Relationship Between DDX39 And Prognosis Of Colorectal Cancer And Its Roles In The Polarization Of Macrophage

Background: Colorectal cancer is a common malignant tumor with high risk of digestive system.The incidence rate of colorectal cancer is second in malignant tumors in females,and it is third in malignant tumors in the males.The growth of colorectal cancer cells depends on the characteristics of the tumor itself and the surrounding microenvironment.The largest number of cells in tumor microenvironment are macrophages.Therefore,the effect of macrophages on the growth of tumor cells is the current research hotspot.Macrophage includes 2 subtypes,namely M1 type and M2 type,M1 macrophages release cytokines and promote immune response to inhibit tumor cell proliferation and adjacent tissue damage.Type M2 macrophages release cytokines that promote the proliferation and repair of adjacent cells,and inhibit the immune response and promote tumor growth.DDX39 is a Dead-Box protein,is a kind of ATP dependent RNA helicase.It is found that the expression level of DDX39 in tumor epithelium has an effect on the growth of tumor cells.However,there is no reach about the expression of DDX39 in tumor stroma,especially the macrophages.In this study,we investigated the effect of DDX39 expression on the prognosis of colorectal carcinoma and the roles of DDX39 on the polarization of macrophages。Objective: To investigate the relationship between the expression of DDX39 protein and prognosis of colorectal cancer,analysis the relationship between the expression of DDX39 in mesenchyme and the number of interstitial cells.To observe the expression of DDX39 in macrophages and its effects on the polarization direction of macrophages.Methods:1.The relationship between the expression of DDX39 protein in colorectal carcinoma and the prognosis of patientsAccording to the results of immunohistochemical staining of paraffin sections of 824 patients with colorectal cancer,combined with the clinical pathological parameters and prognosis information,the effect of DDX39 protein expression on the prognosis of colorectal cancer was investigated.2.Relationship between expression of DDX39 and macrophage in colorectal cancerDDX39 and CD163,CD66 b,POSTN,FOXP3 protein expression were observed in the stroma of 824 cases of colorectal cancer,select the most closely related protein immunofluorescence Co localization.3.The effect of DDX39 on macrophages polarizationTHP-1 cell lines were induced into macrophages by PMA 200ng/ml,48 h and IFN 20ng/ml were added into LPS,and then polarized into M1 type macrophages.After adding IL-4 20ng/ml and IL-13 20ng/ml,polarized M2 macrophages.PCR method was used to detect the expression of TNF-,IL-6,IL-1,CD163,and DDX39 in different subtypes of macrophages.To reduce the expression of M0 in type DDX39 macrophages,observe the changes of TNF-,IL-6,IL-1,and CD163,and determine the change of the polarization direction of macrophages.Results:1.The expression of DDX39 protein was detected in the cytoplasm and nucleus of epithelium and stroma by immunohistochemistry.Statistical analysis showed that the expression of DDX39 in epithelial and stromal tissues of colorectal cancer was significantly different from that in normal,para carcinoma and adenoma.The expression and location of tumor,DDX39 protein in epithelial differentiation,TNM,serum CEA,serum CA199,tumor recurrence and metastasis were statistically significant(P < 0.05);statistically significant expression of DDX39 protein in the stroma in tumor location,TNM staging,serum CEA,serum CA199,tumor recurrence and metastasis(P < 0.05).The epithelial cells in the low expression of DDX39 high expression group overall survival rate and disease-free survival rate,poor prognosis,the difference was statistically significant(P=0.000);interstitial DDX39 in low expression group was higher than group total survival rate and disease-free survival rate,poor prognosis,the difference was statistically significant(P=0.000).Cox single factor analysis showed that the overall survival in patients,tumor differentiation,serum CEA,DDX39 expression and epithelial mesenchymal DDX39 expression were prognostic factors in colorectal cancer;Cox multivariate analysis showed that tumor differentiation,expression of DDX39 and epithelial mesenchymal DDX39 expression level were independent factors affecting the prognosis of colorectal cancer.Cox single factor disease free survival analysis showed that the degree of tumor differentiation,TNM staging,serum CEA,DDX39 expression and epithelial mesenchymal DDX39 expression were prognostic factors in colorectal cancer;Cox multivariate analysis showed that tumor differentiation,expression of DDX39 and epithelial mesenchymal DDX39 expression level were independent factors the prognosis of colorectal cancer.2.CD163 markers were selected M2 macrophages,CD66 b labeled neutrophils,POSTN labeled fibroblasts,FOXP3 labeled T cells by immunohistochemistry.To analysis of correlation between the expression of DDX39 in the stroma and the protein.We found the interstitial DDX39 and the expression of CD163 was most closely related,Spearman coefficient r=-0.627,a significant negative correlation.Immunofluorescence staining of paraffin sections of 20 cases of colorectal cancer showed that DDX39 was co localized with F4/80,indicating that macrophages expressed DDX39.However,DDX39 and CD163 immunofluorescence did not co localization,the expression of CD163 in the stroma was not DDX39,but the expression of DDX39 in high expression region was less than that of CD163.3.It was found by PCR that the expression of CD163 was higher than that of type M1 macrophages,while the expression of DDX39 was decreased,indicating that M2 might be a marker of type M1 macrophages.When the DDX39 of M0 macrophages was knocked out,the CD163 increased,indicating that the trend of macrophages to M2 type polarization.Conclusion:1.The expression of DDX39 protein was independent prognostic factors of colorectal cancer.2.The expression of DDX39 in the stroma was most closely related to CD163 and negatively correlated with the protein level.3.DDX39 and CD163 were negatively correlated with RNA level,and the decrease of DDX39 might promote the polarization of M2.