title: Effects of DNMT1 silencing on malignant phenotype and methylated gene expression in cervical cancer cells.
abstract: BACKGROUND
  DNA methylation has been widely used in classification, early diagnosis, therapy and prediction of metastasis as well as recurrence of cervical cancer. DNMT methyltransferase 1 (DNMT1), which plays a significant role in maintaining DNA methylation status and regulating the expression of tumor suppressor genes. The aim of this research was to investigate the relationship between DNMT1 and abnormal methylation of tumor suppressor genes and malignant phenotype in cervical cancer.
  
  
  METHODS
  Levels of DNMT1 mRNA and protein were detected using qPCR and Western blot, respectively. Cell proliferation was analyzed by MTT and apoptosis was performed by Annexin V-FITC/PI double staining flow cytometry, respectively. MeDIP-qPCR and qPCR were performed to measure demethylation status and mRNA re-expression level of 7 tumor-suppressor genes (CCNA1, CHFR, FHIT, PAX1, PTEN, SFRP4, TSLC1) in Hela and Siha cells after silencing DNMT1.
  
  
  RESULTS
  The average expression levels of DNMT1 mRNA and protein in Hela and Siha cells were decreased significantly compared with control group. The flow cytometry and MTT results showed that Hela and Siha cells apoptosis rates and cell viabilities were 19.4 ± 2.90%, 25.7 ± 3.92% as well as 86.7 ± 3.12%, 84.16 ± 2.67% respectively 48 h after transfection (P < 0.01). Furthermore, the promoter methylation of five tumor suppressor genes was decreased with the increased mRNA expression after silencing DNMT1, whereas there were no significant changes in PTEN and FHIT genes in Hela cells, and CHFR and FHIT genes in Siha cells.
  
  
  CONCLUSIONS
  Our experimental results demonstrate that methylation status of DNMT1 can influence several important tumor suppressor genes activity in cervical tumorigenesis and may have the potential to become an effective target for treatment of cervical cancer.
authors: The Second Medical College, Jinan University, Shenzhen Clinical Medical Research Center, Shenzhen People's Hospital, 518020 Shenzhen, PR China.
authors: Yi
authors: Y
authors: Zhang
authors: Fu-qiang
authors: FQ
authors: Chen
authors: Ye-hong
authors: YH
authors: Sun
authors: Shu-yan
authors: SY
authors: Zhou
authors: Ti-yuan
authors: TY
authors: Li
authors: Rui
authors: R
authors: Chen
mesh_terms: D017209:Apoptosis; D015153:Blotting, Western; D002453:Cell Cycle; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D018899:CpG Islands; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000235:genetics* / Q000378:metabolism; D019175:DNA Methylation; D004273:DNA, Neoplasm / Q000235:genetics; D005260:Female; D005434:Flow Cytometry; D015972:Gene Expression Regulation, Neoplastic; D020868:Gene Silencing; D016147:Genes, Tumor Suppressor / Q000502:physiology*; D006801:Humans; D010641:Phenotype; D011401:Promoter Regions, Genetic / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D002583:Uterine Cervical Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000473:pathology*
chemical_list: D004273:DNA, Neoplasm; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases; C000619380:DNMT1 protein, human