Expression Of TRAIL In Esophageal Squamous Cell Carcinoma Promotes Tumor Progression

Background and objectives Esophageal cancer is one of the common malignant gastrointestinal tumors with extremely high morbidity and mortality.In the world,70% of the cases occur in China,of which 90% are esophageal squamous cell carcinoma(ESCC).Currently,the difficulty of esophageal cancer treatment lies in the late detection and recurrence of metastases.Therefore,finding new effective treatment strategies has become the focus of esophageal cancer research today.Tumor Necrosis Factor-Related Apoptosis Inducing Ligand(TRAIL)is a member of the tumor necrosis factor superfamily(TNFSF),encoded by the gene TNFSF10.As a type II transmembrane,extracellular region of TRAIL can be cleaved by proteases to form a trimer,which exists as a soluble protein.There are five TRAIL receptors: four membrane-bound receptors,TRAIL-R1,TRAIL-R2,and TRAIL-R3,TRAIL-R4,and a soluble receptor,osteoprotegerin(OPG).These receptors can be divided into two types in function: DR4 and DR5 are death receptors because they contain death domains(DD)in the cytoplasm and can transmit apoptosis signals.The other three play a role of "decoy".Dc R1 and Dc R2 have high homology with the extracellular regions of DR4 and DR5,but due to the lack of a functional intracellular domain,they cannot transmit apoptosis signals.For OPG,it is low affinity with TRAIL at physiological temperatures.TRAIL can specifically induce the apoptosis of malignant tumor cells,but it is not toxic to normal cells,so it was used for tumor treatment at the beginning of its discovery.However,due to the acute toxicity caused by TRAIL treatment,the clinical use of TRAIL has been reviewed.And TRAIL can activate NF-k B,PI3 K,JNK,p38 and other signaling pathways in colon cancer,non-small cell lung cancer and other tumors to promote tumor proliferation,invasion and metastasis,and can also affect the secretion of CCL2,CXCL8,CXCL1 by tumor cells to change the tumor microenvironment.However,the impact of TRAIL on the progression of esophageal cancer is rare.In this project,we mainly studied the expression of TRAIL in the tumor tissue and normal tissue of patients with esophageal squamous cell carcinoma in the TCGA database,analyzed the relationship between the expression and the clinical parameters of patients,and verified that in the clinical specimens of patients with esophageal squamous cell carcinoma.The effects of TRAIL expression on the biological function and chemokine secretion of esophageal squamous carcinoma cell lines were also explored.This study may provide a novel treatment for immunotherapy.Methods 1.Extract total RNA from tissues and cells by RNAiso Plus,and detect RNA concentration and purity by Nano Drop.2.Detect the expression of TRAIL in paraffin sections of tumor tissues of ESCC by Immunohistochemical.Covered with neutral gum,dried and photographed and scored under using microscope.3.Gene silencing of TRAIL was achieved by si RNA.4.Human recombinant TRAIL was used to treat ESCC cell lines KYSE70 and KYSE150 in vitro.5.Use ultra-low adsorption plates and pelleting medium to test the pelleting ability of tumor cells.6.CCK-8 kit was used to detect the proliferation ability of tumor cells.7.Transwell plate was used to detect tumor cell migration ability and immune cell chemotactic ability.8.Western blot was used to detect the expression of E-cadherin in vitro.9.Use ELISA kit to detect CXCL10 secretion.10.Real-time quantitative PCR was used to detect the m RNA levels.11.Flow cytometry was used to detect the expression of stemness genes on the tumor cell surface and the transfection efficiency of transfected cells in vitro.Results 1.TRAIL is highly expressed in tumor tissues of patients with ESCC,and has a negative correlation with the patient's prognosis and a positive correlation with the patient's disease stage and lymphatic metastasis.2.In ESCC cell lines KYSE70 and KYSE150,TRAIL promoted tumor cell proliferation,spheroidization and migration,up-regulated the expression of stem genes,and affected the expression of EMT-related protein E-cadherin.3.TRAIL inhibited the secretion of CXCL10 in ESCC cell lines KYSE70 and KYSE150.4.In ESCC tumor cell lines KYSE70 and KYSE150,TRAIL reduces the chemotaxis of CD8+ T cells to tumor cells by inhibiting the secretion of CXCL10.Conclusion TRAIL is highly expressed in ESCC tumor tissues and TRAIL expression is positively correlated with lymph node metastasis and disease stage,and negatively correlated with patient prognosis.TRAIL can promote the ability of proliferation,spheroidization and migration in ESCC tumor cells.Inhibiting the expression of TRAIL in tumor cells up-regulates the secretion of tumor cells CXCL10 and therefore increases the chemotaxis of CD8+ T cells to tumor cells.Overexpression of TRAIL can down-regulate the secretion of tumor cells CXCL10 and inhibit the chemotaxis of CD8+ T cells to tumor cells.