The Expression And Carcinogenic Mechanism Of GLRX3 In Nasopharyngeal Carcinoma

BackgroundGLRX3(Glutaredoxin 3)is an oxidoreductase which is a member of the glutaredoxin family which reduces a variety of substrates using glutathione as a cofactor.It was overexpressed in breast cancer,colon cancer and lung cancer.Furthermore,our previous work showed that the transcription level of GLRX3 in both nasopharyngeal carcinoma(NPC)cell lines and NPC tissues was significantly higher than that in the normal nasopharyngeal epithelial cell line NP69 and non-cancerous nasopharynx tissues,respectively;and in vitro studies showed that knockdown of GLRX3 gene could inhibit the growth,migration and colony formation of HONE 1 cells.The aforementioned results suggested that GLRX3 might be an oncogene in NPC.PurposeThe current study aimed at further exploring the expression and function of GLRX3 gene in NPC,by performing experiments in vitro and in vivo experiments.We first tried to establish a cell line,named CNE2-KD GLRX3(GLRX3-KD),in which GLRX3 was knocked down in CNE2.Secondly,we tried to study the biological function of GLRX3 in NPC cells.Furthermore,the expression of protein GLRX3 in NPC,adjacent normal mucosa and chronic inflammation of nasopharyngeal mucosa tissues was carried out,and the relationship between the expression of GLRX3 and clinical parameters was analyzed.Thus,it is possible to investigate the potential role of GLRX3 in the occurrence and development of NPC,and to verify the hypothetical feasibility of GLRX3 gene as an oncogene of NPC.Methods1.Retroviral vectors targeted GLRX3 genes were successfully constructed and then transfected into PA317,collected the virus liquid to infection in NPC cell line CNE2,which has a high expression of GLRX3,with RNA interference(RNAi)technology.The stably transfected cell line targeting GLRX3 gene was established for the subsequent work.A series of functional experiments were performed to investigate the biological features of GLRX3 gene,including growth inhibition experiments,colony formation assay,wound-healing assay and invasion assay,as well as in vivo xenograft experiments in nude mice.2.The paraffin blocks of 59 cases of NPC and 46 cases of their adjacent normal nasopharyngeal epithelium tissues,and 30 cases of chronic inflammation of nasopharyngeal mucosa tissues were collected from the Department of Pathology of the First Affiliated Hospital of GuangXi Medical University.All samples were confirmed by pathological diagnosis.The clinic data of these cases was collected as well.3.Immunohistochemistry(IHC)was applied to detect the expression of GLRX3 in all the above samples.The localization of the positive signals and the expression level were compared among the aforementioned three groups of samples.The correlation between GLRX3 and the clinicopathological character was also analyzed,to explore the function of GLRX3 gene in the occurrence and development of NPC.Results1.The GLRX3-silenced cell line,GLRX3-KD-3 and GLRX3-KD-4,was successfully constructed in CNE2.Compared to the control CNE2,GLRX3-KD-3 cells showed a significantly reduced capacity in clone formation,proliferation,migration,invasion,and tumor growth in nude mice.2.IHC results showed that GLRX3 protein located in the cytoplasm.The expression of GLRX3 protein in NPC was significantly higher than that in adjacent mucosa(P<0.05),but no significant relationship with gender,age,clinical stage and lymphonode metastasis(P>0.05).Conclusions1.GLRX3 is overexpressed in NPC tissues.2.GLRX3 gene silence can inhibit the malignant behavior of the NPC cells.3.GLRX3 is a candidate cancer gene in NPC.