Effect of Qigesan on Proliferation， Apoptosis and miR-133a/Akt/mTOR Signaling Pathway of Esophageal Cancer EC9706 Cells / 中国实验方剂学杂志

ABSTRACT

Objective:To observe the effect of Qigesan on the proliferation and apoptosis of the human esophageal cancer cell EC9706， and the effect on miR-133a/protein kinase B（Akt）/mammalian target of rapamycin （mTOR） signaling pathway. Method:The effective constituent of Qigesan was extracted by ethyl acetate. Thiazolyl blue tetrazolium bromide（MTT） colorimetric assay was used to determine the dosage of Qigesan on cells and to detect the effect of Qigesan on the proliferation of EC9706 cells. The effect of Qigesan on apoptosis of EC9706 cells was detected by flow cytometry. The effect of Qigesan on miR-133a and insulin-like growth factor 1 receptor（IGF-1R） mRNA expression was detected by Real-time quantitative polymerase chain reaction （Real-time PCR） . The protein expression of Akt and mTOR in EC9706 cells was detected by Western blot. Result:Qigesan can inhibit the proliferation of EC9706 cells in a dose-dependent manner（<italic>P</italic><0.01）. Inhibitory concentrations 30% inhibition concentration（IC₃₀） 40 mg·L^-1 and median inhibition concentration（IC₅₀） 80 mg·L^-1 were selected for follow-up experiments. Compared with the blank group， both the inhibitor group and the combination drug group can inhibit the proliferation of EC9706 cells （<italic>P</italic><0.01）. The inhibitor at 0.25 μmol·L^-1 was selected for subsequent experiments. Compared with the blank group， Qigesan 80 mg·L^-1 dose group could significantly promote the late apoptosis rate and total apoptosis rate of EC9706 cells（<italic>P</italic><0.05）， and the 40 mg·L^-1 dose group could significantly promote the late apoptosis rate of EC9706 cells（<italic>P</italic><0.05）， which shows synergistic effect after concomitant use with Akt/mTOR inhibitor（<italic>P</italic><0.05）. Compared with the blank control group， each group can effectively increase expression of miR-133a（<italic>P</italic><0.05）. The combination of inhibitor and traditional Chinese medicine（TCM） has obvious promotion effect. Compared with blank control group， the expressions of Akt and mTOR were significantly decreased in each group（<italic>P</italic><0.05）. Compared with single medication， the expressions of Akt and mTOR were decreased in combination of inhibitor and TCM group. Conclusion:Qigesan can inhibit the growth of EC9706 cells and promote apoptosis， and its inhibitory mechanism may be related to the Akt/mTOR signaling pathway by regulating the expression of miR-133a.