Effect of folate in modulating the expression of DNA methyltransferase 1 and methyl-CpG-bingding protein 2 in cervical cancer cell lines / 中华流行病学杂志

Objective To explore the effects of folate on the expression of DNA methyltransferase 1 (DNMT1) and methyl-CpG-bingding protein 2 (MeCP2) in cervical cancer cell lines.Methods Experimental study was carried out in vitro.Human cervical cancer cell lines,including C33A cell with HPV negative and Caski cell with HPV16 positive,were treated with different concentration of folate.The expression of DNMT1 and MeCP2 protein (by Western blot)and mRNA (by real-time PCR) were then detected in the two cell lines.Results It was found that supplement of folate was able to reduce the cell proliferation in C33A cell (r=0.984,P＜0.001) and Caski cell (r=0.978,P=0.002),as well as induced the cell apoptosis (C33A:r=0.989,P＜0.001 ;Caski:r=0.994,P＜0.001).Results showed that the expression levels of DNMT1 protein (C33A:r=-0.914,P＜ 0.001 ; Caski:r=-0.859,P=0.003) and MeCP2 protein (C33A:r=-0.830,P=0.005 ;Caski:r=-0.981,P＜0.001) decreased gradually with the increase of folate concentrations,but the expression of DNMT1 and MeCP2 mRNA was not observed in Caski or C33A cell.When at the same levels of folate,the expression of DNMT1 protein or mRNA was higher in Caski cell than in C33A cell.However,the expression of MeCP2 protein or mRNA was higher in C33A cell than in Caski cell.Conclusion Our fimding indicated that adequate foleta could effectively inhibit the proliferation of cervical cancer cells and facilitate their apoptosis in vitro,thus would reverse the aberration protein expression of DNMTl and MeCP2.That there might be a synergistic action between HPV16 infection and parafunction of DNMT l in cervical cancer,being noticed.

Detection of p16(INK4A) in the Mixed Cell Populations of Normal Peripheral Blood Mononuclear Cells and Cervical Cancer Cell Lines / Journal of the Korean Cancer Association, 대한암학회지

PURPOSE: Human papilloma viruses (HPVs) play a central role in the pathogenesis of neoplastic lesions of the uterine cervix. The viral oncoprotein HPV E6 degrades the p53 protein, and the HPV E7 protein inactivates pRB and increases the expression of the CDK inhibitor, p16(INK4A). We investigated the usefulness of p16(INK4A) as a biologic marker for the cervical dysplastic and neoplastic cells. MATERIALS AND METHODS: We examined the expression of p16(INK4A) and cytokeratin in a mixed population of normal peripheral blood mononuclear cells (PBMC) and the cervical cancer cell lines (HeLa, SiHa, and CasKi) using flow cytometry. RESULTS: The DNA indices of the HeLa, SiHa and CasKi cell lines were 1.89, 1.53 and 1.75, respectively, indicating that these cells are aneuploid cells. Furthermore, the positive rate of p16(INK4A) expression was 86.7% for the HeLa mixed population, 85.6% for the SiHa mixed population, and 92.2% for the CasKi mixed population. According to the FL3A vs FL3W histogram, electrical gating of the HeLa, SiHa and CasKi mixed populations showed the expression levels of both cytokeratin and p16(INK4A) to be identical, at 86.6%, 84.8% and 85.0%, respectively. These findings revealed that almost all cells selected through electrical gating were cervical cancer cells originating from the epithelium and which expressed cytokeratin and p16(INK4A). On the other hand, when each mixed population was electrically gated for normal PBMC, we found that the PBMCs expressed neither cytokeratin nor p16(INK4A). CONCLUSION: Using flow cytometry, we observed the enhanced expression of p16(INK4A) in cervical cancer cell lines. These RESULTS suggest the usefulness of p16(INK4A) for the selective detection of cervical dysplastic and cancer cells in the liquid-based samples, which are taken from the cervices and contaminated with blood and stromal cells.

The Mechanisms of the Antiproliferative Effect by Interferon- a in Cervical Cancer Cell Lines / 대한부인종양콜포스코피학회잡지

Interferons(IFNs) exhibit an antiproliferative effect on many normal and transformed cells and have in vivo antitumor activity in a variety of cancers. Recent clinical studies have suggested major activity with IFNs, especially in advanced squamous cell carcinoma of the skin and cervix. With the objective of exploring how the IFNs might work in squamous carcinoma cell line, we studied the effect of IFN-a on cervical cancer cell lines. The effect of IFNs on apoptosis and cell cycle of cervical cancer cell lines(C33A, CaSki, SiHa, HeLa, ME-180) was analysed by flow cytometry in time dependent manner. Results were as follows: (1) According to cell count of studied cancer cell lines treated with 2,000 IU/ml IFN-a for 7 days exposure, IFN-a had the antiproliferative effect on all five tested cervical cancer cell lines. Also this antiproliferative effect was confirmed by WST-1 assay. (2) The effect of IFN-a on apoptosis of each cultute was analysed by flow cytometry after 3 days and 7 days exposure with 2,000 IU/ml IFN-a, Apoptosis was not induced by IFN-a treatment. (3) The effect of IFN-a on the cell cycle of each culture was analysed by flow cytometry after 3 days exposure with 2,000 IU/ml IFN-a. As compared to control cells, treatment with IFN-a resulted in a higher proportion of cells in S phase with lower portion of cells with G2/M phase. (4) Time course of IFN-a effect on HPV 16 and HPV 18 E6 mRNA levels was evaluated by northern blot analysis. In CaSki cell line, HPV 16 E6 mRNA expression induced by IFN-a was not inhibited. But in HeLa cell line, HPV 18 E6 inRNA expression was inhibited. IFN-a appears to have the antiproliferative effect on all five tested cervical cancer cell lines and the antiproliferative effect of IFN-a seemed to be induced not by apoptosis but by disruption on specific cell cycle. Also regulation of HPV E6 mRNA expression induced by IFN-a is not directly related to the mechanisms of the antiproliferative effect of IFN-a in cancer cell lines.