OBJECTIVETo explore the interaction of folate deficiency and aberration of DNA methyltransferase 1 (DNMT1) in the progression of cervix carcinogenesis.

METHODSAll clinical samples were collected from 80 patients with cervix squamous cell carcinoma (SCC), 105 patients with cervical intraepithelial neoplasm (CINI, n = 52; CINII/III, n = 53) and 53 patients with cervix inflammation (CI). The participants were diagnosed by histology at Shanxi Province Tumor Hospital and Second Hospital of Shanxi Medical University during the period of September 2009 to May 2010. Meanwhile, cervical cancer cell lines Caski and C33A were treated with different concentration of folate. Radioimmunoassay (RIA), Western blotting and real-time PCR were used to detect the levels of serum folate, the expression of DNMT1 protein and mRNA, respectively. The data were analyzed by Student t test, ANOVA, chi-square test and Spearman correlation using SPSS statistical package. The correlation strength between factors and cervical canceration was calculated by OR and 95%CI value. Interaction effect was evaluated by the application of additive effect model.

RESULTSThe levels of serum folate (median inter-quartile range) were (2.66 ± 1.82), (2.83 ± 2.23), (3.17 ± 1.91) and (3.21 ± 1.74) ng/ml, the levels of DNMT1 protein (x(-) ± s) were 2.28 ± 0.55, 1.84 ± 0.37, 1.33 ± 0.38 and 0.92 ± 0.29, the Ct-ratio (Ct value of DNMT1/Ct value of β-actin) of DNMT1 mRNA (x(-) ± s) were 1.26 ± 0.13, 1.27 ± 0.12, 1.27 ± 0.12 and 1.33 ± 0.11 in the group of SCC, CINII/III, CINIand CI, respectively. The results showed that the serum folate levels were descended, and the expression levels of DNMT1 protein (χ(2)(tend) = 50.80, P < 0.05) and mRNA (χ(2)(tend) = 17.63, P < 0.05) were increased steadily with the severity of the cervix lesions. Moreover, our results revealed that there was an additive interaction between folate deficiency and high-expression of DNMT1 protein related to the risk of CIN and SCC. And it showed that the relative excess risk of interaction (RERI), attributable proportion of interaction (API) and synergy index(S) was 0.27, 0.14 and 1.40 in CINI group, 0.47, 0.19, 1.46 in CINII/III group, 1.60, 0.31, 1.61 in SCC group, respectively. It was found that folate was able to reduce the proliferation of Caski and C33A cells (r values were 0.954 and 0.969, all P values < 0.05), with 11.4% and 13.6% of growth inhibition at the concentration of 10 µg/ml, 64.8% and 49.4% at 1000 µg/ml in Caski and C33A cells, respectively. The result showed there was an inverse correlation between the levels of folate and DNMT1 protein (r values were -0.859 and -0.914, all P values < 0.05), with 1.96 and 1.92 of expression levels at the concentration of 10 µg/ml, and 1.60 and 1.38 at 1000 µg/ml in Caski and C33A cells, respectively. At folate concentration of 1000 µg/ml, the expression of DNMT1 protein or mRNA was higher in Caski cell than in C33A cell (t values were -4.22 and 3.50, all P values < 0.05).

CONCLUSIONOur finding indicated that the low levels of serum folate and high-expression of DNMT1 protein or mRNA seemed to be associated with high risk of cervical cancer and cervix precancerous lesion. Sufficient folate is able to effectively inhibit the growth of cervical cancer cells in vitro, and would counteract transcriptional and posttranscriptional aberration of DNMT1. It suggested that there might be a synergistic action between folate deficiency and aberration of DNMT1 in the progression of cervix carcinogenesis.