Background: Environmental epidemiologists have shown increasing interest for alternative epigenetic markers, such as 5-hydroxymethylcytosine (5hmC) and methylation of mitochondrial DNA. Experimental models are needed to identify environmentally sensitive markers before their translation to epidemiology studies. Aims: We measured 5hmC and methylation of mitochondrial and nuclear genes related to behavioral and brain functions on rat brain from a toxicity study of PBDEs, bioaccumulative flame retardants that have been linked with neurodevelopmental effects. Methods: Wistar dams were exposed to BDE-47 (2 and 200μg/Kg body weight) from gestation day 9 and 16 to postnatal day 1, 8, and 15. Frontal lobes of offspring were used to measure: 1) global 5hmC and methylcytosine (5mC) by fluorometric quantification; 2) mitochondrial gene methylation, including COX1, COX2, and COX3, and 3) Nr3c1, Crhr1, Mc2r, Bdnf and Snca through bisulfite pyrosequencing. Results: Pre- and perinatal exposure to BDE-47 had significant effects on genomic and mitochondrial DNA methylation in rat brain. Global 5hmC methylation was significantly increased (p-value=0.024) with 200μg/Kg of BDE-47 exposure, while global L1Rn UTR methylation levels in frontal lobe decreased with 2μg/Kg of BDE-47 exposure (p-value=0.024). Mitochondrial COX1 and 2 genes showed a marginal decrease in methylation with 200μg/Kg of BDE-47 exposure (p-value=0.096 in overall CpGs from COX1 and p-value=0.009 in the 1st CpG from COX2) in frontal lobe. Mitochondrial COX3 showed a marginal decrease in methylation with 2μg/Kg of BDE-47 exposure in frontal lobe (p-value=0.09 in overall CpGs). A CpG site-specific decrease in methylation was observed in Bdnf and Nr3cl from 2 and 200μg/Kg BDE-47 exposed rats, not in a dose dependent manner. Conclusions: Our data provide molecular evidence of epigenetie mechanisms of PBDE toxicity that may affect neurodevelopment and neuronal functions during vulnerable periods of brain differentiation.
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