[Effect of Hydralazine on demethylation status and expression of APC gene, proliferation and apoptosis of human cervical cancer cell lines].
OBJECTIVE
To investigate the correlation between methylation status and gene expression of APC (adenomatous polyposis coli) gene in HeLa, CaSki and SiHa cell lines of cervical carcinoma, and explore the effect of hydralazine on the transcription regulation of the 5'CpG island demethylation of APC gene and the proliferation and apoptosis of the cell lines.


METHODS
Methylation status and the expression of APC gene were analyzed using methylated specific PCR, RT-PCR and FQ-PCR methods. The expression of beta-catenin protein which correlates closely with APC was detected by SP method after treatment with Hydralazine. MTT and FCM assays were used to observe the changes of proliferation activity and apoptosis of the cells after Hydralazine treatment.


RESULTS
(1) APC gene was methylated or hemimethylated respectively in HeLa and CaSki cell lines, at the same time, APC gene was not methylated in SiHa cell. (2) After having been treated by 40 micromol/L Hydralazine for 72 hours, growth inhibitory ratios of HeLa, CaSki and SiHa cell lines were (52.12 +/- 3.78)%, (44.31 +/- 2.59)% and (47.73 +/- 4.73)% respectively, on the contrary, normal cell HECV's growth inhibitory ratio was only (27.18 +/- 0.79)%. APC gene in HeLa and CaSki cell lines which were treated by 40 micromol/L Hydralazine for 72 hours was demethylated and expressed positively, the expression of APC mRNA in HeLa, CaSki and SiHa cell lines increased to 10.35, 11.40 and 0.73 times respectively. (3) Hydralazine, when used at the concentration of 40 micromol/L for 72 hours, induced S phase and G2/M phase arrest and apoptosis in HeLa and CaSki cells. beta-catenin protein can be expressed in cell membrane after treatment with Hydralazine.


CONCLUSIONS
APC gene methylation plays an important role in the carcinogenesis of cervical cells and can re-express after the treatment with Hydralazine which also could inhibit the growth of the cervical cancer cells.
Song
Yin-hong
YH
Department of Immunology, Medical College of Three Gorges University, Yichang, Hubei 443002, China.
Zhang
Chang-jü
CJ
D025601:Adenomatous Polyposis Coli Protein / Q000235:genetics / Q000378:metabolism*; D000970:Antineoplastic Agents / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects*; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects*; D018899:CpG Islands / Q000235:genetics; D019175:DNA Methylation; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D017491:Genes, APC; D006367:HeLa Cells; D006801:Humans; D006830:Hydralazine / Q000494:pharmacology*; D012333:RNA, Messenger / Q000378:metabolism; D002583:Uterine Cervical Neoplasms / Q000378:metabolism / Q000473:pathology; D051176:beta Catenin / Q000378:metabolism
D025601:Adenomatous Polyposis Coli Protein; D000970:Antineoplastic Agents; D012333:RNA, Messenger; D051176:beta Catenin; D006830:Hydralazine