Demethylation in the promoter area by the antisense of human DNA MTase gene / 中华肿瘤杂志

ABSTRACT

<p><b>OBJECTIVE</b>To investigate the change in the expression of E-Cadherin of human hepatocarcinoma cell line SMMC-7721 after transfection by antisense human DNA MTase gene.</p><p><b>METHODS</b>DNA MTase gene eukaryotic expression vectors, including sense and antisense fragments, were constructed with recombinant technology and transfected into the hepatocarcinoma cell line SMMC-7721 with liposome DOTAP. The expression of DNA MTase gene mRNA and E-Cadherin gene mRNA was examined with RT-PCR and the expression of E-Cadherin with immunohistochemical and flow cytometry. The status of methylation in E-Cadherin gene promoter area was examined with methylation specific PCR (MSP).</p><p><b>RESULTS</b>The sense and antisense eukaryotic expression vectors were successfully constructed and then the constructed recombinant plasmids were successfully transfected into SMMC-7721 cell with liposome DOTAP. The expression of endogenous DNA MTase mRNA was obviously decreased with E-Cadherin gene mRNA and its activity increased in the SMMC-7721 cell, which was tranfected with antisense DNA MTase gene fragment. Moreover, demethylation in the promoter area of E-Cadherin gene was observed with MSP.</p><p><b>CONCLUSION</b>Demethylation in the promoter area and increasing mRNA level of E-Cadherin gene can be induced by expression inhibition of DNA MTase gene of SMMC-7721 cell line.</p>