ABSTRACT
<p><b>OBJECTIVE</b>To determine DNA methylation status of ZIC1 and KLOTHO gene in colorectal carcinomas and its relationship with clinicopathological features of patients.</p><p><b>METHODS</b>The mRNA expression of ZIC1 and KLOTHO genes in colorectal carcinomas was detected by real-time quantitative RT-PCR, and the promoter methylation status was detected by methylation specific PCR (MSP). The relationship of ZIC1 and KLOTHO methylation status with clinicopathological features of colorectal carcinoma was analyzed.</p><p><b>RESULT</b>The mRNA expression levels of ZIC1 and KLOTHO genes were significantly down-regulated in tumor tissues when compared to adjacent nontumor tissues (P<0.001). ZIC1 and KLOTHO methylation was detected in 80.0%(20/25) and 76.0%(19/25) of colorectal tumor tissues, respectively, and the both positive rate was 64.0%(16/25).</p><p><b>CONCLUSION</b>The down-regulated expression of ZIC1 and KLOTHO in colorectal carcinoma may relate to promoter methylation. The detection of methylation of ZIC1 and KLOTHO gene potentially provides biomarkers for diagnosis of colorectal carcinoma.</p>
Subject(s)
Adult , Aged , Female , Humans , Male , Middle Aged , Colorectal Neoplasms , Genetics , DNA Methylation , Gene Expression Regulation, Neoplastic , Glucuronidase , Genetics , Promoter Regions, Genetic , Genetics , Transcription Factors , GeneticsABSTRACT
<p><b>OBJECTIVE</b>To construct COL1A1-targeted short hairpin RNA (shRNA) vector with pSilencer 4.1-CMV neo siRNA expression vector and to evaluate its effect on proliferation and migration of gastric cancer BGC-823 cells in vitro.</p><p><b>METHODS</b>Three COL1A1-shRNA plasmids (COL1A1-shRNA-1, COL1A1-shRNA-2, COL1A1-shRNA-3), targeting different sites of COL1A1 gene, were constructed using pSilencer 4.1-CMV neo siRNA expression vector and transfected into gastric cancer BGC-823 cells. Real time quantitative RT-PCR and Western blot were performed to detect expression levels of COL1A1. MTT and Transwell migration assays were employed to evaluate the effects of COL1A1 gene silence on cell proliferation and migration.</p><p><b>RESULT</b>Three recombinant plasmids targeting COL1A1 were constructed successfully. The expressions of COL1A1 in BGC-823 cells, including mRNA and protein levels, were significantly inhibited by the COL1A1-shRNA transfectants, which resulted in a clear reduction of cell proliferation and migration capacity.</p><p><b>CONCLUSION</b>The COL1A1-shRNA can effectively knock down gene expression and inhibit proliferation and migration of gastric cancer BGC-823 cells.</p>