Circ＿0004674 Mechanisms Regulating Glycolysis And Biological Behavior Of Osteosarcoma Via The MiR-140-3p/TCF4 Pathway

Background/Objective: Circ RNAs,as a subclass of non-coding RNAs,play an important role in the development of tumours.Circ＿0004674 is one of the many Circ RNAs,which is closely related to the formation and progression of oral squamous cell carcinoma and cervical cancer.However,the role of Circ＿0004674 in osteosarcoma is poorly understood,especially with regard to the mechanisms of glycolysis.Therefore,the role of Circ＿0004674 in glycolysis and biological behavior of osteosarcoma and its potential regulatory mechanism were investigated in this paper.Methods:(1)The expressions of circ＿0004674,miR-140-3p and TCF4 in 9osteosarcoma tissues(without preoperative chemotherapy or radiotherapy)and 9paracancer normal tissues,osteoblasts h FOB1.19 and MG63,U2 osteosarcoma cell lines were detected by real-time quantitative PCR The difference.(2)The effects of circ＿0004674 knockdown and down-regulation of miR-140-3p on proliferation,apoptosis,migration and invasion of osteosarcoma cells were detected by CCK8,flow cytometry,wound healing assay and Transwell method.(3)The effects of circ＿0004674 knockdown and down-regulation of miR-140-3p on glucose consumption,lactate production and glycolytic related protein expression in osteosarcoma cells were detected by glucose uptake assay,lactate production assay and Western blotting assay(Wb).(4)Prediction of possible downstream target genes TCF4 of circ＿0004674 and miR-140-3p in osteosarcoma using miRWalk,miRDB,Target Scan and miRTar Base bioinformatics databases,and verification of direct binding between circ＿0004674 and miR-140-3p and TCF4 by dual luciferase reporter analysis assay.(5)The role of circ＿0004674/miR-140-3p/TCF4 signalling axis on malignant biological behaviours such as glycolysis and proliferation of osteosarcoma cells was examined by functional rescue assays such as CCK8.Results: The expression level of circ＿0004674 was increased in osteosarcoma tissues and cell lines compared to normal tissues adjacent to cancer and human osteoblasts h FOB1.19.circ＿0004674 knockdown inhibited proliferation,invasion and migration of osteosarcoma cells,induced apoptosis and showed a significant weaken effect on glycolytic processes.miR-140-3p is lowly expressed in osteosarcoma tissues and cell lines.miR-140-3p downregulation reverses the effects of circ＿0004674knockdown on the malignant biological behavior of osteosarcoma cells such as glycolysis and proliferation.Dual luciferase reporter assays confirm that binding between circ＿0004674 and miR-140-3p and downstream target genes is direct.TCF4 was overexpressed in osteosarcoma tissues and cell lines,while miR-140-3p was negatively correlated with the gene expression levels of both circ＿0004674 and TCF4.Overexpression of TCF4 reversed the effects of circ＿0004674 knockdown on glycolysis and biological behavior of osteosarcoma cells.Conclusions: circ＿0004674 knockdown can inhibit the proliferation,migration and invasion of osteosarcoma cells,promote apoptosis and weaken the glycolysis process of osteosarcoma.TCF4 is the target gene of miR-140-3p.circ＿0004674promotes glycolysis and biological behavioral process of osteosarcoma through miR-140-3p/TCF4 pathway,which is expected to be a new therapeutic target for osteosarcoma.