The Role And Mechanism Of Rock2 Regulating IFITM1 In Osteosarcoma Cell Proliferation

Background and Objective:Osteosarcoma(osteosarcoma,OS)is the most common primary malignant bone tumor,rapid progression,poor prognosis.Osteosarcoma is easy to occur early transfer owing to the high degree of malignancy.At present,the accurate treatment of osteosarcoma is to find an effective target for prevention and treatment.Rhoassociated kinase 2(Rock2)is a key signal molecule of Rho / ROCK signaling pathway,and many research reveals that Rock2 has an important role in osteosarcoma proliferation.And Interferon induced transmembrane protein 1(IFITM1)has been shown highly expressed in a variety of malignant tumors and associated with tumor proliferation.Our previous studies found that the reduction of Rock2 protein could cause IFITM1 decreased expression in osteosarcoma cells by c DNA microarray.And osteosarcoma is one of great ability proliferation tumor,so we speculate Rock2 regulating IFITM1 expression affecting tumor proliferation ability in osteosarcoma.In this study we will firstly investigate the impact on the biological activity of Rock2 regulating IFITM1 on osteosarcoma proliferation ability in vivo and vitro.Then further to observe the molecular biology mechanism of Rock2 regulating IFITM1.Including detecting the expression Rock2 and IFITM1 in osteosarcoma tissues and cells,and separately analyzing their association with survival time.Methods: 1,to detect Rock2 and IFITM1 expression levels of mRNA and protein in multiple strains of osteosarcoma cells by qRT-PCR and Western blot.2,When reduced Rock2 expressing in U2-OS and MG-63,to detect IFITM1 mRNA and protein expression using qRT-PCR and Western blot,while taking advantage of CCK8 and EDU experiment to observe capacity of cell proliferation changes.3,used stably transfected sh Rock2-MG-63 cell line to construct osteosarcoma tumor model in subcutaneous of nude mice,and observe tumor growth.4,In osteosarcoma cells steady both silence Rock2 and simultaneously overexpress IFITM1 or both overexpress Rock2 and reduce IFITM1,to observe osteosarcoma cell proliferation by the CCK8 and EDU experiment.5,confirmed that Rock2 regulate expression of IFITM1 by p GSK-3?/?-catenin pathway by western blot and qRT-PCR.6,detected Rock2 and IFITM1 expression levels in osteosarcoma tissue of 62 cases by immunohistochemistry,and analyze the correlation with survival time by survival curves.Result:1,qRT-PCR and Western blot results showed that mRNA and protein expression levels of Rock2 and IFITM1 elevated in multiple strains of osteosarcoma cell lines.2,mRNA and protein levels of IFITM1 decreased after transfected sh Rock2 or added Rock2 inhibitor(Y27632)in U2-OS and MG-63,while cell proliferation was significantly decreased(p <0.05).Furthermore,in vivo experiments,we confirmed that stable low expression Rock2 group U2-OS cells formed tumors,and tumor volume and weight were significantly reduced for the control group(p <0.05).In addition,we further found that elevated IFITM1 can reverse the situation that tumor proliferation decreased by silencing Rock2,while reducing IFITM1 can suppress the proliferation of osteosarcoma that was enhanced by overexpressing Rock2.3,We confirmed Rock2 regulated the expression of IFITM1 by p GSK-3? / ?-catenin pathway in osteosarcoma using qRT-PCR and Western blot.4,Immunohistochemistry showed,Rock2 expression in osteosarcoma was higher than normal bone tissue(p <0.001),and IFITM1 was also higher than normal bone tissue(p <0.05).Rock2 and IFITM1 expression levels was negatively correlated with patient survival.Conclusion:Rock2 adjusted IFITM1 by GSK-3? / ?-catenin pathway and further affect osteosarcoma cell proliferation.Our studys for the study of osteosarcoma proliferation mechanism provided a new theoretical basis and will provide a new direction for the prevention and treatment targets of osteosarcoma.