Naïve pluripotent embryonic stem (ESCs) cells and embryonic germ (EGCs) cells are derived from the preimplantation epiblast and primordial germ cells (PGCs), respectively. We investigated whether differences exist between ESCs and EGCs in view of their distinct developmental origins. PGCs are programmed to undergo global DNA demethylation; however we find that EGCs exhibit equivalent levels of global DNA methylation to ESCs. Importantly, inhibition of MEK and Gsk3b by 2i conditions leads to a pronounced reduction in DNA methylation in both cell types. This is driven by Prdm14 and is associated with downregulation of Dnmt3a and Dnmt3b. However, genomic imprints are maintained in 2i and we report derivation of EGCs with intact genomic imprints. Collectively, our findings establish that culture in 2i instils a naïve pluripotent state with a distinctive epigenetic configuration that parallels molecular features observed in both the preimplantation epiblast and nascent PGCs.

This study demonstrates a correlation between chronic alcohol use and demethylation of normally hypermethylated imprinted regions in sperm DNA. We hypothesize that, should these epigenetic changes in imprinted genes be transmitted through fertilization, they would alter the critical gene expression dosages required for normal prenatal development resulting in offspring with features of FASD.

Imprinted loci play a critical role in fetal development. Their expression is often regulated by CCCTC-binding factor (CTCF) protein binding at imprinting control regions (ICRs). Prenatal alcohol exposure has been shown to reduce global DNA methylation in the developing mouse fetus. This study explored the effect of preconception paternal alcohol exposure on DNA methylation at two paternally methylated ICRs (H19 and Rasgrf1) in the sperm of exposed males and somatic DNA of sired offspring. Significant reductions at the H19 CTCF 1 (p = 0.0027) and CTCF 2 (p = 0.0009) binding sites were observed in the offspring of ethanol-treated sires, which was significantly correlated with reduced weight at postnatal days 35–42 (p < 0.05). As birth weight was unaffected and growth was only delayed during the postnatal weaning period, with subsequent re-convergence, we hypothesize that this may be the result of a mental deficit causing delayed establishment of independent feeding following weaning and would explain why this effect is transient. No difference in DNA methylation was observed in the sperm of alcohol-exposed males, indicating that the transmission of the epigenetic signal at conception is not due to altered methylation, but may be the result of an RNA-mediated mechanism or altered chromatin remodeling.