| Background and purposeColon cancer(COAD)is a common malignant tumor in the digestive system.Its high morbidity and mortality make it one of the main causes of cancer death in the world.Studies have shown that tumor microenvironment plays an important role in the development of cancer.Therefore,we studied the microenvironment of colon cancer in order to find new diagnostic markers and immunotherapy targets for colon cancer.MethodsThe Estimate algorithm was used to obtain the tumor purity of each colon cancer sample in the TCGA database.And then we analyzed the relationship between each component in the tumor microenvironment with the prognosis and clinicopathological characteristics of the patient.After that,117 common differentially expressed genes(DEGs)were obtained by comparing the gene expression profiles of DEGs between tumor and normal samples and the high and low immune score groups.Then we analyzed the function,molecular pathway and prognostic value of 117 DEGs.The expression of DNASE 1L3 in colon cancer was verified by multiple online websites or databases.CIBERSORT algorithm was used to obtain the immune cell content of each colon cancer sample,and differential expression analysis and correlation test were used to screen the immune cells associated with DNASE1L3 expression level.In the experimental section,real-time PCR was applied for confirming the expression of DNASE1L3 in normal samples of colon cancer and adjacent cancer.The correlation between DNASE1L3 and EMT related markers was detected in SW480、SW48、HT-29、HCT-116 and SW620 after overexpression of DNASE1L3.The protein expression of DNASE1L3 in the above 5 cells was detected by western blot,and 2 cells(SW480 and SW48)with the lowest expression were selected for subsequent scratch experimentand cell proliferation experiment.Results1.There were significant differences between Immune score in tumor microenvironment with stage(P=0.011)and the M classification in TMN(P=0.001)in colon cancer patients.So we chose the Immune score for subsequent analysis.2.A total of 117 genes were obtained from the intersection of the DEGs associated with the immune score of colon cancer and the DEGs in colon cancer and normal samples.The survival analysis was carried out in 117 DEGs,and 4 statistically significant genes were obtained,which were GPX3,DNASE 1L3,SLC11A1 and PRELP,the target gene DNASE 1L3 was finally determined.3.The results of single-gene GSEA showed that the gene function and enrichment pathway of high DNASE1L3 expression group were mainly related to immunity.4.Immune cell correlation analysis results showed that there were 10 immune cells that were related to the expression level of DNASE 1L3.Among them,it is most related to macrophages M0.5.Compared with normal tissues adjacent to cancer,DNASE1L3 was low expression in colon cancer and negative correlated with patient prognosis.The results of cell experiment showed that DNASE 1L3 had no obvious correlation with EMT.It can inhibite the migration.but there was no significant statistical difference in inhibition of cell proliferation.ConclusionDNASE 1L3 may be a marker related to the prognosis and immune infiltration in colon cancer,or it may provide guidance for the follow-up search of the diagnosis and treatment targets in colon cancer. |
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