Hypomethylation of DNA repeats has been linked to diseases and cancer predisposition. Human studies suggest that higher blood concentrations of environmental contaminants (EC) correlate with levels of hypomethylation of DNA repeats in blood. The objective of this study was to examine the effect of in utero and/or lactational exposure to EC on the methylation of DNA repeats (LINE-1 and identifier element) in Sprague-Dawley rat pups at birth, at postnatal day (PND) 21, and in adulthood (PND78-86). From gestation day 0 to PND20, dams were exposed to a mixture "M" of polychlorinated biphenyls (PCB), pesticides, and methylmercury (MeHg), at 0.5 or 1 mg/kg/d (0.5M and M). At birth, some control (C) and M litters were cross-fostered to create the following in utero/postnatal exposure groups: C/C, M/C, C/M, M/M. Additional dams received 1.8 ng/kg/d of a mixture of aryl-hydrocarbon receptor (AhR) agonists (non-ortho-PCB, PC-dibenzodioxins, and PC-dibenzofurans) without or with 0.5M (0.5MAhR). Measurements of EC residue levels confirmed differences in their accumulation across treatments, age, and tissues. Although induction of hepatic detoxification enzyme activities (cytochrome P-450) demonstrated biological effects of treatments, the assessment of methylation in DNA repeats by sodium bisulfite pyrosequencing of liver, spleen, and thymus samples revealed no marked treatment-related effects but significant tissue- and age-related methylation differences. Further studies are required to determine whether absence of significant observable treatment effects on methylation of DNA repeats in the rat relate to tissue, strain, or species differences.
DNA methylation is an important epigenetic mechanism contributing to the regulation of gene expression and to the stability of DNA repeated sequences. Abnormal DNA methylation is a characteristic of cancer cells and there are scientific concerns that exposure to environmental contaminants (EC) might contribute to carcinogenesis by altering DNA methylation mechanisms. This study compared DNA methylation of repeated elements and cancerrelated genes in normal human liver, and in cancer (HepG2) and non-cancer (HC-04) cell lines, and investigated the effects of 5-aza-2'-deoxycytidine (5adC; a demethylation control) and polychlorinated biphenyl-126 (PCB126), a non-genotoxic rodent hepatocarcinogen. The results revealed striking celltype differences in DNA methylation of repeated elements (AluYb8, LINE-1, Sat-alpha) and 7/9 cancerrelated genes (CCND2, DAB2IP, DLEC1, GSTp1, OPCML, RASSF1, RUNX3, but not SFRP2 and SOCS1). In 72-h dose-response experiments, 5adC induced "U" shape demethylation responses in the nine investigated genes, but associated with elevated mRNA expression only in 6/9 and 3/9 genes in HC-04 and HepG2 cells, respectively. DNA methylation was resistant to PCB126 in most genes in both cell lines, except for reduced promoter methylation and increased expression of GSTp1 in HepG2 cells, which further support a role for oxidative stress in PCB126induced oncogenesis/toxicity. Finally, the different DNA methylation patterns and gene expression responses between the non-cancer HC-04 and cancer HepG2 cell lines provide alternative models to further explore epigenetic divergence in gene expression regulation.
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