The Role And Mechanism Of Activated Bile Acid Receptor TGR5 In Esophageal Squamous Carcinoma Cells

Objective: This study explores the role and mechanism of activating TGR5 in esophageal squamous cell carcinoma.Methods:(I)Analysis of esophageal cancer and TGR5 according to TCGA database.(II)Role of activation of TGR5 in human esophageal squamous cell carcinoma: 1.By culturing esophageal squamous cell carcinoma cells,RNA was extracted and the expression of TGR5 gene in different esophageal cancer cell lines was detected using qRTPCR technology;2.The effect of activation of TGR5 on proliferation of esophageal squamous cell carcinoma by MTT assay;3.Cell scratch test to detect the effect of activated TGR5 on the migration of esophageal squamous cell carcinoma;4.Using q-RTPCR to detect the effects of TGR5 activation on the proliferation and migration-related genes of esophageal squamous cell carcinoma;5.The effect of activating TGR5 on the apoptosis of esophageal squamous carcinoma cells was detected by flow cytometry.Results:(I)Analysis of esophageal cancer and TGR5 according to TCGA database: 1.There is a clear correlation between the stage of esophageal cancer and the survival rate of patients.As the stage progresses,the survival rate of patients declines significantly;2.TGR5 is widely expressed in various tissues of the normal body and is highly expressed in the esophagus;3.TGR5 is expressed in normal esophagus and cancer tissues,but the expression of TGR5 in cancer tissues is generally lower than that in normal tissues.Therefore,TGR5 can be used as a potential target in the treatment of esophageal cancer.(II)Activation of TGR5 in human esophageal squamous cell carcinoma: 1.TGR5 expression in different esophageal cancer cell lines: TGR5 is expressed in different esophageal cancer cell lines,and the expression in KYSE150 cell line is higher than in EC109 cell line;2.Activation of TGR5 can inhibit the proliferation of esophageal squamous carcinoma cells: according to the results of MTT experiments,the proliferation of esophageal squamous cell carcinoma cells was inhibited after treatment with different concentrations of INT777.The higher the concentration,the more effective the inhibition is;3.Activation of TGR5 can inhibit the migration of human esophageal cancer cell lines: the results of cell scratch experiments showed that compared with the control group,after adding INT777 to KYSE150 and EC109 cell lines to activate TGR5,the cell scratch healing ability was significantly reduced;4.Effect of TGR5 on gene expression of esophageal cancer cells:(1)q-RTPCR results show that after 100 μM INT777 treatment,in KYSE150 cells,the pro-inflammatory and oncogenes TNF-α,IL-6,IP-10,and the migration-promoting genes MMP1 and MMP12 were significantly inhibited,and the differences were statistically significant(P <0.05);(2)The q-RTPCR results of EC109 cells showed that the proinflammatory and oncogenes TNF-α and IP-10 in EC109 cells were significantly inhibited,and the difference was statistically significant(P <0.05).However,there were no significant differences in other inflammatory cancers and migrationrelated genes;5.The effect of TGR5 on the apoptosis of esophageal cancer cells was detected by flow cytometry.The results showed that there was no significant difference between the esophageal cancer cells treated with INT777 at 24 h and 48 h compared with the negative control group.Therefore,TGR5 had no effect on the apoptosis of esophageal cancer cells at 24 h and 48 h.The molecular mechanism that activates TGR5 to inhibit tumor proliferation and migration is being further explored.Conclusion: 1.TGR5 is widely expressed in various tissues,and its expression level in esophageal cancer tissues is generally lower than that in normal esophageal tissues;2.Activation of TGR5 can inhibit the proliferation of human esophageal squamous cell carcinoma KYSE150,EC109 and TE-1;3.Activation of TGR5 can inhibit the migration of human esophageal squamous carcinoma cells KYSE150 and EC109;4.Activation of TGR5 can inhibit the expression of proinflammatory and pro-oncogenes TNF-α,IL-6,IP-10,etc.,and the migration-promoting genes MMP1,MMP12 in esophageal cancer cells.