Childhood adversity increases methylation in the GRIN2B gene.

Childhood adversity is an early life stressor associated with increased risk of several psychiatric disorders such as depression. Epigenetic changes, primarily DNA methylation, can be affected by early life stress, which in turn might contribute to altered disease susceptibility later in life. One plausible biomarker of early life stress is methylation of the ionotropic glutamate receptor NMDA type subunit 2B (GRIN2B) gene, which has been previously shown to be epigenetically affected by prenatal environmental stressors. Here, we set out to investigate if stress-inducing adversity during childhood is associated with changes in methylation of GRIN2B in adulthood. We studied 186 individuals from a Swedish naturalistic population-based cohort who had provided saliva samples (DNA) as well as information regarding both childhood adversity (CA) and depressive symptoms (dep) (nCA,dep = 41, nCA,no-dep = 56, nno-CA,dep = 40, Nno-CA,no-dep = 49). Methylation at four CpG sites in a regulatory region of GRIN2B was analysed using bisulfite pyrosequencing. Associations for methylation status to childhood adversity and to depression status were investigated using linear regression models. Our study shows that childhood adversity is associated with increased methylation levels of GRIN2B in adulthood, for three of the measured CpGs (p = 0.007, 0.006 and 5 × 10-14). This indicates that GRIN2B methylation is susceptible to early life stress, and that methylation at this gene is persistent over time. No association was found between GRIN2B methylation and depression status. Yet, this does not rule out a role for alterations in GRIN2B methylation for other neuropsychological outcomes not studied here.

DNA methylation at GRIN2B partially mediates the association between prenatal bisphenol F exposure and cognitive functions in 7-year-old children in the SELMA study.

BACKGROUND: Accumulating evidence suggests that prenatal chemical exposure triggers epigenetic modifications that could influence health outcomes later in life. In this study, we investigated whether DNA methylation (DNAm) levels at the glutamate ionotropic receptor NMDA type subunit 2B (GRIN2B) gene underlies the association between prenatal exposure to an endocrine disrupting chemical (EDC), bisphenol F (BPF), and lower cognitive functions in 7-year-old children. METHODS: Data from 799 children participating in the Swedish Environmental Longitudinal Mother and child Asthma and allergy (SELMA) pregnancy cohort was analyzed. Prenatal BPF exposure was assessed by measuring BPF levels in maternal urine. At age 7, DNAm of three CpG sites in a regulatory region of the GRIN2B gene was analyzed from buccal swabs using bisulfite-Pyrosequencing. Cognitive functions, including full-scale IQ and four subscales, were evaluated using the Wechsler Intelligence Scale for Children (WISC-IV). Associations between prenatal BPF exposure and GRIN2B DNAm, as well as between GRIN2B DNAm and cognitive functions, were determined using regression models adjusted for potential confounders. Generalized structural equation models (gSEM) were used to evaluate if GRIN2B DNAm mediates the association between prenatal BPF exposure and cognitive functions at 7 years of age. RESULTS: Prenatal BPF exposure was positively associated with GRIN2B DNAm levels at the third CpG site (CpG3), while CpG3 methylation was inversely associated with cognitive test scores. Mediation analyses showed that CpG3 methylation exerted 6-9% of the association between BPF exposure and full-scale IQ, as well as verbal comprehension and perceptual reasoning in boys, while not significant in girls. CONCLUSIONS: This study is the first to identify locus-specific DNAm as a mediating factor underlying an epidemiological association between prenatal EDC exposure and cognitive functions in childhood. It also confirms previous findings, that GRIN2B DNAm is responsive to environmental exposures.

Gabapentin increases expression of δ subunit-containing GABAA receptors.

BACKGROUND: Gabapentin is a structural analog of the inhibitory neurotransmitter γ-aminobutyric acid (GABA). Its anticonvulsant, analgesic and anxiolytic properties suggest that it increases GABAergic inhibition; however, the molecular basis for these effects is unknown as gabapentin does not directly modify GABA type A (GABAA) receptor function, nor does it modify synaptic inhibition. Here, we postulated that gabapentin increases expression of δ subunit-containing GABAA (δGABAA) receptors that generate a tonic inhibitory conductance in multiple brain regions including the cerebellum and hippocampus. METHODS: Cell-surface biotinylation, Western blotting, electrophysiologic recordings, behavioral assays, high-performance liquid chromatography and gas chromatography-mass spectrometry studies were performed using mouse models. FINDINGS: Gabapentin enhanced expression of δGABAA receptors and increased a tonic inhibitory conductance in neurons. This increased expression likely contributes to GABAergic effects as gabapentin caused ataxia and anxiolysis in wild-type mice but not δ subunit null-mutant mice. In contrast, the antinociceptive properties of gabapentin were observed in both genotypes. Levels of GABAA receptor agonists and neurosteroids in the brain were not altered by gabapentin. INTERPRETATION: These results provide compelling evidence to account for the GABAergic properties of gabapentin. Since reduced expression of δGABAA receptor occurs in several disorders, gabapentin may have much broader therapeutic applications than is currently recognized. FUND: Supported by a Foundation Grant (FDN-154312) from the Canadian Institutes of Health Research (to B.A.O.); a NSERC Discovery Grant (RGPIN-2016-05538), a Canada Research Chair in Sensory Plasticity and Reconsolidation, and funding from the University of Toronto Centre for the Study of Pain (to R.P.B.).

Understanding Epigenetic Effects of Endocrine Disrupting Chemicals: From Mechanisms to Novel Test Methods.

Endocrine-disrupting chemicals (EDCs) are man-made chemicals that interfere with hormonal signalling pathways. They are used in, for example, production of common household materials, in resin-based medical supplies and in pesticides. Thus, they are environmentally ubiquitous and human beings and wildlife are exposed to them on a daily basis. Early-life exposure to EDCs has been associated with later-life adversities such as obesity, diabetes and cancer. Mechanisms underlying such associations are unknown but are likely to be mediated by epigenetic changes induced by EDCs. Epigenetics is the study of changes in gene function that are heritable but do not entail a change in DNA sequence. EDCs have been shown to affect epigenetic marks such as DNA methylation and histone modifications. The scope of this article was to review today's knowledge about mechanisms involved in EDC-induced epigenetic changes and to discuss how this knowledge could be used for designing novel methods addressing epigenetic effects of EDCs.

Prenatal Bisphenol A Exposure is Linked to Epigenetic Changes in Glutamate Receptor Subunit Gene Grin2b in Female Rats and Humans.

Bisphenol A (BPA) exposure has been linked to neurodevelopmental disorders and to effects on epigenetic regulation, such as DNA methylation, at genes involved in brain function. High doses of BPA have been shown to change expression and regulation of one such gene, Grin2b, in mice. Yet, if such changes occur at relevant doses in animals and humans has not been addressed. We investigated if low-dose developmental BPA exposure affects DNA methylation and expression of Grin2b in brains of adult rats. Furthermore, we assessed associations between prenatal BPA exposure and Grin2b methylation in 7-year old children. We found that Grin2b mRNA expression was increased and DNA methylation decreased in female, but not in male rats. In humans, prenatal BPA exposure was associated with increased methylation levels in girls. Additionally, low APGAR scores, a predictor for increased risk for neurodevelopmental diseases, were associated with higher Grin2b methylation levels in girls. Thus, we could link developmental BPA exposure and low APGAR scores to changes in the epigenetic regulation of Grin2b, a gene important for neuronal function, in a sexual dimorphic fashion. Discrepancies in exact locations and directions of the DNA methylation change might reflect differences between species, analysed tissues, exposure level and/or timing.

Developmental exposure to bisphenol A alters expression and DNA methylation of Fkbp5, an important regulator of the stress response.

Bisphenol A (BPA), an abundant endocrine disruptor, affects stress-responsiveness and related behaviors in children. In rats, perinatal BPA exposure modifies stress response in pubertal offspring via unknown mechanisms. Here we examined possible epigenetic modifications in the glucocorticoid receptor gene and its regulator Fkbp5 in hypothalamus and hippocampus of exposed offspring. We found increased DNA methylation of Fkbp5 and reduced protein levels in the hippocampus of exposed male rats. Similar effects were obtained in a male hippocampal cell line when exposed to BPA during differentiation. The estrogen receptor (ER) antagonist ICI 182,780 or ERß knock-down affected Fkbp5 expression and methylation similarly to BPA. Further, BPA's effect on Fkbp5 was abolished upon knock-down of ERß, suggesting a role for this receptor in mediating BPA's effects on Fkbp5. These data demonstrate that developmental BPA exposure modifies Fkbp5 methylation and expression in male rats, which may be related to its impact on stress responsiveness.

15-deoxy-Δ¹²,¹4-PGJ2 promotes inflammation and apoptosis in cardiomyocytes via the DP2/MAPK/TNFα axis.

BACKGROUND: Prostaglandins (PGs), lipid autacoids derived from arachidonic acid, play a pivotal role during inflammation. PGD2 synthase is abundantly expressed in heart tissue and PGD2 has recently been found to induce cardiomyocyte apoptosis. PGD2 is an unstable prostanoid metabolite; therefore the objective of the present study was to elucidate whether its final dehydration product, 15-deoxy-Δ¹²,¹4-PGJ2 (15d-PGJ2, present at high levels in ischemic myocardium) might cause cardiomyocyte damage. METHODS AND RESULTS: Using specific (ant)agonists we show that 15d-PGJ2 induced formation of intracellular reactive oxygen species (ROS) and phosphorylation of p38 and p42/44 MAPKs via the PGD2 receptor DP2 (but not DP1 or PPARγ) in the murine atrial cardiomyocyte HL-1 cell line. Activation of the DP2-ROS-MAPK axis by 15d-PGJ2 enhanced transcription and translation of TNFα and induced apoptosis in HL-1 cardiomyocytes. Silencing of TNFα significantly attenuated the extrinsic (caspase-8) and intrinsic apoptotic pathways (bax and caspase-9), caspase-3 activation and downstream PARP cleavage and γH2AX activation. The apoptotic machinery was unaffected by intracellular calcium, transcription factor NF-κB and its downstream target p53. Of note, 9,10-dihydro-15d-PGJ2 (lacking the electrophilic carbon atom in the cyclopentenone ring) did not activate cellular responses. Selected experiments performed in primary murine cardiomyocytes confirmed data obtained in HL-1 cells namely that the intrinsic and extrinsic apoptotic cascades are activated via DP2/MAPK/TNFα signaling. CONCLUSIONS: We conclude that the reactive α,ß-unsaturated carbonyl group of 15d-PGJ2 is responsible for the pronounced upregulation of TNFα promoting cardiomyocyte apoptosis. We propose that inhibition of DP2 receptors could provide a possibility to modulate 15d-PGJ2-induced myocardial injury.

Spermidine protects against α-synuclein neurotoxicity.

As our society ages, neurodegenerative disorders like Parkinson`s disease (PD) are increasing in pandemic proportions. While mechanistic understanding of PD is advancing, a treatment with well tolerable drugs is still elusive. Here, we show that administration of the naturally occurring polyamine spermidine, which declines continuously during aging in various species, alleviates a series of PD-related degenerative processes in the fruit fly Drosophila melanogaster and the nematode Caenorhabditis elegans, two established model systems for PD pathology. In the fruit fly, simple feeding with spermidine inhibited loss of climbing activity and early organismal death upon heterologous expression of human α-synuclein, which is thought to be the principal toxic trigger of PD. In this line, administration of spermidine rescued α-synuclein-induced loss of dopaminergic neurons, a hallmark of PD, in nematodes. Alleviation of PD-related neurodegeneration by spermidine was accompanied by induction of autophagy, suggesting that this cytoprotective process may be responsible for the beneficial effects of spermidine administration.

The cell death protease Kex1p is essential for hypochlorite-induced apoptosis in yeast.

Following microbial pathogen invasion, the human immune system of activated phagocytes generates and releases the potent oxidant hypochlorous acid (HOCl), which contributes to the killing of menacing microorganisms. Though tightly controlled, HOCl generation by the myeloperoxidase-hydrogen peroxide-chloride system of neutrophils/monocytes may occur in excess and lead to tissue damage. It is thus of marked importance to delineate the molecular pathways underlying HOCl cytotoxicity in both microbial and human cells. Here, we show that HOCl induces the generation of reactive oxygen species (ROS), apoptotic cell death and the formation of specific HOCl-modified epitopes in the budding yeast Saccharomyces cerevisiae. Interestingly, HOCl cytotoxicity can be prevented by treatment with ROS scavengers, suggesting oxidative stress to mediate the lethal effect. The executing pathway involves the pro-apoptotic protease Kex1p, since its absence diminishes HOCl-induced production of ROS, apoptosis and protein modification. By characterizing HOCl-induced cell death in yeast and identifying a corresponding central executor, these results pave the way for the use of Saccharomyces cerevisiae in HOCl research, not least given that it combines both being a microorganism as well as a model for programmed cell death in higher eukaryotes.