[Global DNA methylation changes during palatal formation in fetal mice induced by 2,3,7,8-tetrachlrodibenzo-p-dioxin].

Objective: To investigate global DNA methylation and DNA methyhransferases participation in the mechanism of cleft palate induced by maternal exposure to 2,3,7,8-tetrachlrodibenzo-p-dioxin (TCDD)in mice.
Methods: 40 pregnant C57BL/6J mice were randomly divided into 2 groups: the control group(n =20) and TCDD-exposure group(n =20).On gestation day 10.5 (GD10.5),the mice in TCDD-group were orally administrated with TCDD 28 μg/kg, while the mice in the control group received equivalent corn oil. The pregnant mice were sacrificed on GD13.5,GD14.5,GD15.5,GD16.5,GD17.5,fetal palates were collected for analysis. Global DNA methylation levels were detected by Methylamp TM Global DNA Methylation Quantification Ultra Kit through an ELISA-like reaction. The expression levels of DNA methyltransferases were examined by quantitative real-time PC R(q-PCR).IBM SPSS 20.0 software was applied for statistical analysis. Kolmogorov-Smirnov test was used for normal distribution check, and the distribution was normal. Independent t-test was carried out among two groups. P ＜ 0.05 was considered statistically significant.
Results: The global DNA methylation level in TCDD-exposure group was significantly higher than that in control group on GD13.5 (49.52％ ±4.03％ vs 33.42％ ± 6.78％,P ＜ 0.01),while lower on GD14.5 (24.10％ ±2.29％ vs 30.12％ ±3.92％,P ＜0.05) and on GD16.5 (32.77％ ±0.98％ vs 36.45％ ± 3.27％,P ＜ 0.05).The expression level of Dnmt1 mRNA in TCDD-exposure group was higher than that in control group on GD13.5(1.28±0.11 vs 1.01 ±0.10,P＜0.05) and on GD16.5(1.04 ±0.05 vs 0.81 ±0.01,P ＜0.01).The expression level of Dnmt3a mRNA in TCDD-exposure group was higher than that in control group on GD13.5 (1.15 ±0.17 vs 0.81 ±0.02,P ＜0.05)and on GD16.5 (1.11 ± 0.06 vs 0.96 ± 0.06,P ＜ 0.05).The expression level of Dnmt3b mRNA in TCDD-exposure group was higher than that in control group on GD14.5(0.97 ±0.06 vs 0.72 ±0.06,P ＜0.01).
Conclusions: It is supposed that complicated mechanisms are exist to regulate global DNA methylation levels in palatal tissue of fetal mice. The significant increased DNA methylation level on GD13.5 resulting from up-expression of Dnmt1 and Dnmt3a may be one of the epigenetic mechanisms which cause palate malformation in fetal mice induced by maternal exposure to TCDD.