Experimental research on methylation status of CpG islands in promoter region of TGF-β3 and Dnmts during TCDD-induced fetal palatogenisis / 中华整形外科杂志

ABSTRACT

Objective@#To investigate the correlation between CpG islands methylation statuses of TGF-β3, Dnmts and their expression during TCDD-induced mouse embryonic palatal development.@*Methods@#Eithtteen pregnant C57BL/6J mice were randomly divided into 2 groups the control group(n=9) and TCDD-exposure group(n=9). At 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 at GD13.5, GD14.5, GD15.5, fetal palates were collected. CpG island methylation statuses were analysed by methylation specific polymerase chain reaction(MSP). IBM SPSS 20.0 software was applied for statistical analysis. Kolmogorov-Smirnov test was used for normal distribution check, and the distributions were normal. Independent t-test was carried out between two groups. P<0.05 was considered statistically significant.@*Results@#CpG island in promoter region of gene TGF-β3 were all at low methylation level at all GDs of both groups, there were no differences at same GD between two groups [GD13.5 (8.6±0.8)% vs (8.7±0.8)%, P>0.05; GD14.5 (11.5±1.4)%vs (11.7±1.0)%, P>0.05; GD15.5 (12.0±0.7)% vs (12.1±0.5)%, P>0.05]. CpG island in promoter region of gene Dnmt1 were all highly methylated with no differences showed at same GD between two groups [GD13.5 (73.9±1.1)%vs (72.6±0.8)%, P>0.05; GD14.5 (70.8±1.7)% vs (70.7±1.0)%, P>0.05; GD15.5 (69.4±2.2)% vs (69.7±0.5)%, P>0.05]. The methylation level of CpG island 1 in promoter region of gene Dnmt3a in TCDD group was higher than that in control group at GD13.5 and GD15.5 [(21.9±1.1)% vs (8.1±0.6)%, P<0.01, (43.4±0.4)% vs(32.9±0.7)%, P<0.01], while lower at GD14.5[(33.2±0.5)% vs (42.9±0.3)%, P<0.01]. The methylation level of CpG island 2 in promoter region of gene Dnmt3a in control group was higher than that in TCDD group at all GDs [GD13.5 (82.0±0.7)% vs (32.3±0.6)%, P<0.01; GD14.5 (62.7±1.0)%vs (25.5±1.4)%, P<0.01; GD15.5 (47.2±0.4)% vs (30.3±1.4)%, P<0.01].@*Conclusions@#Low methylation level of CpG island 2 which is close to the first exon in promoter region of gene Dnm3a may be the cause of highly expressed Dnmt3a mRNA at GD13.5 during mice palatogenesis induced by TCDD, thus the global DNA methylation is extremely high to induce cleft palate. TCDD-treatment doesn′t influence the CpG methylation statuses in promoter region of TGF-β3 and Dnmt1.