Metabolism Methylation Research Articles

Methyl Group Metabolism in Differentiation, Aging, and Cancer.

Methyl group metabolism belongs to a relatively understudied field of research. Its importance lies in the fact that methyl group metabolic pathways are crucial for the successful conversion of dietary nutrients into the basic building blocks to carry out any cellular methylation reaction. Methyl groups play essential roles in numerous cellular functions such as DNA methylation, nucleotide- and protein biosynthesis. Especially, DNA methylation is responsible for organizing the genome into transcriptionally silent and active regions. Ultimately, it is this proper annotation that determines the quality of expression patterns required to ensure and shape the phenotypic integrity and function of a highly specialized cell type. Life is characterized by constantly changing environmental conditions, which are addressed by changes in DNA methylation. This relationship is increasingly coming into focus as it is of fundamental importance for differentiation, aging, and cancer. The stability and permanence of these metabolic processes, fueling the supplementation of methyl groups, seem to be important criteria to prevent deficiencies and erosion of the methylome. Alterations in the metabolic processes can lead to epigenetic and genetic perturbations, causative for diverse disorders, accelerated aging, and various age-related diseases. In recent decades, the intake of methyl group compounds has changed significantly due to, e.g., environmental pollution and food additives. Based on the current knowledge, this review provides a brief overview of the highly interconnected relationship between nutrition, metabolism, changes in epigenetic modifications, cancer, and aging. One goal is to provide an impetus to additionally investigate changes in DNA methylation as a possible consequence of an impaired methyl group metabolism.

Lipid metabolism and m6A RNA methylation are altered in lambs supplemented rumen-protected methionine and lysine in a low-protein diet

BackgroundMethionine or lysine has been reported to influence DNA methylation and fat metabolism, but their combined effects in N6-methyl-adenosine (m6A) RNA methylation remain unclarified. The combined effects of rumen-protected methionine and lysine (RML) in a low-protein (LP) diet on lipid metabolism, m6A RNA methylation, and fatty acid (FA) profiles in the liver and muscle of lambs were investigated. Sixty-three male lambs were divided into three treatment groups, three pens per group and seven lambs per pen. The lambs were fed a 14.5% crude protein (CP) diet (adequate protein [NP]), 12.5% CP diet (LP), and a LP diet plus RML (LP + RML) for 60 d.ResultsThe results showed that the addition of RML in a LP diet tended to lower the concentrations of plasma leptin (P = 0.07), triglyceride (P = 0.05), and non-esterified FA (P = 0.08). Feeding a LP diet increased the enzyme activity or mRNA expression of lipogenic enzymes and decreased lipolytic enzymes compared with the NP diet. This effect was reversed by supplementation of RML with a LP diet. The inclusion of RML in a LP diet affected the polyunsaturated fatty acids (PUFA), n-3 PUFA, and n-6 PUFA in the liver but not in the muscle, which might be linked with altered expression of FA desaturase-1 (FADS1) and acetyl-CoA carboxylase (ACC). A LP diet supplemented with RML increased (P < 0.05) total m6A levels in the liver and muscle and were accompanied by decreased expression of fat mass and obesity-associated protein (FTO) and alkB homologue 5 (ALKBH5). The mRNA expressions of methyltransferase-like 3 (METTL3) and methyltransferase-like 14 (METTL14) in the LP + RML diet group were lower than those in the other two groups. Supplementation of RML with a LP diet affected only liver YTH domain family (YTHDF2) proteins (P < 0.05) and muscle YTHDF3 (P = 0.09), which can be explained by limited m6A-binding proteins that were mediated in mRNA fate.ConclusionsOur findings showed that the inclusion of RML in a LP diet could alter fat deposition through modulations of lipogenesis and lipolysis in the liver and muscle. These changes in fat metabolism may be associated with the modification of m6A RNA methylation.Graphical abstractA systematic graph illustrates the mechanism of dietary methionine and lysine influence on lipid metabolism and M6A. The green arrow with triangular heads indicates as activation and brown-wine arrows with flat heads indicates as suppression.

Comparative Proteomic Analyses of Poorly Motile Swamp Buffalo Spermatozoa Reveal Low Energy Metabolism and Deficiencies in Motility-Related Proteins.

Simple SummaryProteomic analysis of normal and poorly motile buffalo sperm indicates that the formation of poorly motile buffalo sperm is associated with inefficient energy metabolism, decreased sperm protamine proteins, a lack of motility-related proteins, and changes in tail structural proteins. The results provide clues for finding molecular mechanisms for poor motility of buffalo sperm.The acquisition of mammalian sperm motility is a main indicator of epididymal sperm maturation and helps ensure fertilization. Poor sperm motility will prevent sperm cells from reaching the fertilization site, resulting in fertilization failure. To investigate the proteomic profiling of normal and poorly motile buffalo spermatozoa, a strategy applying liquid chromatography tandem mass spectrometry combined with tandem mass targeting was used. As a result, 145 differentially expressed proteins (DEPs) were identified in poorly motile spermatozoa (fold change > 1.5), including 52 upregulated and 93 downregulated proteins. The upregulated DEPs were mainly involved in morphogenesis and regulation of cell differentiation. The downregulated DEPs were involved with transport, oxidation-reduction, sperm motility, regulation of cAMP metabolism and regulation of DNA methylation. The mRNA and protein levels of PRM1 and AKAP3 were lower in poorly motile spermatozoa, while the expressions of SDC2, TEKT3 and IDH1 were not correlated with motility, indicating that their protein changes were affected by transcription or translation. Such changes in the expression of these proteins suggest that the formation of poorly motile buffalo spermatozoa reflects a low efficiency of energy metabolism, decreases in sperm protamine proteins, deficiencies in motility-related proteins, and variations in tail structural proteins. Such proteins could be biomarkers of poorly motile spermatozoa. These results illustrate some of the molecular mechanisms associated with poorly motile spermatozoa and provide clues for finding molecular markers of these pathways.

S-Adenosylmethionine-responsive cystathionine β-synthase modulates sulfur metabolism and redox balance in Mycobacterium tuberculosis.

Methionine and cysteine metabolisms are important for the survival and pathogenesis of Mycobacterium tuberculosis (Mtb). The transsulfuration pathway converts methionine to cysteine and represents an important link between antioxidant and methylation metabolism in diverse organisms. Using a combination of biochemistry and cryo–electron microscopy, we characterized the first enzyme of the transsulfuration pathway, cystathionine β-synthase (MtbCbs) in Mtb. We demonstrated that MtbCbs is a heme-less, pyridoxal-5′-phosphate–containing enzyme, allosterically activated by S-adenosylmethionine (SAM). The atomic model of MtbCbs in its native and SAM-bound conformations revealed a unique mode of SAM-dependent allosteric activation. Further, SAM stabilized MtbCbs by sterically occluding proteasomal degradation, which was crucial for supporting methionine and redox metabolism in Mtb. Genetic deficiency of MtbCbs reduced Mtb survival upon homocysteine overload in vitro, inside macrophages, and in mice coinfected with HIV. Thus, the MtbCbs-SAM axis constitutes an important mechanism of coordinating sulfur metabolism in Mtb.

MiR-143-Mediated Responses to Betaine Supplement Repress Lipogenesis and Hepatic Gluconeogenesis by Targeting MAT1a and MAPK11.

The liver as the central organ is responsible for lipogenesis, gluconeogenesis and one-carbon metabolism. Methyl donors (e.g., betaine) modulate metabolic homeostasis and gene regulation through one-carbon metabolism. MiR-143 regulates DNA methylation by targeting DNMT3A, thereby suggesting that this miRNA participates in one-carbon metabolic pathways. However, the effect and mechanism that regulate glucose and lipid metabolism via the methyl group metabolism pathway remain elusive. In this study, we found that a betaine supplement and miR-143 KO significantly promoted lipolysis and glucose utilization and repressed lipogenesis and gluconeogenesis through enhancing energy consumption and thermogenesis, repressing GPNMB and targeting MAPK11, respectively. We further explored the relationship between miR-143 and a methyl donor (betaine) and the miR-143-mediated responses to the betaine supplement regulating the mechanism of the glucose and lipid metabolism. The results showed that betaine significantly down-regulated the expression of miR-143 that subsequently increased SAM levels in the liver by targeting MAT1a. In brief, the regulations of glucose and lipid metabolism are related to the miR-143-regulation of one-carbon units, and the relationship between betaine and miR-143 in the methionine cycle is a typical yin-yang type of regulation. Thus, betaine and miR-143 function together as key regulators and biomarkers for preventing and diagnosing metabolic diseases such as fatty liver disease, obesity, and diabetes.

Analysis of S-Adenosylmethionine and S-Adenosylhomocysteine: Method Optimisation and Profiling in Healthy Adults upon Short-Term Dietary Intervention.

S-adenosylmethionine (SAM) is essential for methyl transfer reactions. All SAM is produced de novo via the methionine cycle. The demethylation of SAM produces S-adenosylhomocysteine (SAH), an inhibitor of methyltransferases and the precursor of homocysteine (Hcy). The measurement of SAM and SAH in plasma has value in the diagnosis of inborn errors of metabolism (IEM) and in research to assess methyl group homeostasis. The determination of SAM and SAH is complicated by the instability of SAM under neutral and alkaline conditions and the naturally low concentration of both SAM and SAH in plasma (nM range). Herein, we describe an optimised LC-MS/MS method for the determination of SAM and SAH in plasma, urine, and cells. The method is based on isotopic dilution and employs 20 µL of plasma or urine, or 500,000 cells, and has an instrumental running time of 5 min. The reference ranges for plasma SAM and SAH in a cohort of 33 healthy individuals (age: 19–60 years old; mean ± 2 SD) were 120 ± 36 nM and 21.5 ± 6.5 nM, respectively, in accordance with independent studies and diagnostic determinations. The method detected abnormal concentrations of SAM and SAH in patients with inborn errors of methyl group metabolism. Plasma and urinary SAM and SAH concentrations were determined for the first time in a randomised controlled trial of 53 healthy adult omnivores (age: 18–60 years old), before and after a 4 week intervention with a vegan or meat-rich diet, and revealed preserved variations of both metabolites and the SAM/SAH index.

Methyl Metabolism and the Clock: An Ancient Story With New Perspectives.

Methylation, that is, the transfer or synthesis of a –CH3 group onto a target molecule, is a pervasive biochemical modification found in organisms from bacteria to humans. In mammals, a complex metabolic pathway powered by the essential nutrients vitamin B9 and B12, methionine and choline, synthesizes S-adenosylmethionine, the methyl donor in the methylation of nucleic acids, proteins, fatty acids, and small molecules by over 200 substrate-specific methyltransferases described so far in humans. Methylations not only play a key role in scenarios for the origin and evolution of life, but they remain essential for the development and physiology of organisms alive today, and methylation deficiencies contribute to the etiology of many pathologies. The methylation of histones and DNA is important for circadian rhythms in many organisms, and global inhibition of methyl metabolism similarly affects biological rhythms in prokaryotes and eukaryotes. These observations, together with various pieces of evidence scattered in the literature on circadian gene expression and metabolism, indicate a close mutual interdependence between biological rhythms and methyl metabolism that may originate from prebiotic chemistry. This perspective first proposes an abiogenetic scenario for rhythmic methylations and then outlines mammalian methyl metabolism, before reanalyzing previously published data to draw a tentative map of its profound connections with the circadian clock.

Excess S-adenosylmethionine inhibits methylation via catabolism to adenine

The global dietary supplement market is valued at over USD 100 billion. One popular dietary supplement, S-adenosylmethionine, is marketed to improve joints, liver health and emotional well-being in the US since 1999, and has been a prescription drug in Europe to treat depression and arthritis since 1975, but recent studies questioned its efficacy. In our body, S-adenosylmethionine is critical for the methylation of nucleic acids, proteins and many other targets. The marketing of SAM implies that more S-adenosylmethionine is better since it would stimulate methylations and improve health. Previously, we have shown that methylation reactions regulate biological rhythms in many organisms. Here, using biological rhythms to assess the effects of exogenous S-adenosylmethionine, we reveal that excess S-adenosylmethionine disrupts rhythms and, rather than promoting methylation, is catabolized to adenine and methylthioadenosine, toxic methylation inhibitors. These findings further our understanding of methyl metabolism and question the safety of S-adenosylmethionine as a supplement.

Abstract MP45: Metabolomic Profiling Provides Novel Insights Into The Biochemical Mechanism Of Mental Stress-induced Ischemia

Introduction: Mental stress-induced myocardial ischemia (MSI) is a marker of future cardiovascular events. The underlying biochemical mechanisms of this cardiovascular condition are poorly understood. Our objective was to use metabolic profiling to identify biochemical changes associated with mental stress (MS) in patients with stable coronary artery disease (CAD) and investigate differential pathways that are involved in MSI. Methods: A total of 110 Individuals with stable CAD who participated in the Mental Stress Ischemia Prognosis Study (MIPS) underwent a standardized mental stress to provoke of myocardial ischemia, detected with single-photon emission tomography. Plasma samples were obtained before and after MS at 45 and 90 minutes and metabolomic profiling was performed using liquid chromatography/mass spectrometry using a confirmed 500 compounds. Metabolomic features and pathways associated with ischemia using metabolome-wide association studies and Mummichog, respectively. Results: The sample involved a subset case-control for (MSI+ vs MSI-). At 45 minutes and 90 minutes, there were significant stress-induced changes in several compounds. There was a significant upregulation of metabolites associated with lipid signaling et ( p = 0.025, Inosine diphosphate). A comparison between the MSI+ and MS- groups showed differential upregulation of DNA methylation metabolites ( p = .036, 5-Methylcytosine hydrochloride) at 45 minutes and microbiome-related metabolites at 90 minutes (P=0.041, N, N-dimethyl-1,4-phenylenediamine). Pathway analyses were consistent with the aforementioned findings. Conclusion: Upregulation of metabolic pathways involved in DNA methylation and microbial metabolism is differentially associated with MSI. These findings provide novel mechanistic insights into the association between acute mental stress and myocardial ischemia as a marker for cardiovascular vulnerability in patients with CAD

Untargeted 2D NMR Metabolomics of [13C-methyl]Methionine-Labeled Tumor Models Reveals the Non-DNA Methylome and Provides Clues to Methyl Metabolism Shift during Tumor Progression.

For more than a decade, DNA and histone methylations have been the focus of extensive work, although their relationship with methyl group metabolism was overlooked. Recently, it has emerged that epigenetic methylations are influenced by methyl donor nutrient availability, cellular levels of S-adenosyl-methionine (SAM), and cytoplasmic methyltransferase activities. SAM-dependent methyltransferases methylate a wide range of targets, from small molecules to proteins and nucleic acids. However, few investigations of the global methylome of tumors have been performed. Here, untargeted NMR metabolomics of two mouse tumor models labeled with [13C-methyl]methionine were used to search for the NMR-visible set of cellular methyl acceptors denoted the global methylome. Tumor models were B16 melanoma cell cultures and B16 melanoma tumors, which may be considered as two stages of B16 tumor development. Based on 2D 1H–13C NMR spectra and orthogonal partial least squares discriminant analysis of spectra, our study revealed markedly different global methylomes for melanoma models. The methylome of B16 melanoma cell cultures was dominated by histone methylations, whereas that of B16 melanoma tumors was dominated by cytoplasmic small-molecule methylations. Overall, the technique gave access to the non-DNA methylome. Comparison of tumor models also exhibiting differential expression of aerobic glycolysis provided clues to a methyl metabolism shift during tumor progression.

Effects of maternal feed intake restriction on the blood parameters, fatty acid profile and lipogenetic genes expression of perirenal fat in offspring kids

Maternal nutritional restriction impacted lipogenic gene expression in adipose tissue of offspring, but the association of this programming with DNA methylation is not yet clear. Therefore, this experiment aimed to investigate the effects of maternal feed intake restriction on offsprings’ blood indexes, lipid metabolism of perirenal adipose tissue (PAT) and DNA methylation of lipogenic genes. Blood and PAT were collected from the fetuses (at d 100 of gestation) and kids (at d 90 after birth). Maternal undernutrition (at d 45 to d 100) decreases heart and kidney weight of kid goats after birth. Maternal undernutrition decreased the content of leptin in the umbilical cord blood of fetuses. The expression of uncoupling protein 1 (UCP1) was decreased in fetal PAT of the restriction group, but fatty acid synthetase (FASN) was increased. Protein abundance of UCP1 and FASN was down-regulated in the fetal PAT from the restriction group. Furthermore, only the methylation level of the CpG site (from −756 to −757 bp) in the promoter region of FASN gene was increased for the fetal PAT from the restriction group. These results revealed that maternal feed intake restriction influenced the leptin secretion, changed the expression of FASN and UCP1 gene of fetal PAT. This alteration was able to recover in the kids after removing intake restriction, but the growth performance and visceral organ mass were still impaired, suggesting abnormal fat metabolism may happen in the future.

The palliative effects of folic acid on retinal microvessels in diabetic retinopathy via regulating the metabolism of DNA methylation and hydroxymethylation

ABSTRACT Diabetic retinopathy (DR) is one of the severe microvascular complications of diabetes. The protective effects of FA on retinal vascular endothelial cells against high glucose levels involve in multiple aspects in DR; however, the underlying mechanism is not fully elucidated. In present study, we investigated the transcriptome as well as genome-wide DNA methylation and hydroxymethylation signature in human retinal microvascular endothelial ACBRI 181 cells cultured within high glucose (HG) medium supplemented with or without FA by RNA-seq, MeDIP-seq, and hMeDIP-seq. Total 3308 differential expressed genes (DEGs) were involved in multiple biological processes and molecular functions containing angiogenesis, inflammation, S-adenosyl methionine metabolism, and hypoxia response. Moreover, the global DNA methylation and hydroxymethylation in ACBRI 181 cells with FA treatment were both compromised compared to HG. Combined with transcriptome data, four subclusters of DEGs with hyper- or hypomethylated promoters were further verified. Unexpectedly, promoters of these 487 genes all displayed a pattern of increased DNA hydroxymethylation. Furthermore, hyperglycemia rat model was established and administered with FA. The DNA methylation and hydroxymethylation changes of selected target genes COL1A1, ITGA7, MMP-14, and VEGFB confirmed by MeDIP-qPCR were consistent with the results in human ACBRI 181 cells. Finally, the presence of activated DNMT1 and TET2 induced by FA was determined in ACBRI 181 cells and hyperglycemia rat. Taken together, this research provided a resource of expression and epigenetic profiles in retinal microvascular endothelial cell, emphasizing a pharmacological mechanism of FA on DNA methylation and hydroxymethylation regulation in retinal microvessel cells of DR.

Plasma choline and betaine and risks of cardiovascular events and recurrent stroke after ischemic stroke

Choline and betaine have been suggested to play a pivotal role in neurotransmitter synthesis, cell membrane integrity, and methyl-group metabolism, exerting neuroprotective effects in patients with various neurological disorders. However, population-based evidence on choline and betaine with subsequent cardiovascular events after stroke is rare. We aimed to prospectively investigate the relationships of circulating choline and betaine with cardiovascular events and recurrent stroke in patients with ischemic stroke. We performed a nested case-control study within the China Antihypertensive Trial in Acute Ischemic Stroke. A total of 323 cardiovascular events (including 264 recurrent strokes) and 323 controls (free of recurrent cardiovascular events) matched for age (±1 y), sex, and treatment group were included. The primary endpoint was a composite of cardiovascular events after ischemic stroke. Plasma choline and betaine were measured at baseline by ultra-high-performance LC-MS/MS. Conditional logistic regression models were applied, and discrimination, reclassification, and calibration of models with choline pathway metabolites were evaluated. Plasma choline and betaine were inversely associated with cardiovascular events and recurrent stroke after ischemic stroke. Specifically, in fully adjusted models, each additional SD of choline and betaine was associated with 35% (95% CI: 20%-48%) and 30% (95% CI: 14%-43%) decreased risks of subsequent cardiovascular events, respectively, and 34% (95% CI: 16%-48%) and 29% (95% CI: 12%-43%) decreased risks of recurrent stroke, respectively. In addition, both choline and betaine offered substantial risk discrimination and reclassification improvement for cardiovascular events and recurrent stroke beyond traditional risk factors, as evidenced by an increase in C statistics, the net reclassification index, and integrated discrimination improvement. Plasma choline pathway metabolites, including choline and betaine, were associated with decreased risks of cardiovascular events and recurrent stroke and provided incremental value in risk discrimination and stratification in patients with ischemic stroke. This nested case-control study was based on the China Antihypertensive Trial in Acute Ischemic Stroke, which is registered at clinicaltrials.gov as NCT01840072.

S-adenosyl-L-homocysteine hydrolase FgSah1 is required for fungal development and virulence in Fusarium graminearum

ABSTRACT The S-adenosyl-L-homocysteine hydrolase (Sah1) plays a crucial role in methylation and lipid metabolism in yeast and mammals, yet its function remains elusive in filamentous fungi. In this study, we characterized Sah1 in the phytopathogenic fungus F. graminearum by generating knockout and knockout-complemented strains of FgSAH1. We found that the FgSah1-GFP fusion protein was localized to the cytoplasm, and that deletion of FgSAH1 resulted in defects in vegetative growth, asexual and sexual reproduction, stress responses, virulence, lipid metabolism, and tolerance against fungicides. Moreover, the accumulations of S-adenosyl-L-homocysteine (AdoHcy) and S-adenosyl-L-methionine (AdoMet) (the methyl group donor in most methyl transfer reactions) in ΔFgSah1 were seven- and ninefold higher than those in the wild-type strain, respectively. All of these defective phenotypes in ΔFgSah1 mutants were rescued by target gene complementation. Taken together, these results demonstrate that FgSah1 plays essential roles in methylation metabolism, fungal development, full virulence, multiple stress responses, lipid metabolism, and fungicide sensitivity in F. graminearum. To our knowledge, this is the first report on the systematic functional characterization of Sah1 in F. graminearum.

Understanding Choline Bioavailability and Utilization: First Step Toward Personalizing Choline Nutrition.

Choline is an essential macronutrient involved in neurotransmitter synthesis, cell-membrane signaling, lipid transport, and methyl-group metabolism. Nevertheless, the vast majority are not meeting the recommended intake requirement. Choline deficiency is linked to nonalcoholic fatty liver disease, skeletal muscle atrophy, and neurodegenerative diseases. The conversion of dietary choline to trimethylamine by gut microbiota is known for its association with atherosclerosis and may contribute to choline deficiency. Choline-utilizing bacteria constitutes less than 1% of the gut community and is modulated by lifestyle interventions such as dietary patterns, antibiotics, and probiotics. In addition, choline utilization is also affected by genetic factors, further complicating the impact of choline on health. This review overviews the complex interplay between dietary intakes of choline, gut microbiota and genetic factors, and the subsequent impact on health. Understanding of gut microbiota metabolism of choline substrates and interindividual variability is warranted in the development of personalized choline nutrition.

Folic acid supplementation during pregnancy modulates hepatic methyl metabolism and genes expression profile of neonatal lambs of different litter sizes.

Maternal folic acid (FA) plays an important role in the fetus development, but it is unknown the response of hepatic metabolism in the offspring from different litter sizes to maternal FA supplementation. In the present study, this was done by feeding the ewes with 0, 16 and 32 mg/(kg·DM) FA supplemented diet during pregnancy and analysing the hepatic one-carbon metabolism-related indices and gene expression in the neonatal lambs of different litter sizes (twins, TW; triplets, TR). Regardless of litter sizes, the concentrations of folate, methionine, S-adenosylmethionine and DNA methyltransferase increased significantly, but homocysteine and S-adenosylhomocysteine decreased in the liver of newborn lambs from ewes whose diet was supplemented with FA. In TW, maternal FA status has little effect on hepatic genes expression profile of newborn lambs, and no significant enriched pathway was found. However, DEG involved in cell proliferation such as CCNA2, CCNB2, CCNE2, CDK1 and BUB1 were significantly enriched when the ewes were supplemented with FA in TR groups. In addition, nucleotide synthesis-related genes such as POLD1, POLD2, MCM4 and MCM5 were enriched markedly in DNA replication and pyrimidine metabolism pathways in triplets when a higher FA ingestion [32 mg/(kg·DM)] was implemented in ewes. This finding demonstrated that the hepatic methyl metabolism in TW and TR newborn lambs was regulated by maternal FA status. The hepatic cell proliferation and nucleotide metabolism related genes in TR were more susceptible to maternal dietary FA supplementation during pregnancy.

Moderate Folic Acid Supplementation in Pregnant Mice Results in Altered Methyl Metabolism and in Sex-Specific Placental Transcription Changes.

Many pregnant women have higher folic acid (FA) intake due to food fortification and increased vitamin use. It is reported that diets containing five-fold higher FA than recommended for mice (5xFASD) during pregnancy resulted in methylenetetrahydrofolate reductase (MTHFR) deficiency and altered choline/methyl metabolism, with neurobehavioral abnormalities in newborns. The goal is to determine whether these changes have their origins in the placenta during embryonic development. Female mice are fed control diet or 5xFASD for a month before mating and maintained on these diets until embryonic day 17.5. 5xFASD led to pseudo-MTHFR deficiency in maternal liver and altered choline/methyl metabolites in maternal plasma (increased methyltetrahydrofolate and decreased betaine). Methylation potential (S-adenosylmethionine:S-adenosylhomocysteine ratio) and glycerophosphocholine are decreased in placenta and embryonic liver. Folic acid supplemented diet results in sex-specific transcriptome profiles in placenta, with validation of dietary expression changes of 29 genes involved in angiogenesis, receptor biology or neurodevelopment, and altered methylation of the serotonin receptor 2A gene. Moderate increases in folate intake during pregnancy result in placental metabolic and gene expression changes, particularly in angiogenesis, which may contribute to abnormal behavior in pups. These results are relevant for determining a safe upper limit for folate intake during pregnancy.

Folic Acid Supplementation in a Mouse Model of Gestational Diabetes Alters Insulin Sensitivity in Mothers and Liver Methyl Metabolism and Beta Cell Mass in Fetal Offspring

ObjectivesGestational diabetes (GDM) is associated with adverse outcomes in both mother and child. Women with diabetes and obesity (BMI ≥ 30kg/m2) are recommended to take more folic acid than recommended for healthy women, to prevent birth defects. However, little is known about the effects of high folic acid supplementation in GDM. The objective of this study was to determine if folic acid supplementation in a mouse model of GDM affects maternal and fetal metabolic health. MethodsFemale (C57BL/6J) mice were fed from weaning a control diet (10% kcal fat; control dams) or western diet (45% kcal fat; western dams; model of GDM) with (10mg/kg diet) or without (2mg/kg diet) supplemental folic acid (SFA). Dams were fed for 13 weeks prior to breeding. Physiological assessments of glucose homeostasis were conducted before breeding. Tissues were collected at gestational day 18.5 ResultsWestern dams had greater adiposity (p < 0.05), glucose intolerance (p < 0.01), and impaired β cell function (p < 0.05), confirming the model of GDM. Insulin sensitivity was reduced in SFA control dams (p < 0.05) but improved in SFA western dams (p < 0.05) compared to unsupplemented dams. Western dams had lower liver S-adenosylmethionine (SAM) and S-adenosylhomocysteine (SAH) (p < 0.05) compared to control dams; no effect of SFA was observed. SFA control and SFA western male offspring, but not female, had greater β cell mass (p = 0.058) compared to those from unsupplemented dams. Higher liver SAM was observed in SFA control and SFA western female offspring (p = 0.04), but not in male offspring. RNA sequencing identified 203 and 243 differentially expressed genes in liver from male SFA western offspring compared to western male offspring and SFA control male offspring, respectively (p < 0.05, FDR = 0.1). The differentially expressed genes were enriched in lipid metabolism pathways. In contrast, female SFA western offspring had only 12 differentially expressed genes in liver compared to SFA control offspring (p < 0.05, FDR = 0.1). ConclusionsFolic acid supplementation improved insulin sensitivity in dams fed a western diet and has sex-specific effects on fetal offspring β cell mass and liver methyl metabolism and gene expression. Funding SourcesCIHR.

Homocysteine-induced neural tube defects in chick embryos via oxidative stress and DNA methylation associated transcriptional down-regulation of miR-124.

Although moderate homocysteine (HCY) elevation is associated with neural tube defects (NTDs), the underlying mechanisms have not been elucidated. In this study, we aimed to investigate that whether HCY-induced NTDs were associated with oxidative stress and methyl metabolism in chick embryos. The potential role of miR-124 in neurogenesis was also investigated. In this study, increased intracellular oxidative species and alterations in DNA methylation were observed following HCY treatment. This alteration coincided with decreases of Mn superoxide dismutase and glutathione peroxidase activities, as well as the expression of anti-rabbit DNA methyltransferase (DNMT) 1 and 3a. In addition, HCY induced significant decreases of S-adenosylmethionine (SAM)/S-adenosylhomocysteine (SAH) (P<0.05). N-acetyl-L-cysteine and choline ameliorated global DNA hypomethylation induced by HCY. MiR-124 levels were significantly suppressed by HCY (P<0.05), while elevated by 5-aza-2'-deoxycytidine (5-aza-dC). MiR-124 knockdown resulted in spina bifida occulta. Our research suggests that HCY-induced NTDs were associated with oxidative stress and methyl metabolism in chick embryos. MiR-124 down-regulation may occur via epigenetic mechanisms and contribute to HCY-induced NTDs in chick embryo models.

Unique adaptations in neonatal hepatic transcriptome, nutrient signaling, and one-carbon metabolism in response to feeding ethyl cellulose rumen-protected methionine during late-gestation in Holstein cows

BackgroundMethionine (Met) supply during late-pregnancy enhances fetal development in utero and leads to greater rates of growth during the neonatal period. Due to its central role in coordinating nutrient and one-carbon metabolism along with immune responses of the newborn, the liver could be a key target of the programming effects induced by dietary methyl donors such as Met. To address this hypothesis, liver biopsies from 4-day old calves (n = 6/group) born to Holstein cows fed a control or the control plus ethyl-cellulose rumen-protected Met for the last 28 days prepartum were used for DNA methylation, transcriptome, metabolome, proteome, and one-carbon metabolism enzyme activities.ResultsAlthough greater withers and hip height at birth in Met calves indicated better development in utero, there were no differences in plasma systemic physiological indicators. RNA-seq along with bioinformatics and transcription factor regulator analyses revealed broad alterations in ‘Glucose metabolism’, ‘Lipid metabolism, ‘Glutathione’, and ‘Immune System’ metabolism due to enhanced maternal Met supply. Greater insulin sensitivity assessed via proteomics, and efficiency of transsulfuration pathway activity suggested beneficial effects on nutrient metabolism and metabolic-related stress. Maternal Met supply contributed to greater phosphatidylcholine synthesis in calf liver, with a role in very low density lipoprotein secretion as a mechanism to balance metabolic fates of fatty acids arising from the diet or adipose-depot lipolysis. Despite a lack of effect on hepatic amino acid (AA) transport, a reduction in metabolism of essential AA within the liver indicated an AA ‘sparing effect’ induced by maternal Met.ConclusionsDespite greater global DNA methylation, maternal Met supply resulted in distinct alterations of hepatic transcriptome, proteome, and metabolome profiles after birth. Data underscored an effect on maintenance of calf hepatic Met homeostasis, glutathione, phosphatidylcholine and taurine synthesis along with greater efficiency of nutrient metabolism and immune responses. Transcription regulators such as FOXO1, PPARG, E2F1, and CREB1 appeared central in the coordination of effects induced by maternal Met. Overall, maternal Met supply induced better immunometabolic status of the newborn liver, conferring the calf a physiologic advantage during a period of metabolic stress and suboptimal immunocompetence.