Analysis The Correlations Of Dc-sign/ DC-SIGNR With EBV Infection And Susceptibility Of Nasopharyngeal Carcinoma

Objective: The aim of this study is to explore the interaction between DC-SIGN/DC-SIGNR and Epstein-Barr virus( EBV) infection by constructing EBV infection different cell types models and observing the expression ofDC-SIGN/DC-SIGNR in cells after EBV infection. To further investigate the relations between genetic polymorphism of DC-SIGN/DC-SIGNR and susceptibility to nasopharyngeal carcinoma(NPC),we perform a ease-control study, and thereby giving a deeper insight into the role of EBV infection in the mechanism of NPC occurrence and improve the effectiveness of its early diagnosis.Methods: 1. Monocytes and lymphocytes were isolated from peripheral blood mononuclear cells from healthy volunteers and developed by stimulation with cytokine in vitro. Peripheral blood mononuclear-derived monocytes,lymphocytes,KMH2 cells and nasopharyngeal carcinoma cells were directly infected by EBV which were prepared by AGS EBV(+) cell line. At the same time four nasopharyngeal carcinoma epithelial cell lines were co-cultured with KMH2 EBV(+) cells respectively to constructing EBV infection nasopharyngeal carcinoma cell model.2. The expression of DC-SIGN/DC-SIGNR gene in different cell types which had been infected by EBV were detected by Real-time Quantitative-PCR(RT-qPCR) and In-cell Western.3. The competitive inhibitor Mannan of DC-SIGN/DC-SIGNR was used in competitive inhibition of EBV infection experiment: monocytes were blocked by mannan for one hour before EBV infection. And then EBV infection related molecules in infected cells were detected by RT-qPCR.4. The mRNA expression level of DC-SIGN/DC-SIGNR gene under nasopharyngeal mucosal tissue in nasopharyngeal carcinoma patients and chronicity blennymenitis of pars nasalis pharynges patients were detected by RT-qPCR.5. Technologies of Sequenom MassArray, PCR, agarose gel electrophoresis and DNA sequencing were used to analyze the genetic polymorphism of DC-SIGN and DC-SIGNR. Chi-square test and multivariate unconditional logistic regression analysis were used to analyze the associations between the polymorphism of DC-SIGN/DC-SIGNR and susceptibility of nasopharyngeal carcinoma.Results: 1. Peripheral blood mononuclear-derived monocytes and lymphocytes, KMH2 cells were able to be directly infected by EBV. And the EBV infection of monocytes was inhibited by mannan. The expression of DC-SIGN was down-regulated after EBV infected monocytes, while the expression of DC-SIGNR was up-regulated. At the same time the expression of DC-SIGN and DC-SIGNR were all up-regulated after EBV infected lymphocytes and KMH2 cells.2. The efficiency of EBV particles directly infected nasopharyngeal carcinoma cells was low. Nasopharyngeal carcinoma cells could be infected by EBV after co-cultured with KMH2 EBV (+) cells. And the expression of DC-SIGNR were up-regulated after EBV infected nasopharyngeal carcinoma cells.3.There were no significant differences of mRNA expression level of DC-SIGN and DC-SIGNR under nasopharyngeal mucosal tissue between nasopharyngeal carcinoma patients and chronicity blennymenitis patients(P>0.05).4. The differences of genotypes/alleles distribution of DC-SIGN's neck domain found in Chinese health control from Guangxi Province and Caucasians weren't statistical significance, as well as in health control and nasopharyngeal carcinoma patients(P>0.05). The differences of genotypes distribution and alleles frequency of DC-SIGNR's neck domain between Chinese health controls from Guangxi Province and Caucasian population were all extremely distinct(P<0.05). The differences of genotypes distribution of DC-SIGNR's neck domain found in Chinese NPC patients from Guangxi Province and health controls weren't statistical significance (P>0.05), but alleles distribution frequency were extremely distinct(P<0.05). And the 9-repeat DC-SIGNR's neck domain allele was more frequently found in patients with nasopharyngeal carcinoma compared to health controls (P<0.05).5. The differences of genotypes distribution of DC-SIGN rs7252229 between Chinese NPC patients from Guangxi Province and health controls were statistically significant (P<0.05). Genotypes GC and GG at DC-SIGN rs7252229 were associated with a decreased risk of NPC with adjusted Odds Ratios (ORs)of 0.076(95%CI:0.008-0.690,P=0.022) and 0.056 (95%CI:0.006-0.487,P=0.009)respectively.Conclusions: 1 .As the expression of DC-SIGN and DC-SIGNR were changed in monocytes, lymphocytes, KMH2 cells and NPC cells after EBV infection. And the changes of DC-SIGN/DC-SIGNR expression in different cell types were significantly different, so we propose that DC-SIGN and DC-SIGNR may play an important role for EBV infection cells and the roles would be changed follow with cell types.2.The ways of EBV infected epithelial cells were more likely to cell-to-cell contact rather than direct infection.3.The polymorphism of DC-SIGNR' s neck domain maybe associate with the susceptibility of NPC.4. DC-SIGN rs7252229 polymorphism may influence susceptibility to NPC.