豆根管食通口服液对食管鳞癌细胞miR-377的影响研究

目的观察豆根管食通口服液对食管鳞癌细胞株KYSE450的细胞增殖、细胞凋亡及对miR-377、CD133和VEGF表达的影响。方法先制备豆根管食通口服液含药血清,然后分为高中低三个浓度,分别作用24 h、48 h、72 h,筛选出最佳浓度和最佳作用时间,然后将KYSE450细胞株分为空白组、豆根管食通中剂量组、氟尿嘧啶组。采用MTT方法检测豆根管食通口服液含药血清对KYSE450细胞增殖的影响,采用流式细胞仪检测豆根管食通口服液含药血清对KYSE450细胞凋亡的影响,采用RT-PCR方法检测豆根管食通口服液含药血清对KYSE450细胞miR-377的影响,采用WB方法检测豆根管食通口服液含药血清对KYSE450细胞CD133和VEGF表达的影响。结果筛选出最佳浓度为豆根管食通中剂量组,最佳作用时间为48 h;MTT结果显示,豆根管食通口服液对KYSE450细胞能够明显抑制细胞增殖,豆根管食通口服液含药血清中剂量组的细胞凋亡率为(18.8±2.66)%,高于空白组(P <0.01),促进miR-377表达,与空白对照组相比(P <0.05),抑制CD133表达(P <0.05),抑制VEGF表达(P <0.01)。结论豆根管食通口服液对食管鳞癌KYSE450细胞株有抗增殖、促凋亡的作用,其机制可能是通过上调miR-377的表达,进而抑制CD133和VEGF表达。

Effect of Dougen Guangshitong Oral Liquid on miR-377 Expression in Esophageal Squamous Carcinoma Cells

Objective To observe the effect of Dougen Guanshitong Oral Liquid on the proliferation and apoptosis of esophageal squamous cell line KYSE450 and on the expression of miR-377,CD133 and vascular endothelial growth factor(VEGF)proteins in the cell line KYSE450.Methods The drug serum of Dougen Guanshitong Oral Liquid was prepared firstly,and then the KYSE450 cells were co-cultured with high-,middle-,and lowconcentration drug serum for 24,48,and 72 hours respectively to screen out the optimal concentration and the optimal medication time.MTT method was used to detect the effect on the proliferation of KYSE450 cells,flow cytometry was used to detect the effect on the apoptosis of KYSE450 cells,RT-PCR method was used to detect the effect on the miR-377 expression in KYSE450 cells,and Western blotting method was used to detect the effect on CD133 and VEGF expression in KYSE450 cells of the blank group,middle-dose Dougen Guanshitong Oral Liquid group and fluorouracil group.Results The optimal concentration was the middle dosage of Dougen Guanshitong Oral Liquid,and the optimal medication time was 48 hours.MTT results showed that Dougen Guanshitong Oral Liquid could significantly inhibit the proliferation of KYSE450 cells,the apoptotic rate of the middle-dose group was(18.8±2.66)%(P<0.01)as compared with the blank group.Dougen Guanshitong Oral Liquid could also significantly promote the expression of miR-377(P<0.05)compared with the blank group,and inhibit the expression of CD133(P<0.05)and the expression of VEGF(P<0.01).Conclusion Dougen Guanshitong Oral Liquid has certain effect on inhibiting the proliferation and promoting the apoptosis of esophageal squamous carcinoma KYSE450 cells.The mechanism may be related with the up-regulation of the expression of miR-377 and with the inhibition of the expression of CD133 and VEGF.