Folate and Cobalamin Deficiencies during Pregnancy Disrupt the Glucocorticoid Response in Hypothalamus through N-Homocysteinilation of the Glucocorticoid Receptor.

Vitamin B9 (folate)/B12 (cobalamin) deficiency is known to induce brain structural and/or functional retardations. In many countries, folate supplementation, targeting the most severe outcomes such as neural tube defects, is discontinued after the first trimester. However, adverse effects may occur after birth because of some mild misregulations. Various hormonal receptors were shown to be deregulated in brain tissue under these conditions. The glucocorticoid receptor (GR) is particularly sensitive to epigenetic regulation and post-translational modifications. In a mother-offspring rat model of vitamin B9/B12 deficiency, we investigated whether a prolonged folate supplementation could restore the GR signaling in the hypothalamus. Our data showed that a deficiency of folate and vitamin B12 during the in-utero and early postnatal periods was associated with reduced GR expression in the hypothalamus. We also described for the first time a novel post-translational modification of GR that impaired ligand binding and GR activation, leading to decrease expression of one of the GR targets in the hypothalamus, AgRP. Moreover, this brain-impaired GR signaling pathway was associated with behavioral perturbations during offspring growth. Importantly, perinatal and postnatal supplementation with folic acid helped restore GR mRNA levels and activity in hypothalamus cells and improved behavioral deficits.

Brain Susceptibility to Methyl Donor Deficiency: From Fetal Programming to Aging Outcome in Rats.

Deficiencies in methyl donors, folate, and vitamin B12 are known to lead to brain function defects. Fetal development is the most studied but data are also available for such an impact in elderly rats. To compare the functional consequences of nutritional deficiency in young versus adult rats, we monitored behavioral outcomes of cerebellum and hippocampus circuits in the offspring of deficient mother rats and in adult rats fed a deficient diet from 2 to 8 months-of-age. We present data showing that the main deleterious consequences are found in young ages compared to adult ones, in terms of movement coordination and learning abilities. Moreover, we obtained sex and age differences in the deleterious effects on these functions and on neuronal layer integrity in growing young rats, while deficient adults presented only slight functional alterations without tissue damage. Actually, the cerebellum and the hippocampus develop and maturate according to different time lap windows and we demonstrate that a switch to a normal diet can only rescue circuits that present a long permissive window of time, such as the cerebellum, whereas the hippocampus does not. Thus, we argue, as others have, for supplements or fortifications given over a longer time than the developmental period.

Methyl Donor Deficiency during Gestation and Lactation in the Rat Affects the Expression of Neuropeptides and Related Receptors in the Hypothalamus.

The micronutrients vitamins B9 and B12 act as methyl donors in the one-carbon metabolism involved in transmethylation reactions which critically influence epigenetic mechanisms and gene expression. Both vitamins are essential for proper development, and their deficiency during pregnancy has been associated with a wide range of disorders, including persisting growth retardation. Energy homeostasis and feeding are centrally regulated by the hypothalamus which integrates peripheral signals and acts through several orexigenic and anorexigenic mediators. We studied this regulating system in a rat model of methyl donor deficiency during gestation and lactation. At weaning, a predominance of the anorexigenic pathway was observed in deficient pups, with increased plasma peptide YY and increased hypothalamic pro-opiomelanocortin (POMC) mRNA, in line with abnormal leptin, ghrelin, and insulin secretion and/or signaling during critical periods of fetal and/or postnatal development of the hypothalamus. These results suggest that early methyl donor deficiency can affect the development and function of energy balance circuits, resulting in growth and weight deficits. Maternal administration of folic acid (3 mg/kg/day) during the perinatal period tended to rectify peripheral metabolic signaling and central neuropeptide and receptor expression, leading to reduced growth retardation.

Developmental Impairments in a Rat Model of Methyl Donor Deficiency: Effects of a Late Maternal Supplementation with Folic Acid.

Vitamins B9 (folate) and B12 act as methyl donors in the one-carbon metabolism which influences epigenetic mechanisms. We previously showed that an embryofetal deficiency of vitamins B9 and B12 in the rat increased brain expression of let-7a and miR-34a microRNAs involved in the developmental control of gene expression. This was reversed by the maternal supply with folic acid (3 mg/kg/day) during the last third of gestation, resulting in a significant reduction of associated birth defects. Since the postnatal brain is subject to intensive developmental processes, we tested whether further folate supplementation during lactation could bring additional benefits. Vitamin deficiency resulted in weaned pups (21 days) in growth retardation, delayed ossification, brain atrophy and cognitive deficits, along with unchanged brain level of let-7a and decreased expression of miR-34a and miR-23a. Whereas maternal folic acid supplementation helped restore the levels of affected microRNAs, it led to a reduction of structural and functional defects taking place during the perinatal/postnatal periods, such as learning/memory capacities. Our data suggest that a gestational B-vitamin deficiency could affect the temporal control of the microRNA regulation required for normal development. Moreover, they also point out that the continuation of folate supplementation after birth may help to ameliorate neurological symptoms commonly associated with developmental deficiencies in folate and B12.

N-homocysteinylation of tau and MAP1 is increased in autopsy specimens of Alzheimer's disease and vascular dementia.

The pathomechanisms that associate a deficit in folate and/or vitamin B12 and the subsequent hyperhomocysteinemia with pathological brain ageing are unclear. We investigated the homocysteinylation of microtubule-associated proteins (MAPs) in brains of patients with Alzheimer's disease or vascular dementia, and in rats depleted in folate and vitamin B12, Cd320 KO mice with selective B12 brain deficiency and H19-7 neuroprogenitors lacking folate. Compared with controls, N-homocysteinylated tau and MAP1 were increased and accumulated in protein aggregates and tangles in the cortex, hippocampus and cerebellum of patients and animals. N-homocysteinylation dissociated tau and MAPs from ß-tubulin, and MS analysis showed that it targets lysine residues critical for their binding to ß-tubulin. N-homocysteinylation increased in rats exposed to vitamin B12 and folate deficit during gestation and lactation and remained significantly higher when they became 450 days-old, despite returning to normal diet at weaning, compared with controls. It was correlated with plasma homocysteine (Hcy) and brain expression of methionine tRNAsynthetase (MARS), the enzyme required for the synthesis of Hcy-thiolactone, the substrate of N-homocysteinylation. Experimental inactivation of MARS prevented the N-homocysteinylation of tau and MAP1, and the dissociation of tau and MAP1 from ß-tubulin and PSD95 in cultured neuroprogenitors. In conclusion, increased N-homocysteinylation of tau and MAP1 is a mechanism of brain ageing that depends on Hcy concentration and expression of MARS enzyme. Its irreversibility and cumulative occurrence throughout life may explain why B12 and folate supplementation of the elderly has limited effects, if any, to prevent pathological brain ageing and cognitive decline. Copyright © 2019 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Late Maternal Folate Supplementation Rescues from Methyl Donor Deficiency-Associated Brain Defects by Restoring Let-7 and miR-34 Pathways.

The micronutrients folate and vitamin B12 are essential for the proper development of the central nervous system, and their deficiency during pregnancy has been associated with a wide range of disorders. They act as methyl donors in the one-carbon metabolism which critically influences epigenetic mechanisms. In order to depict further underlying mechanisms, we investigated the role of let-7 and miR-34, two microRNAs regulated by methylation, on a rat model of maternal deficiency. In several countries, public health policies recommend periconceptional supplementation with folic acid. However, the question about the duration and periodicity of supplementation remains. We therefore tested maternal supply (3 mg/kg/day) during the last third of gestation from embryonic days (E) 13 to 20. Methyl donor deficiency-related developmental disorders at E20, including cerebellar and interhemispheric suture defects and atrophy of selective cerebral layers, were associated with increased brain expression (by 2.5-fold) of let-7a and miR-34a, with subsequent downregulation of their regulatory targets such as Trim71 and Notch signaling partners, respectively. These processes could be reversed by siRNA strategy in differentiating neuroprogenitors lacking folate, with improvement of their morphological characteristics. While folic acid supplementation helped restoring the levels of let-7a and miR-34a and their downstream targets, it led to a reduction of structural and functional defects taking place during the perinatal period. Our data outline the potential role of let-7 and miR-34 and their related signaling pathways in the developmental defects following gestational methyl donor deficiency and support the likely usefulness of late folate supplementation in at risk women.

Nutritional models of foetal programming and nutrigenomic and epigenomic dysregulations of fatty acid metabolism in the liver and heart.

Barker's concept of 'foetal programming' proposes that intrauterine growth restriction (IUGR) predicts complex metabolic diseases through relationships that may be further modified by the postnatal environment. Dietary restriction and deficit in methyl donors, folate, vitamin B12, and choline are used as experimental conditions of foetal programming as they lead to IUGR and decreased birth weight. Overfeeding and deficit in methyl donors increase central fat mass and lead to a dramatic increase of plasma free fatty acids (FFA) in offspring. Conversely, supplementing the mothers under protein restriction with folic acid reverses metabolic and epigenomic phenotypes of offspring. High-fat diet or methyl donor deficiency (MDD) during pregnancy and lactation produce liver steatosis and myocardium hypertrophy that result from increased import of FFA and impaired fatty acid ß-oxidation, respectively. The underlying molecular mechanisms show dysregulations related with similar decreased expression and activity of sirtuin 1 (SIRT1) and hyperacetylation of peroxisome proliferator-activated receptor-γ coactivator 1α (PGC-1α). High-fat diet and overfeeding impair AMPK-dependent phosphorylation of PGC-1α, while MDD decreases PGC-1α methylation through decreased expression of PRMT1 and cellular level of S-adenosyl methionine. The visceral manifestations of metabolic syndrome are under the influence of endoplasmic reticulum (ER) stress in overnourished animal models. These mechanisms should also deserve attention in the foetal programming effects of MDD since vitamin B12 influences ER stress through impaired SIRT1 deacetylation of HSF1. Taken together, similarities and synergies of high-fat diet and MDD suggest, therefore, considering their consecutive or contemporary influence in the mechanisms of complex metabolic diseases.

Folate and fetal programming: a play in epigenomics?

Folate plays a key role in the interactions between nutrition, fetal programming, and epigenomics. Maternal folate status influences DNA methylation, inheritance of the agouti phenotype, expression of imprinting genes, and the effects of mycotoxin FB1 on heterochromatin assembly in rodent offspring. Deficiency in folate and other methyl donors increases birth defects and produces visceral manifestations of fetal programming, including liver and heart steatosis, through imbalanced methylation and acetylation of PGC1-α and decreased SIRT1 expression, and produces persistent cognitive and learning disabilities through impaired plasticity and hippocampal atrophy. Maternal folate supplementation also produces long-term epigenomic effects in offspring, some beneficial and others negative. Deciphering these mechanisms will help understanding the discordances between experimental models and population studies of folate deficiency and supplementation.

[Folates and fetal programming: role of epigenetics and epigenomics]. / Folates et programmation foetale: rôle des mécanismes nutrigénomiques et épigénomiques.

Folates are needed for synthesis of methionine, the precursor of S-adenosyl methionine (SAM). They play therefore a key role in nutrition and epigenomics by fluxing monocarbons towards synthesis or methylation of DNA and RNA, and methylation of gene transregulators, respectively. The deficiency produces intrauterine growth retardation and birth dejects. Folate deficiency deregulates epigenomic mechanisms related to fetal programming through decreased cellular availability of SAM. Epigenetic mechanisms of folate deficiency are illustrated by inheritance of coat colour of agouti mice model and altered expression of Igf2/H19 imprinting genes. Dietary exposure to fumonisin FB1 acts synergistically with folate deficiency on alterations of heterochromatin assembly. Deficiency in folate and vitamin B12 produces impaired fatty acid oxidation in liver and heart through imbalanced methylation and acetylation of PGC1-alpha and decreased expression of SIRT1, and long-lasting cognitive disabilities through impaired hippocampal cell proliferation, differentiation and plasticity and atrophy of hippocampal CA1. Deciphering these mechanisms will help understand the discordances between experimental models and population studies on folate supplementation.