Search for down-regulated genes by epigenetic changes in human hepatoma via DNA demethylation approaches

• Main Authors: Chiu-Ping Wu, 吳秋萍
• Other Authors: Sen-Yung Hsieh
• Format: Others
• Language: zh-TW
• Published: 2005
• Online Access: http://ndltd.ncl.edu.tw/handle/46457128615367037365

碩士 === 長庚大學 === 基礎醫學研究所 === 93 === Gene silencing due to aberrant DNA methylation plays an important role in carcinogenesis. Recently, it was reported that synergy DNA methyl transferase inhibitor and histone deacetylase inhibition would lead to re-expression of genes silenced in cancer. To identify methylation-mediated silencing of genes in hepatocellular carcinoma (HCC), we surveyed genes induced by treatment with 5-aza-2’-deoxycytidine (5-Aza-dC) and Trichostatin A (TSA) in human hepatoma cell lines, Hep3B and Mahlavu, using cDNA microarray analysis and determined the methylation status of 5’ CpG islands by bisulfite genomic sequencing. Gene expression profile was altered after drug treatment and a total of 131 genes were identified based on two times individual cDNA microarray experiments. The up-regulation identified by cDNA microarray was further confirmed by quantitative real-time reverse transcriptase-polymerase chain reaction in six selected genes. Of the six, the demethylation effects by 5-Aza-dC treatment was further confirmed in MT1F, MT1X, SNCAIP and NFASC genes. Furthermore, MT1F and MT1X were highly down-regulated in HCC tissues. Of interests, MT (metallothionein) genes have been reported to be correlated with cell proliferation and cell differentiation. Down-regulation of MT1F and MT1X was very likely implicated in hepatocarcinogenesis. These findings suggest that combination of 5-Aza-dC and TSA treatment with cDNA microarray successfully facilitated identifying the down-regulated genes by epigenetic mechanism in human hepatoma.