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ORIGINAL ARTICLE
Year : 2015  |  Volume : 11  |  Issue : 4  |  Page : 818-822

Regulation of demethylation and re-expression of RASSF1A gene in hepatocellular carcinoma cell lines treated with NCTD in vitro


1 Department of Intervential Radiology, Affiliated Hospital of Jiangsu University, Zhenjiang, Jiangsu, P. R. China
2 Department of Gastroenterology, Affiliated Hospital of Jiangsu University, Zhenjiang, Jiangsu, P. R. China

Correspondence Address:
Min Xu
Department of Gastroenterology, Affiliated Hospital of Jiangsu University, Zhenjiang, 212000, Jiangsu
P. R. China
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/0973-1482.146126

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Background: Hepatocellular carcinoma, a lethal malignant neoplasm with poor prognosis, has dismal results of surgical resection and chemoradiotherapy. Norcantharidin (NCTD), the demethylated analog of cantharidin derived from a traditional Chinese medicine, Mylabris, has been used in the treatment of cancer. However, the detailed mechanisms underlying this process are generally unclear. Purpose: The aim of this study was to investigate the mechanism of NCTD-induced apoptosis in HepG2 cells. Materials and Methods: Human HepG2 cell lines were treated with NCTD at different concentrations (2.50, 5.00, 10.00, 20.00, 40.00 μg/mL) for 24 hours. Cell proliferation was evaluated by measurement of cellular 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT). The methylation levels of RASSF1A (Ras-association domain family 1 A) in HepG2 cells were detected by methylation-specific PCR (MSP). The mRNA levels of RASSF1A in HepG2 cells were detected by real-time fluorescent quantitative PCR (RT-PCR). The levels of RASSF1A protein expression of HepG2 cells were detected by Western blotting assay. Results: The inhibition of cell proliferation was observed when treated with NCTD at concentrations (2.5 μg/mL), and as concentration increased, the proliferation of HepG2 cells was markedly inhibited by NCTD in dose-dependent manners. The levels of methylation of RASSF1A decreased at the increasing concentration of 10, 20 and 40 μg/mL. The levels of RASSF1A mRNA and protein were decreased when treated with NCTD at the concentrations of 10, 20 and 40 μg/mL, which were also in a dose-dependent manner. Conclusion: NCTD can reverse the methylation state of RASSF1A gene and induce its re-expression, which will provide the theoretical basis for the clinical practice.


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