Using miRNA microarray analysis, we identified 31 miRNAs that were significantly up-regulated or down-regulated in colon cancer tissues. We chose MIR196B, which was specifically up-regulated in colon cancer, for further study. We identified 18 putative MIR196B target genes by comparing between the mRNAs down-regulated in MIR196B-overexpressed cells and the assumed MIR196B target genes predicted by public bioinformatics tools. The association between MIR196B and FAS was verified in this study. FAS expression was constitutively elevated in normal human colorectal tissues. However, its expression was often reduced in human colorectal cancer. The decrease in FAS expression could be responsible for the reduction of apoptosis in colorectal cancer cells. In colorectal cancer tissue, we showed that MIR196B up-regulation was mutually followed by down regulation of FAS expression. We also showed that MIR196B directly repressed FAS expression in colorectal cells. Furthermore, anti-MIR196B up-regulated FAS expression and increased apoptosis in colorectal cancer cell lines. Our results suggest that the up-regulation of MIR196B modulates apoptosis in colorectal cancer cells by partially repressing FAS expression and that anti-MIR196B could be a potential candidate as an anti-cancer drug in colorectal cancer therapy.

MiRNAs are non-coding RNAs that play important roles in the pathogenesis of human diseases by regulating target gene expression in specific cells or tissues. We aimed to detect miRNAs related to ulcerative colitis [UC], identify their target molecules, and analyse the correlation between the miRNAs and their target genes in colorectal cells and dextran sulphate sodium [DSS]-induced mouse colitis. UC-associated miRNAs were identified by miRNA microarray analysis using DSS-induced colitis and normal colon tissues. The results were validated by quantitative real-time polymerase chain reaction [RT-PCR]. We identified target genes of MIR429, a colitis-associated miRNA, from our screen by comparing the mRNA microarray analysis in MIR429-overexpressed cells with predicted candidate target genes. We constructed luciferase reporter plasmids to confirm the effect of MIR429 on target gene expression. The protein expression of the target genes was measured by western blot,enzyme-linked immunosorbent assay [ELISA] analysis, or immunohistochemistry. We identified 37 DSS-induced colitis associated miRNAs. We investigated MIR429 that is down-regulated in DSS-induced colitis, and identified 41 target genes of MIR429. We show that the myristoylated alanine-rich protein kinase C substrate [MARCKS] is a direct target of MIR429. MARCKS mRNA and protein expression levels are down-regulated by MIR429, and MIR429 regulates the expression of MARCKS and MARCKS-mediated mucin secretion in colorectal cells and DSS-induced colitis. In addition, anti-MIR429 up-regulates MARCKS expression in colorectal cell lines. Our findings suggest that MIR429 modulates mucin secretion in human colorectal cells and mouse colitis tissues by up-regulating of MARCKS expression, thereby making MIR429 a candidate for anti-colitis therapy in human UC.

The human microRNA 452 (MIR452) was identified as a colorectal cancer (CRC)-associated micro RNA (miRNA) by miRNA expression profiling of human CRC tissues versus normal colorectal tissues. It was significantly up-regulated in human CRC tissues. However, the functional mechanisms of MIR452 and its target genes in CRC remain unclear. We identified 27 putative MIR452 target genes, and found that the vascular endothelial growth factor A (VEGFA) was a direct target gene of MIR452. Both cellular and extracellular VEGFA levels were significantly downregulated in CRC cells upon their transfection with MIR452 or siVEGFA. VEGFA expression was frequently downregulated in human CRC tissues in comparison with that in their healthy counterparts. We showed that MIR452 regulated the expression of genes in the VEGFA-mediated signal transduction pathways vascular endothelial growth factor receptor 1 (VEGFR2)–mitogen-activated protein kinase (MAPK) and VEGFR2–SRC proto-oncogene non-receptor tyrosine kinase (SRC) in CRC cells. Immunohistological analyses of xenografted MIR452-overexpressing CRC cells in mice showed that MIR452 regulated cell proliferation and angiogenesis. Furthermore, aortic ring angiogenesis assay in rats clearly showed that the number of microvessels formed was significantly reduced by MIR452 transfection. Our findings suggest that MIR452 regulates cell proliferation, cell migration, and angiogenesis by suppressing VEGFA expression in early CRC progression; therefore, MIR452 may have therapeutic value in relation to human CRC.