Genes Regulated by Nkx2-3 in Sporadic and Inflammatory Bowel Disease-Associated Colorectal Cancer Cell Lines

Nkx2-3 has been reported to be up-regulated in B cell lines and intestinal tissues from Crohn's disease patients and down-regulated in colorectal cancer. The purpose of the current study is to determine genes regulated by Nkx2-3 in sporadic (CRS61) and inflammatory bowel disease-associated (CRS4) colorectal cancer cell lines. Small interfering RNA-mediated knockdown of Nkx2-3 in both cell lines was generated and high-density cDNA microarrays representing over 25,000 genes were performed. Microarray results were validated by RT-PCR and immunofluorescence. Pathway analysis was used to identify gene networks associated with Nkx2-3 knockdown in these cell lines. A total of 1,677 genes were regulated by Nkx2-3 in CRS4 cells; 1,375 genes were regulated by Nkx2-3 in CRS61 cells. Among those genes regulated by Nkx2-3, 254 genes were similarly regulated by Nkx2-3 knockdown in both cell lines; 159 genes were differentially regulated by Nkx2-3 knockdown between the two lines. Genes regulated by Nkx2-3 were grouped primarily within the following two functional categories: (1) immune and inflammatory response; and (2) cell proliferation, growth, and oncogenesis. Among the genes with similarly changed expression in the two cell lines, the top affected pathways included antigen presentation and cell-cell signaling. Among the genes with differentially changed expression between the two cell lines, ingenuity pathway analysis indicated that the top affected pathway included genes directly involved in Wnt signaling. Nkx2-3 may contribute to the pathogenesis of IBD-associated CRC and sporadic CRC by regulating the Wnt signaling pathway.