活剂LPS, ER+LPS组先加入100 nmol/L TLR4抑制剂Eritoran处理2 h后再加入2μg/m L LPS, LFOF、MFOF、HFOF+LPS组先分别加入10、20、40μg/m L FOF处理2 h后再加入2μg/m L LPS, 空白组加入等量溶媒.处理48 h后, 用流式细胞仪测算细胞凋亡率, Western blotting法检测细胞中的细胞凋亡相关蛋白 (Bax、Bcl-2、Survivin) 、TLR4/MyD88/NF-κB通路相关蛋白 (TLR4、p-TLR4、MyD88、p-MyD88、p65、p-p65) , 细胞免疫荧光法观察细胞p65蛋白进核情况.结果 与空白组比较, LPS组SGC7901细胞凋亡率下降, Bax蛋白表达降低, Bcl-2、Survivin、p-TLR4、p-MyD88及p-p65蛋白表达上升, p65进核量增加 (P均<0. 05) ;与LPS组比较, ER+LPS组、LFOF+LPS组、MFOF+LPS组和HFOF+LPS组SGC7901细胞凋亡率上升, Bax蛋白表达升高, Bcl-2、Survivin、p-TLR4、p-MyD88及p-p65蛋白表达降低, p65进核量减少 (P均<0. 05) .结论 FOF可诱导胃癌SGC7901细胞凋亡, 该作用与其抑制TLR4/MyD88/NF-κB通路活性有关.
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