Research On The Effects And Mechanism Of CD147 And CAS In Testicular Germ Cell Tumors

Objective:To investigate the expression level and localization of CD147 and CAS in different types of testicular germ cell tumors,and to study the effects of two kinds of molecules on the biological behaviors of testicular germ cell tumorsr cells in cell proliferation,cell migration and invasion at the cellular level,as well as the molecular mechanism of their effects so as to provide a basis for the early diagnosis and the treatment of testicular germ cell tumors.Methods:(1)The expression levels and localization of CD147 and CAS in human testicular tissues were detected by immunohistochemical SP method,and the expression levels and localization of the two molecules were compared.(2)The expression of CD147 and CAS in the testicular germ cell tumors cell line was silenced by adeno-associated virus(AAV)in the experimental group,and the empty vector was transfected in the control group.The silencing effect of AAV on CD147 and CAS in NT-2 cells was verified by PCR,qPCR and western blot at mRNA and protein levels.(3)Cell viability assay kit was used to detect the difference in cell viability between the control group and the experimental group.(4)The effects of CD147 and CAS on the cell migration and invasion abilities of NT-2 cells were evaluated by Transwell cell migration and invasion experiments.(5)Western blotting technique was used to detect the changes of signal molecules related to cell division and proliferation in the experimental group and the control group.Results:(1)The immunohistochemical staining results showed that CD147 was not expressed in normal testicular tissue,but was expressed on the cell membrane to varying degrees in different types of TGCTs.In seminoma,yolk sac tumor and teratoma,CD147 showed low to high expression.By analyzing and comparing the expression levels of CD147 in normal testicular tissues and different types of TGCTs,it was found that the expression levels of CD147 in different types of TGCTs were significantly higher than those in normal testicular tissues.In seminoma,the expression level of CD147 in T1N0M0 wassignificantly lower than that of the seminoma tissues in T2N0M0 and T4N0M0(p=0.0173 and p=0.0467).There was no significant difference in CD147 between T2N0M0 stage and T4N0M0 stage in seminoma tissues.There was also no significant difference in CD147 expression between seminoma and non-seminoma(2)The location of CAS was different in normal testicular tissues and different types of testicular germ cell tumors.In normal testicular tissues,CAS was mainly located in the nucleus of spermatozoa,but it was not found in mature sperm.In seminoma and embryonal carcinoma,CAS was mainly located in cytoplasm.However,CAS was located in cytoplasm and nucleus in teratoma.While in yolk sac tumor,CAS was mainly expressed in nucleus.There was no significant difference in CAS expression between normal testicular tissues and seminoma(p>0.9999).In normal testicular tissues and seminoma,the expression level of CAS was significantly higher than that in non-seminoma.(3)The results of PCR,qPCR and western blot showed that the knockdown effects of CD147 and CAS in AAV-mediated NT-2 cells were significant.(4)The cell viability assay showed that the proliferation ability of NT-2 cells was significantly decreased by silencing the expressions of CD147 and CAS.(5)Cell migration and invasion experiment showed that silencing the expression of CD147 and CAS in NT-2 cells had no effect on the migration and invasion ability of testicular cancer cells.(6)The results of Western blotting technique showed that the expression of TRAF2,a molecule regulating the NF-κB signaling pathway,was down-regulated after CD147 knockdown in NT-2 cells in the experimental group compared with the control group.The expression level of classical NF-κB signaling pathway molecule p100 did not change,and the expression levels of p50 and p65 were down-regulated.The expression level of non-classical NF-κB signaling pathway molecule RelB was significantly decreased.(7)The expression of anti-apoptotic factors Bcl-2 and BCL-xl were down-regulated after the knockdown of CAS in NT-2 cells.The expression level of cell cycle regulating molecule p53 did not change significantly.In the NF-κB signaling pathway,the expression levels of classical NF-κB signaling molecule p65 and non-classical NF-κB signaling molecule RelB were significantly decreased.Conclusion:(1)CD147 was expressed in different level in TGCTs,but was not found in normal testicular tissues.(2)In seminoma,the expression of CD147 in T2N0M0 and T4N0M0 grades was higher than that in T1N0M0 grades.(3)In normal testicular tissues,CAS was mainly located in spermatocyte nucleus.In seminoma and embryonal carcinoma,CAS wasmainly located in cytoplasm.(4)CD147 and CAS could promote the growth and proliferation of testicular germ cell tumor cells through the NF-κB signaling pathway.(5)CD147 and CAS had no effect on the migration and invasion ability of testicular germ cell tumor cells.