Folate-deficient Diet Research Articles

Fumonisin B1induced neural tube defects were not increased in LM/Bc mice fed folate-deficient diet

Fumonisin B1 (FB1 ) is found in corn-based foods and is a possible risk factor for neural tube defects (NTDs). The mechanism(s) underlying NTD induction by FB1 in LM/Bc mice is not understood; however, evidence suggests disrupted folate transport is involved. Female LM/Bc mice were fed folate-sufficient (control) or folate-deficient diet beginning 5 wk before mating, treated with 0 (vehicle), 2.5 or 10 mg/kg FB1 by intraperitoneal injection on embryonic days 7 (E7) and E8, and their fetuses examined on E16. Dose-dependent NTD induction was found in groups fed the control diet: 3 of 13 low-dose and 10 of 11 high-dose litters were affected. Among groups fed folate-deficient diet, NTDs were found only in 4 of 11 high-dose litters. In another trial, consumption of folate-deficient diet also resulted in fewer NTDs at a dose of 10 mg/kg FB1 and reduced maternal red blood cell folate levels by 80%. In utero death did not fully account for the differences in NTD rates. Folate deficiency does not exacerbate NTD induction by FB1 in LM/Bc mice. Interactions between folate, other nutritional factors, and FB1 in this mouse model for NTDs are complex and require further investigation.

Maternal micronutrients, omega-3 fatty acids, and placental PPARγ expression.

An altered one-carbon cycle is known to influence placental and fetal development. We hypothesize that deficiency of maternal micronutrients such as folic acid and vitamin B12 will lead to increased oxidative stress, reduced long-chain polyunsaturated fatty acids, and altered expression of peroxisome proliferator activated receptor (PPARγ) in the placenta, and omega-3 fatty acid supplementation to these diets will increase the expression of PPARγ. Female rats were divided into 5 groups: control, folic acid deficient, vitamin B12 deficient, folic acid deficient + omega-3 fatty acid supplemented, and vitamin B12 deficient + omega-3 fatty acid supplemented. Dams were dissected on gestational day 20. Maternal micronutrient deficiency leads to lower (p < 0.05) levels of placental docosahexaenoic acid, arachidonic acid, PPARγ expression and higher (p < 0.05) levels of plasma malonidialdehyde, placental IL-6, and TNF-α. Omega-3 fatty acid supplementation to a vitamin B12 deficient diet normalized the expression of PPARγ and lowered the levels of placental TNF-α. In the case of supplementation to a folic acid deficient diet it lowered the levels of malonidialdehyde and placental IL-6 and TNF-α. This study has implications for fetal growth as oxidative stress, inflammation, and PPARγ are known to play a key role in the placental development.

A Study of the Effect of Isolated Folic Acid Deficiency on Lens in Adolescent Rats

Background and Objectives: India accounts for 20% of the world's blind population and 90% of the reversible blind people are from developing nations. Micronutrients like folic acid have been linked to cataract and 33% of the Indian population suffers from folate deficiency. This study was done to find evidence of cataractogenesis in the younger age group of folate-deficient people. Methods: After taking animal ethical clearance, 12 Wistar albino rats weighing 120–150 grams and corresponding to 18–20 years of human age, were fed folate-deficient diet for 45 days and sacrificed on the 46th day. The enucleated eyeballs were subjected to histopathological examination, under light microscopy, for any lens abnormality. Results: The study stated that the lens capsule epithelium and the arrangement and regularity of the lens fibers were normal. There was no evidence of thickening, degeneration, swelling or disruption of the lens fibers. Conclusion: It was inferred that the lens cells could store folic acid as intracellular folylpolyglutamate or bound to internalized receptor proteins. Folic acid is essential in the synthesis of an important antioxidant Glutathione from glutamic acid, cysteine and glycine. Glutathione in turn protects the cells from oxidative damage incurred during normal metabolic and optical activities in the eye lens.

Low paternal dietary folate alters the mouse sperm epigenome and is associated with negative pregnancy outcomes

Epidemiological studies suggest that a father’s diet can influence offspring health. A proposed mechanism for paternal transmission of environmental information is via the sperm epigenome. The epigenome includes heritable information such as DNA methylation. We hypothesize that the dietary supply of methyl donors will alter epigenetic reprogramming in sperm. Here we feed male mice either a folate-deficient or folate-sufficient diet throughout life. Paternal folate deficiency is associated with increased birth defects in the offspring, which include craniofacial and musculoskeletal malformations. Genome-wide DNA methylation analysis and the subsequent functional analysis identify differential methylation in sperm of genes implicated in development, chronic diseases such as cancer, diabetes, autism and schizophrenia. While >300 genes are differentially expressed in offspring placenta, only two correspond to genes with differential methylation in sperm. This model suggests epigenetic transmission may involve sperm histone H3 methylation or DNA methylation and that adequate paternal dietary folate is essential for offspring health.

Myasthenia gravis: Newer therapies offer sustained improvement

Myasthenia gravis is a prototypical antibody-mediated autoimmune neuromuscular disorder. Treatments have improved over the past 30 years, leading to significantly fewer deaths and better quality of life. Future research should further elucidate its pathogenesis, reveal better ways to diagnose it, and yield new treatments.

Genes with Aberrant Expression in Murine Preneoplastic Intestine Show Epigenetic and Expression Changes in Normal Mucosa of Colon Cancer Patients

An understanding of early genetic/epigenetic changes in colorectal cancer would aid in diagnosis and prognosis. To identify these changes in human preneoplastic tissue, we first studied our mouse model in which Mthfr⁺/⁻ BALB/c mice fed folate-deficient diets develop intestinal tumors in contrast to Mthfr⁺/⁺ BALB/c mice fed control diets. Transcriptome profiling was performed in normal intestine from mice with low or high tumor susceptibility. We identified 12 upregulated and 51 downregulated genes in tumor-prone mice. Affected pathways included retinoid acid synthesis, lipid and glucose metabolism, apoptosis and inflammation. We compared murine candidates from this microarray analysis, and murine candidates from an earlier strain-based comparison, with a set of human genes that we had identified in previous methylome profiling of normal human colonic mucosa, from colorectal cancer patients and controls. From the extensive list of human methylome candidates, our approach uncovered five orthologous genes that had shown changes in murine expression profiles (PDK4, SPRR1A, SPRR2A, NR1H4, and PYCARD). The human orthologs were assayed by bisulfite-pyrosequencing for methylation at 14 CpGs. All CpGs exhibited significant methylation differences in normal mucosa between colorectal cancer patients and controls; expression differences for these genes were also observed. PYCARD and NR1H4 methylation differences showed promise as markers for presence of polyps in controls. We conclude that common pathways are disturbed in preneoplastic intestine in our animal model and morphologically normal mucosa of patients with colorectal cancer, and present an initial version of a DNA methylation-based signature for human preneoplastic colon.

Maternal folic acid–deficient diet causes congenital malformations in the mouse eye

The eye is a very complex structure derived from the neural tube, surface ectoderm, and migratory mesenchyme from a neural crest origin. Because structures that evolve from the neural tube may be affected by a folate/folic acid (FA) deficiency, the aim of this work was to investigate whether a maternal folic acid-deficient diet may cause developmental alterations in the mouse eye. Female C57BL/6J mice (8 weeks old) were assigned into two different folic acid groups for periods ranging between 2 and 16 weeks. Animals were killed at gestation day 17. Hepatic folate was analyzed, and the eyes from 287 fetuses were macroscopically studied, sectioned and immunolabeled with anti-transforming growth factor (TGF)-β2 and anti-TGF-βRII. Mice exposed to a FA-deficient diet exhibited numerous eye macroscopic anomalies, such as anophthalmia and microphthalmia. Microscopically, the eye was the most affected organ (43.7% of the fetuses). The highest incidence of malformations occurred from the 8th week onward. A statistically significant linear association between the number of maternal weeks on the FA-deficient diet and embryonic microscopic eye malformations was observed. The optic cup derivatives and structures forming the eye anterior segment showed severe abnormalities. In addition, TGF-β2 and TGF-βRII expression in the eye was also altered. This study suggests that an adequate folic acid/folate status plays a key role in the formation of ocular tissues and structures, whereas a vitamin deficiency is negatively associated with a normal eye development even after a short-term exposure.

Dietary and genetic manipulations of folate metabolism differentially affect neocortical functions in mice

Converging evidence suggests that folate-mediated one-carbon metabolism may modulate cognitive functioning throughout the lifespan, but few studies have directly tested this hypothesis. This study examined the separate and combined effects of dietary and genetic manipulations of folate metabolism on neocortical functions in mice, modeling a common genetic variant in the MTHFD1 gene in humans. Mutant (Mthfd1gt/+) and wildtype (WT) male mice were assigned to a folate sufficient or deficient diet at weaning and continued on these diets throughout testing on a series of visual attention tasks adapted from the 5-choice serial reaction time task. WT mice on a deficient diet exhibited impulsive responding immediately following a change in task parameters that increased demands on attention and impulse control, and on trials following an error. This pattern of findings indicates a heightened affective response to stress and/or an inability to regulate negative emotions. In contrast, Mthfd1gt/+ mice (regardless of diet) exhibited attentional dysfunction and a blunted affective response to committing an error. The Mthfd1gt/+ mice also showed significantly decreased expression levels for genes encoding choline dehydrogenase and the alpha 7 nicotinic cholinergic receptor. The effects of the MTHFD1 mutation were less pronounced when combined with a deficient diet, suggesting a compensatory mechanism to the combined genetic and dietary perturbation of folate metabolism. These data demonstrate that common alterations in folate metabolism can produce functionally distinct cognitive and affective changes, and highlight the importance of considering genotype when making dietary folate recommendations.

Reduced nerve growth factor levels in stress-related brain regions of folate-deficient mice

Folate deficiency has been linked to neurodegenerative and stress-related diseases such as stroke, dementia and depression. The role of the neurotrophins nerve growth factor (NGF) and neurotrophin-3 (NT-3) in stress-related disorders and neurodegeneration has garnered increasing attention in recent years. Uracil misincorporation is involved in the neuropsychiatric dysfunction induced by experimental folate deprivation. However, the effects of folate deficiency on the expression of NGF and NT-3 in brain tissue have not yet been investigated. In a 2×2 design, aged mice lacking uracil-DNA N-glycosylase (Ung−/−) versus wild-type (Ung+/+) controls were subjected to a folate-deficient diet versus a regular diet for three months. Independent of genotype, folate deficiency led to decreased NGF protein levels in the frontal cortex and amygdala. In the hippocampus, NGF levels were increased in UNG−/− mice on the normal diet, but not under folate deficiency, while in UNG+/+ mice, folate deprivation did not affect hippocampal NGF content. NT-3 protein concentrations were neither affected by genotype nor by folate deficiency. Altogether, the results of our study show that folate deficiency affects NGF levels in the frontal cortex, amygdala and hippocampus. The decrease in NGF content in the hippocampus in response to folate deficiency in Ung−/− mice may contribute to their phenotype of enhanced anxiety and despair-like behavior as well as to selective hippocampal neurodegeneration.

Preventable decrements of cerebral microvascular volume and vasodilatation in a rat model of diet‐induced Vascular Cognitive Impairment: non‐invasive detection by absolute Near Infrared Spectroscopy

The brain microvascular circulation has considerable structural plasticity and can expand or regress in response to metabolic and dietary challenges We hypothesized that such changes might underlie the association of folate deficiency and hyperhomocysteinemia with cerebrovascular and neurodegenerative disease. Previously, we found that dietary folate deficiency can impair cognition in Sprague Dawley rats and that this can be mitigated by adding methionine to the folate‐deficient diet. Separately, we used non‐invasive absolute Near Infrared Spectroscopy (NIRS) to obtain evidence of brain microvascular changes in folate‐deficient rats, finding diminished cerebral blood volume and vasoreactivity compared to controls. Here we extend these findings by showing that folate deficiency reproducibly diminishes cerebral blood volume and vascular reactivity, and that these changes can be reversed by methionine supplementation. Our findings are consistent with the hypothesis that reversible microvascular changes may contribute to cognitive impairment. The ability to non‐invasively manipulate, detect and measure such changes can help to understand the role of microvascular plasticity in dementia.Support: Israel Science Foundation and EU‐IRG

Strain‐dependent dysregulation of one‐carbon metabolism in male mice is associated with choline‐ and folate‐deficient diet‐induced liver injury

Dysregulation of one-carbon metabolism-related metabolic processes is a major contributor to the pathogenesis of nonalcoholic fatty liver disease (NAFLD). It is well established that genetic and gender-specific variations in one-carbon metabolism contribute to the vulnerability to NAFLD in humans. To examine the role of one-carbon metabolism dysregulation in the pathogenesis and individual susceptibility to NAFLD, we used a "population-based" mouse model where male mice from 7 inbred were fed a choline- and folate-deficient (CFD) diet for 12 wk. Strain-dependent down-regulation of several key one-carbon metabolism genes, including methionine adenosyltransferase 1α (Mat1a), cystathionine-β-synthase (Cbs), methylenetetrahydrofolate reductase (Mthfr), adenosyl-homocysteinase (Ahcy), and methylenetetrahydrofolate dehydrogenase 1 (Mthfd1), was observed. These changes were strongly associated with interstrain variability in liver injury (steatosis, necrosis, inflammation, and activation of fibrogenesis) and hyperhomocysteinemia. Mechanistically, the decreased expression of Mat1a, Ahcy, and Mthfd1 was linked to a reduced level and promoter binding of transcription factor CCAAT/enhancer binding protein β (CEBPβ), which directly regulates their transcription. The strain specificity of diet-induced dysregulation of one-carbon metabolism suggests that interstrain variation in the regulation of one-carbon metabolism may contribute to the differential vulnerability to NFLD and that correcting the imbalance may be considered as preventive and treatment strategies for NAFLD.

Reduced MTHFD1 Activity in Male Mice Perturbs Folate- and Choline-Dependent One-Carbon Metabolism as Well as Transsulfuration

Impaired utilization of folate is caused by insufficient dietary intake and/or genetic variation and has been shown to prompt changes in related pathways, including choline and methionine metabolism. These pathways have been shown to be sensitive to variation within the Mthfd1 gene, which codes for a folate-metabolizing enzyme responsible for generating 1-carbon (1-C)–substituted folate derivatives. The Mthfd1gt/+ mouse serves as a potential model of human Mthfd1 loss-of-function genetic variants that impair MTHFD1 function. This study investigated the effects of the Mthfd1gt/+ genotype and folate intake on markers of choline, folate, methionine, and transsulfuration metabolism. Male Mthfd1gt/+ and Mthfd1+/+ mice were randomly assigned at weaning (3 wk of age) to either a control (2 mg/kg folic acid) or folate-deficient (0 mg/kg folic acid) diet for 5 wk. Mice were killed at 8 wk of age following 12 h of food deprivation; blood and liver samples were analyzed for choline, methionine, and transsulfuration biomarkers. Independent of folate intake, mice with the Mthfd1gt/+ genotype had higher hepatic concentrations of choline (P = 0.005), betaine (P = 0.013), and dimethylglycine (P = 0.004) and lower hepatic concentrations of glycerophosphocholine (P = 0.002) relative to Mthfd1+/+ mice. Mthfd1gt/+ mice also had higher plasma concentrations of homocysteine (P = 0.0016) and cysteine (P < 0.001) as well as lower plasma concentrations of methionine (P = 0.0003) and cystathionine (P = 0.011). The metabolic alterations observed in Mthfd1gt/+ mice indicate perturbed choline and folate-dependent 1-C metabolism and support the future use of Mthfd1gt/+ mice as a tool to investigate the impact of impaired 1-C metabolism on disease outcomes.

Green Tea Is a Poor Contributor to Tissue Folate in a Folate Depletion-Repletion Rat Model

Green tea contains folate and epigallocatechin gallate (EGCg), which is suggested to be an antifolate. In this study, we examined whether green tea was a good source of folate in a folate depletion/repletion rat model. Rats fed a folate deficient diet for 4 weeks were supplied with folate for 7 days, and then folate repletion effects were evaluated in terms of increase in total folate level in plasma, liver, and bone marrow and decrease in plasma homocysteine level. In this model, the folic acid treatment effect was observed to be dose-dependent and an appropriate dose of folic acid was 40 μg/kg/day or greater. Based on this finding, green tea as well as spinach, chicken liver, and folic acid as a reference were applied to this model. Increase in tissue folate level in response to the food samples varied among tissues, with bone marrow showing the smallest response. Increase in tissue folate level was in the order of spinach > chicken liver > green tea, which produced only a slight increase in tissue folate level and further diminished bone marrow folate level. EGCg administered by intragastric gavage at an approximate dose of 8 mg/kg did not attenuate the increase in tissue folate level when repletion was performed with folic acid. These results suggested that green tea is a poor source of food folate, but EGCg in green tea at a low dose has little effect on folic acid absorption.

Deletion of Mthfd1l causes embryonic lethality and neural tube and craniofacial defects in mice

Maternal supplementation with folic acid is known to reduce the incidence of neural tube defects (NTDs) by as much as 70%. Despite the strong clinical link between folate and NTDs, the biochemical mechanisms through which folic acid acts during neural tube development remain undefined. The Mthfd1l gene encodes a mitochondrial monofunctional 10-formyl-tetrahydrofolate synthetase, termed MTHFD1L. This gene is expressed in adults and at all stages of mammalian embryogenesis with localized regions of higher expression along the neural tube, developing brain, craniofacial structures, limb buds, and tail bud. In both embryos and adults, MTHFD1L catalyzes the last step in the flow of one-carbon units from mitochondria to cytoplasm, producing formate from 10-formyl-THF. To investigate the role of mitochondrial formate production during embryonic development, we have analyzed Mthfd1l knockout mice. All embryos lacking Mthfd1l exhibit aberrant neural tube closure including craniorachischisis and exencephaly and/or a wavy neural tube. This fully penetrant folate-pathway mouse model does not require feeding a folate-deficient diet to cause this phenotype. Maternal supplementation with sodium formate decreases the incidence of NTDs and partially rescues the growth defect in embryos lacking Mthfd1l. These results reveal the critical role of mitochondrially derived formate in mammalian development, providing a mechanistic link between folic acid and NTDs. In light of previous studies linking a common splice variant in the human MTHFD1L gene with increased risk for NTDs, this mouse model provides a powerful system to help elucidate the specific metabolic mechanisms that underlie folate-associated birth defects, including NTDs.

Folate Deficiency Is Associated With Oxidative Stress, Increased Blood Pressure, and Insulin Resistance in Spontaneously Hypertensive Rats

The role of folate deficiency and associated hyperhomocysteinemia in the pathogenesis of metabolic syndrome is not fully established. In the current study, we analyzed the role of folate deficiency in pathogenesis of the metabolic syndrome in the spontaneously hypertensive rat (SHR). Metabolic and hemodynamic traits were assessed in SHR/Ola rats fed either folate-deficient or control diet for 4 weeks starting at the age of 3 months. Compared to SHRs fed a folate-replete diet, SHRs fed a folate-deficient diet showed significantly reduced serum folate (104 ± 5 vs. 11 ± 1 nmol/L, P < 0.0005) and urinary folate excretion (4.3 ± 0.6 vs. 1.2 ± 0.1 nmol/16 h, P < 0.0005) together with a near 3-fold increase in plasma total homocysteine concentration (4.5 ± 0.1 vs 13.1 ± 0.7 μmol/L, P < 0.0005), ectopic fat accumulation in liver, and impaired glucose tolerance. Folate deficiency also increased systolic blood pressure by approximately 15 mm Hg (P < 0.01). In addition, the low-folate diet was accompanied by significantly reduced activity of antioxidant enzymes and increased concentrations of lipoperoxidation products in liver, renal cortex, and heart. These findings demonstrate that the SHR model is susceptible to the adverse metabolic and hemodynamic effects of low dietary intake of folate. The results are consistent with the hypothesis that folate deficiency can promote oxidative stress and multiple features of the metabolic syndrome that are associated with increased risk for diabetes and cardiovascular disease.

Dietary Folate Deficiency in Pseudopregnant Mice Has No Effect on Homeobox A10 Promoter Methylation or Expression

During the reproductive cycle, a number of genes controlling endometrial changes are regulated by DNA methylation, a common epigenetic modification. Because dietary folate affects DNA methylation, we determined whether a folate-deficient diet (FDD) alters DNA methylation in endometria of pseudopregnant mice, focusing on the homeobox A10 (Hoxa10) promoter. Mice were given an FDD or control diet for 40 to 45 days and examined on day 5 of pseudopregnancy. Compared to control mice, FDD mice had lower folate levels in liver and serum (P = .004). However, the FDD did not significantly affect DNA methylation within the cytosine-guanine dinucleotide (CpG)-rich Hoxa10 promoter, even when specific CpG sites were examined (P > .05). In endometrial tissue sections, the localization of anti-Hoxa10 staining was unchanged in FDD mice. Therefore, folate deficiency did not significantly affect promoter methylation or expression of Hoxa10.

Low dietary folate and methylenetetrahydrofolate reductase deficiency may lead to pregnancy complications through modulation of ApoAI and IFN‐γ in spleen and placenta, and through reduction of methylation potential

Genetic or nutritional disturbances in folate metabolism lead to hyperhomocysteinemia and adverse reproductive outcomes. Folate-dependent homocysteine remethylation is required for methylation reactions and may influence choline/betaine metabolism. Hyperhomocysteinemia has been suggested to play a role in inflammation. The goal of this study was to determine whether folate-related pregnancy complications could be due to altered expression of some inflammatory mediators or due to disturbances in methylation intermediates. Pregnant mice with or without a deficiency of methylenetetrahydrofolate reductase (MTHFR) were fed control diets or folate-deficient (FD) diets; tissues were collected at embryonic day 14.5. FD decreased plasma phosphocholine and increased plasma glycerophosphocholine and lysophosphatidylcholine. Liver betaine, phosphocholine, and S-adenosylmethionine:S-adenosylhomocysteine ratios were reduced in FD. In liver, spleen, and placenta, the lowest levels of apolipoprotein AI (ApoAI) were observed in Mthfr(+/-) mice fed FD. Increased interferon-gamma (IFN-γ) was observed in spleen and placentae due to FD or Mthfr genotype. Plasma homocysteine correlated negatively with liver and spleen ApoAI, and positively with IFN-γ. Low dietary folate or Mthfr deficiency during pregnancy may result in adverse pregnancy outcomes by altering expression of the inflammatory mediators ApoAI and IFN-γ in spleen and placenta. Disturbances in choline metabolism or methylation reactions may also play a role.

Arsenic urinary speciation in Mthfr deficient mice injected with sodium arsenate

In most mammalian species, arsenic biotransformation occurs primarily by biomethylation and reduction reactions, with dimethylarsinic acid being the predominant metabolite excreted in the urine. Methylenetetrahydrofolate reductase (Mthfr) plays a key role in folate metabolism by channeling one-carbon units between nucleotide synthesis and methylation reactions. In the study on transgenic Mtfhr knockout mice we investigated: (1) whether Mthfr is an important determinant in arsenic biotransformation by performing urinary arsenic speciation, and (2) whether dietary folate deficiency alters arsenic biotransformation in these mice. The Mthfr mice fed folate replete or folate deficient diet were injected with sodium arsenate 1mg/kg, and placed in metabolic cages for a urine collection. The urine was analyzed for arsenic species. Additionally, folate and homocysteine plasma level was analyzed in Mthfr mice. When fed a folate control diet, the Mthfr−/− mice excreted significantly less of the total arsenic in urine than did the Mthfr+/+ and Mthfr+/− mice. The Mthfr−/− had significantly lower levels of pentavalent arsenic in their urine than did the Mthfr+/+mice. The wild type mice excreted significantly less pentavalent arsenic when they were fed folate deficient diet comparing to control diet. The current data suggest that both the Mthfr status and food folate level modulate in a significant manner excretion of arsenic in mice, following intraperitoneal administration of sodium arsenate.

Interstrain differences in the severity of liver injury induced by a choline‐ and folate‐deficient diet in mice are associated with dysregulation of genes involved in lipid metabolism

Nonalcoholic fatty liver disease (NAFLD) is a major health problem and a leading cause of chronic liver disease in the United States and developed countries. In humans, genetic factors greatly influence individual susceptibility to NAFLD. The goals of this study were to compare the magnitude of interindividual differences in the severity of liver injury induced by methyl-donor deficiency among individual inbred strains of mice and to investigate the underlying mechanisms associated with the variability. Feeding mice a choline- and folate-deficient diet for 12 wk caused liver injury similar to NAFLD. The magnitude of liver injury varied among the strains, with the order of sensitivity being A/J ≈ C57BL/6J ≈ C3H/HeJ < 129S1/SvImJ ≈ CAST/EiJ < PWK/PhJ < WSB/EiJ. The interstrain variability in severity of NAFLD liver damage was associated with dysregulation of genes involved in lipid metabolism, primarily with a down-regulation of the peroxisome proliferator receptor α (PPARα)-regulated lipid catabolic pathway genes. Markers of oxidative stress and oxidative stress-induced DNA damage were also elevated in the livers but were not correlated with severity of liver damage. These findings suggest that the PPARα-regulated metabolism network is one of the key mechanisms determining interstrain susceptibility and severity of NAFLD in mice.

Folate Deficiency Alters the Sperm Epigenome and Reproductive Outcomes.

Data from epidemiological and animal studies suggests that the paternal epigenetic contribution in sperm may serve as a route to altered offspring phenotypes. However to date there is little to no evidence of direct effects of diets on the sperm epigenome or consequences for the offspring. Epigenetics refers to heritable changes in gene expression that are regulated by DNA methylation and the covalent modification of histones. Given that dietary folate can alter the supply of methyl donors, we hypothesized that male germ cells may be sensitive to dietary folate and that this may manifest as altered epigenetic programming and have downstream consequences for spermatogenesis and reproductive outcomes. The objective of this study was to determine the impact of exposure to a low folate diet, beginning during embryonic development through to adulthood on spermatogenesis, the sperm epigenome, fertility, and offspring. Inbred C57/BL6 mice were fed either a folate-sufficient (FS, 2 mg of folate/kg of diet), or a folate-deficient diet (FD, 0.3 mg of folate/kg of diet). Adult FS (n=54) and FD (n=49) males remained on the diets for either 2 or 4-4.5 months and were assessed for fertility parameters and overall health. In adults, spermatogenesis and sperm counts were not altered by diet nor were body, testis and epididymis weights. Epigenetic programming was disturbed in the sperm of folate depleted mice. Histone H3 methylation was significantly reduced in sperm from FD versus FS mice. Notably histone H3 methylation has been shown to mark genes for embryonic development. Genome wide DNA methylation (MeDIP-chip) analysis revealed alterations at more than 50 gene promoter regions. Bisulfite sequencing confirmed that 6 of these regions including genes implicated in developmental processes had significant decreases in DNA methylation at specific CpG locations in the sperm of FD males (n=5 for both groups). FD (n=29) and FS (n=27) males were then mated to females on control diets and pregnancy outcomes at embryonic day 18.5 were analyzed. Embryonic loss was 2 fold greater in the litters sired by FD vs FS mice and developmental abnormalities were observed in 27% of the litters sired by FD mice vs 3% for those sired by FS mice. This is the first study to show that diet can alter the sperm epigenome and suggests that an altered sperm epigenome leads to negative pregnancy outcomes. This research was supported by NSERC, and the Réseau Québécois en Reproduction (RQR-FQRNT).