Expression And Biological Function Of IRF2BP2 In Colorectal Cancer

Objective:To investigate the expression and biological function of Interferon regulatory factor 2binding protein 2 in colorectal Interferon regulatory factors.Methods:(1)The expression of IRF2BP2 in colorectal cancer cells was analyzed by searching the TCGA and GTEx databases.(2)142 pairs of cancer and paracancer tissue specimens from colorectal cancer patients were collected,tissue microarrays were prepared,and the expression of IRF2BP2 protein in the tissues was detected using immunohistochemistry.The immunohistochemical results were correlated with the clinicopathological data to analyze the relationship between the expression of IRF2BP2 and age,gender,pathological stage,tumor size,depth of infiltration,differentiation,and lymph node metastasis.(3)Extracted total protein and total RNA from six types of colorectal cancer cells,SW480,SW620,LoVo,HCT116,HT29 and SW116,and normal colorectal mucosal epithelial cells,NCM460,and detected the expression of IRF2BP2 protein and RNA in the above cells using protein blotting assay and reverse transcription quantitative PCR.(4)The IRF2BP2 knockdown SW480,SW620,and LoVo colorectal cancer cell lines were transfected with IRF2BP2 shRNA lentiviral particles and IRF2BP2 scramble shRNA Ientiviral particles,respectively,to construct IRF2BP2 knockdown SW480,SW620,and LoVo cell lines,respectively,SW480-shIRF2BP2 and its control SW480-shIRF2BP2(negative control),SW620-shIRF2BP2 and its control SW620-shnc,and LoVo-shIRF2BP2 and its control LoVo-shnc;SW480,a relatively low IRF2BP2-expressing intestinal cancer cell,was further transfected with IRF2BP2 overexpressing viral particles and IRF2BP2 overexpression negative control viral particles to construct the IRF2BP2 gene overexpression SW480 cell line SW480-IRF2BP2 and its control SW480-mock,and to observe the transfection efficiency qualitatively by fluorescence microscopy and quantitatively by flow cytometry,and to detect the transfection efficiency by Western blot and RT-qCR to further detect the efficiency of IRF2BP2 overexpression and knockdown.(5)Cell function experiments:CCK-8 cell proliferation assay,plate cell colony formation assay and Transwell assay were used to detect the effect of IRF2BP2 expression on the growth,proliferation and migration ability of colorectal cancer cells.Results:(1)The expression of IRF2BP2 was significantly higher in colorectal cancer tissues compared with normal tissues.(2)Immunohistochemical results showed that the expression of IRF2BP2 protein in colorectal cancer tissues was significantly higher than that in normal mucosal epithelial tissues adjacent to the cancer.Correlation with clinicopathology suggested that the expression of IRF2BP2 was related to the degree of differentiation and lymph node metastasis,but not to age,gender,pathological stage,tumor size and depth of infiltration.(3)In contrast to normal colonic mucosal epithelial cells NCM460,IRF2BP2 expression of protein and RNA was elevated in colorectal cancer cell lines.(4)The viral transfection efficiency was detected by fluorescence microscopy and flow cytometry,and the results showed that the fluorescence rate was high,which proved that the viral transfection was successful;meanwhile,the Western blot and RT-qPCR results indicated that the expression of IRF2BP2 protein and RNA was significantly reduced in the knockdown group compared with the negative control group,and significantly increased in the overexpression group compared with the negative control group,which means that the IRF2BP2 knockdown SW480,SW620 and LoVo intestinal cancer cell lines and IRF2BP2 overexpression SW480 intestinal cancer cell lines were successfully constructed.(5)The proliferation,cloning and migration ability of colorectal cancer cells were reduced after IRF2BP2 knockdown;the opposite result was observed for IRF2BP2 overexpression.Conlusions:(1)IRF2BP2 expression was increased in colorectal cancer,and the expression of IRF2BP2 was associated with the degree of differentiation and lymph node metastasis,but not with age,gender,pathological stage,tumor size and depth of infiltration.(2)Expression of IRF2BP2 promotes the growth,proliferation and migration ability of colorectal cancer cells.(3)IRF2BP2 may be a target for colorectal cancer.