Sodium bisulfite pyrosequencing revealed that developmental exposure to environmental contaminant mixtures does not affect DNA methylation of DNA repeats in Sprague-Dawley rats

Abstract

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.

Effects of cross-fostering and developmental exposure to mixtures of environmental contaminants on hepatic gene expression in prepubertal 21 days old and adult male Sprague-Dawley rats

“…In contrast to rats at PND21, mRNA levels of detoxification enzymes at PND78-86 were not significantly different from control, which is in agreement with the low chemical residue levels and absence of induced p450 activities in these rats (Desaulniers et al 2017). Nevertheless, DEG at PND78-86 suggest effects on energy (lipid/ glucose) and retinoid metabolism, molecular transport, small molecule biochemistry, bile acid biosynthesis, pro-inflammatory mechanisms, cell cycle, and contribution to diseases (Tables 2 and 3).…”

“…In general, the early gene sets at PND21 were associated with xenobiotic detoxification, oxidative stress, energy and retinoid metabolism, and immune responses, whereas at PND78-86 different gene sets were associated with retinoids and energy metabolism as potential longterm consequences. The chronic effects were subtle, perhaps given that the chemical residue levels in these rats declined to concentrations similar to those encountered in some human populations (Desaulniers et al 2017). Although species differences obscure interspecies extrapolation (Fader and Zacharewski 2017), the DEG at PND78-86 might assist in predicting mechanisms conducive to disorders previously described in EC exposed human populations (Andersen et al 2014;Kuwatsuka et al 2014;Mitoma et al 2015;Warner et al 2013), including dyslipidemia in Northern Canadian populations (Marushka et al 2018;Singh and Chan 2018).…”

“…The IPA prediction of induction of oxidative stress is in agreement with studies observing elevated hepatic glutathione S-transferase (GST) activities in Sprague-Dawley rats exposed to TCDD (Baars, Jansen, and Breimer 1978) or to non-ortho or ortho-chlorinated PCB (Lamartiniere, Dieringer, and Lucier 1979;Twaroski, O'Brien, and Robertson 2001), which are present in the mixture. The current rats exhibited increased activities of the major liver enzymes involved in xenobiotic metabolism, including CYP1A, 2B, and 3A (Desaulniers et al 2017). These enzymes alter steroid metabolism.…”

“…The HCA shows that samples of the M treatment clustered generally together with the highest degree of similarity. Effects measured at PND78-86 in the AhR and M treatment groups are due to the remaining chemical residues at that time (Desaulniers et al 2017), and to the chronic effects of developmental exposure that may generate data variability.…”

“…The selected doses were (1) low to intermediate relative to those of previous studies (Bowers et al 2004;Chu et al 2008); (2) did not markedly affect dam body weights or reproductive success; and (3) induced no overt signs of toxicity in the offspring. In addition, chemical residue levels in F1 adult males were comparable to those detected in humans (Desaulniers et al 2017;Desaulniers, Xiao, and Cummings-Lorbetskie 2013). Using liver samples from the same rats, the objective of the current follow-up study was to determine the effect of crossfostering in combination with developmental exposure to the EC mixture on hepatic gene expression profile at postnatal day 21 (PND21) and at PND78-86 in F1 males.…”