Discussion on the mechanism of growth inhibition of Liujunzi Decoction on EC9706 based on AKT/mTOR pathway
Objective:To explore the mechanism of Liujunzi Decoction on the growth inhibition of esophageal carcinoma cell EC9706.Methods:MTT assay was used to detect the inhibitory effect of Liujunzi Decoction on EC9706 cells,flow cytometry was used to detect apoptosis and cycle distribution in each group,Seahorse energy metabolism analysis system was used to detect glycolysis rate and mitochondrial pressure of cells in each group,qRT-PCR was used to detect the relative expression of mammalian target of rapamycin(mTOR)and protein kinase Bl(AKT1)mRNA in each group of cells,Western Blot was used to detect the relative expression of p-mTOR,mTOR and AKT1/2/3 protein in each group.Results:Liujunzi Decoction could inhibit the activity of EC9706 cells when the concentration was 200,300,400 μg/mL(P＜0.01);Liujunzi Decoction 150 μg/mL and 200 μg/mL were selected for follow-up experiments.Flow cytometry showed that Liujunzi Decoction could significantly promote the apoptosis of EC9706 cells(P＜0.0l).Liujunzi Decoction 200 μg/mL could prolong the S phase and shorten the G2/M phase(P＜0.05).The results of glycolysis rate test showed that Liujunzi Decoction 200 μg/mL could inhibit basic glycolysis,compensate glycolysis and glycolysis potential of EC9706 cells(P＜0.05).Mitochondrial stress test results showed that Liujunzi Decoction 150 μg/mL inhibited ATP synthesis oxygen consumption,basal respiratory value and non-mitochondrial oxygen consumption in EC9706 cells(P＜0.05,P＜0.01).Liujunzi Decoction 200 μg/mL inhibited ATP synthesis oxygen consumption,mitochondrial maximum oxygen consumption capacity,basic respiratory value,non-mitochondrial oxygen consumption and reserve respiratory capacity of EC9706 cells(P＜0.01).qRT-PCR results showed that Liujunzi Decoction 150,200 μg/mL significantly decreased the relative expression of mTOR mRNA in EC9706 cells(P＜0.05,P＜0.01),Liujunzi Decoction 200 μg/mL significantly decreased the mRNA relative expression of AKT1 in EC9706 cells(P＜0.01).Western Blot results showed that Liujunzi Decoction 50,200 μg/mL significantly inhibited the relative expression of p-mTOR and AKT1/2/3 protein(P＜0.05,P＜0.01),and Liujunzi Decoction 200 μg/mL significantly inhibited the relative expression of mTOR protein(P＜0.01).Conclusion:Liujunzi Decoction can promote apoptosis of EC9706 cells,induce cell cycle arrest,inhibit energy metabolism and cell growth by interfering with AKT/mTOR pathway.
Esophageal cancerLiujunzi DecoctionEC9706Protein kinase B(AKT)Mammalian target of rapamycin(mTOR)
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