Identification of new genetic polymorphisms that alter the dietary requirement for choline and vary in their distribution across ethnic and racial groups.

Effect alleles (alleles with a polymorphism that is associated with the effect being measured) in a small number of single-nucleotide polymorphisms (SNPs) are known to influence the dietary requirement for choline. In this study, we examined a much larger number of SNPs (n=200) in 10 genes related to choline metabolism for associations with development of organ dysfunction (liver or muscle) when 79 humans were fed a low-choline diet. We confirmed that effect alleles in SNPs such as the C allele of PEMT rs12325817 increase the risk of developing organ dysfunction in women when they consume a diet low in choline, and we identified novel effect alleles, such as the C allele of CHKA SNP rs7928739, that alter dietary choline requirements. When fed a low-choline diet, some people presented with muscle damage rather than liver damage; several effect alleles in SLC44A1 (rs7873937, G allele; rs2771040, G; rs6479313, G; rs16924529, A; and rs3199966, C) and one in CHKB (rs1557502, A) were more common in these individuals. This suggests that pathways related to choline metabolism are more important for normal muscle function than previously thought. In European, Mexican, and Asian Americans, and in individuals of African descent, we examined the prevalence of the effect alleles in SNPs that alter choline requirement and found that they are differentially distributed among people of different ethnic and racial backgrounds. Overall, our study has identified novel genetic variants that modulate choline requirements and suggests that the dietary requirement for choline may be different across racial and ethnic groups.-Da Costa, K.-A., Corbin, K. D., Niculescu, M. D., Galanko, J. A., Zeisel, S. H. Identification of new genetic polymorphisms that alter the dietary requirement for choline and vary in their distribution across ethnic and racial groups.

Perinatal manipulation of α-linolenic acid intake induces epigenetic changes in maternal and offspring livers.

Previous studies indicated that the intake of α-linolenic acid (ALA) can alter the concentration of both ω-6 and ω-3 fatty acids in both mother and offspring, with consequences on postnatal brain development. This study describes the association between maternal ALA availability during gestation and lactation, and alterations in the Fads2 DNA methylation in both maternal and offspring livers, at the end of lactation period. Both Fads2 promoter and intron 1 DNA methylation were increased in the groups receiving postnatal flaxseed oil containing 50% ALA (mothers or pups), while bivariate analysis indicated a significant association of the Fads2 epigenetic status in the liver between each mother and its offspring. In addition, Fads2 expression was negatively correlated with promoter methylation at the individual level in maternal livers (P<0.05). This study also indicated that the interplay between ALA availability during gestation and lactation can differentially alter the expression of desaturases and elongases involved in ω-6 and ω-3 metabolic pathways. In summary, when considering the perinatal dietary ALA requirements in mice, both gestation and lactation periods should be considered as having distinct roles in modulating the metabolism of ω-6 and ω-3 fatty acids in maternal mouse livers.

Conceptual shifts needed to understand the dynamic interactions of genes, environment, epigenetics, social processes, and behavioral choices.

Social and behavioral research in public health is often intimately tied to profound, but frequently neglected, biological influences from underlying genetic, environmental, and epigenetic events. The dynamic interplay between the life, social, and behavioral sciences often remains underappreciated and underutilized in addressing complex diseases and disorders and in developing effective remediation strategies. Using a case-study format, we present examples as to how the inclusion of genetic, environmental, and epigenetic data can augment social and behavioral health research by expanding the parameters of such studies, adding specificity to phenotypic assessments, and providing additional internal control in comparative studies. We highlight the important roles of gene-environment interactions and epigenetics as sources of phenotypic change and as a bridge between the life and social and behavioral sciences in the development of robust interdisciplinary analyses.

The epigenetic effects of a high prenatal folate intake in male mouse fetuses exposed in utero to arsenic.

Inorganic arsenic (iAs) is a complete transplacental carcinogen in mice. Previous studies have demonstrated that in utero exposure to iAs promotes cancer in adult mouse offspring, possibly acting through epigenetic mechanisms. Humans and rodents enzymatically convert iAs to its methylated metabolites. This reaction requires S-adenosylmethionine (SAM) as methyl group donor. SAM is also required for DNA methylation. Supplementation with folate, a major dietary source of methyl groups for SAM synthesis, has been shown to modify iAs metabolism and the adverse effects of iAs exposure. However, effects of gestational folate supplementation on iAs metabolism and fetal DNA methylation have never been thoroughly examined. In the present study, pregnant CD1 mice were fed control (i.e. normal folate, or 2.2 mg/kg) or high folate diet (11 mg/kg) from gestational day (GD) 5 to 18 and drank water with 0 or 85 ppm of As (as arsenite) from GD8 to 18. The exposure to iAs significantly decreased body weight of GD18 fetuses and increased both SAM and S-adenosylhomocysteine (SAH) concentrations in fetal livers. High folate intake lowered the burden of total arsenic in maternal livers but did not prevent the effects of iAs exposure on fetal weight or hepatic SAM and SAH concentrations. In fact, combined folate-iAs exposure caused further significant body weight reduction. Notably, iAs exposure alone had little effect on DNA methylation in fetal livers. In contrast, the combined folate-iAs exposure changed the CpG island methylation in 2,931 genes, including genes known to be imprinted. Most of these genes were associated with neurodevelopment, cancer, cell cycle, and signaling networks. The canonical Wnt-signaling pathway, which regulates fetal development, was among the most affected biological pathways. Taken together, our results suggest that a combined in utero exposure to iAs and a high folate intake may adversely influence DNA methylation profiles and weight of fetuses, compromising fetal development and possibly increasing the risk for early-onset of disease in offspring.

Nutritional influence on epigenetics and effects on longevity.

PURPOSE OF REVIEW: This review synthesizes recently published information regarding nutrition and its impact upon epigenetically mediated mechanisms involved in longevity and aging. RECENT FINDINGS: Recent studies enriched considerably our understanding of the relationship between aging and gene-nutrient interactions that continuously shape our phenotype. Epigenetic mechanisms play an important role in mediating between the nutrient inputs and the ensuing phenotypic changes throughout our entire life and seem to be responsible, in part, for the biological changes that occur during aging. Less is known about the epigenetic role that nutrients have in directly influencing longevity and aging. However, recent studies clearly indicated that because nutrition modulates epigenetic events associated with various diseases (e.g., cancer, obesity, and diabetes), there is at least an indirect epigenetic link between nutrition and longevity and, therefore, biologic plausibility to hypothesize the epigenetic role of nutrition in altering longevity. Apart from limited human studies, promising animal studies brought us much closer to understanding how nutrition could have such an impact upon longevity and aging. SUMMARY: Complex epigenetic mechanisms are involved in aging and longevity, directly or indirectly via disease mechanisms. Nutrition has a strong impact upon epigenetic processes and, therefore, holds promise in having important roles in regulating longevity and aging.

Choline deficiency alters global histone methylation and epigenetic marking at the Re1 site of the calbindin 1 gene.

Maternal choline availability is essential for fetal neurogenesis. Choline deprivation (CD) causes hypomethylation of specific CpG islands in genes controlling cell cycling in fetal hippocampus. We now report that, in C57BL/6 mice, CD during gestational days 12-17 also altered methylation of the histone H3 in E17 fetal hippocampi. In the ventricular and subventricular zones, monomethyl-lysine 9 of H3 (H3K9me1) was decreased by 25% (P<0.01), and in the pyramidal layer, dimethyl-lysine 9 of H3 (H3K9me2) was decreased by 37% (P<0.05). These changes were region specific and were not observed in whole-brain preparations. Also, the same effects of CD on H3 methylation were observed in E14 neural progenitor cells (NPCs) in culture. Changes in G9a histone methyltransferase might mediate altered H3K9me2,1. Gene expression of G9a was decreased by 80% in CD NPCs (P<0.001). In CD, H3 was hypomethylated upstream of the RE1 binding site in the calbindin 1 promoter, and 1 CpG site within the calbindin1 promoter was hypermethylated. REST binding to RE1 (recruits G9a) was decreased by 45% (P<0.01) in CD. These changes resulted in increased expression of calbindin 1 in CD (260%; P<0.05). Thus, CD modulates histone methylation in NPCs, and this could underlie the observed changes in neurogenesis.

Phosphatidylethanolamine N-methyltransferase (PEMT) gene expression is induced by estrogen in human and mouse primary hepatocytes.

Choline is an essential nutrient for humans, though some of the requirement can be met by endogenous synthesis catalyzed by phosphatidylethanolamine N-methyltransferase (PEMT). Premenopausal women are relatively resistant to choline deficiency compared with postmenopausal women and men. Studies in animals suggest that estrogen treatment can increase PEMT activity. In this study we investigated whether the PEMT gene is regulated by estrogen. PEMT transcription was increased in a dose-dependent manner when primary mouse and human hepatocytes were treated with 17-beta-estradiol for 24 h. This increased message was associated with an increase in protein expression and enzyme activity. In addition, we report a region that contains a perfect estrogen response element (ERE) approximately 7.5 kb from the transcription start site corresponding to transcript variants NM_007169 and NM-008819 of the human and murine PEMT genes, respectively, three imperfect EREs in evolutionarily conserved regions and multiple imperfect EREs in nonconserved regions in the putative promoter regions. We predict that both the mouse and human PEMT genes have three unique transcription start sites, which are indicative of either multiple promoters and/or alternative splicing. This study is the first to explore the underlying mechanism of why dietary requirements for choline vary with estrogen status in humans.

Lymphocyte gene expression in subjects fed a low-choline diet differs between those who develop organ dysfunction and those who do not.

BACKGROUND: Some humans fed a low-choline diet develop hepatosteatosis, liver and muscle damage, and lymphocyte apoptosis. The risk of developing such organ dysfunction is increased by the presence of single-nucleotide polymorphisms (SNPs) in genes involved in folate and choline metabolism. OBJECTIVE: We investigated whether these changes that occur in the expression of many genes when humans are fed a low-choline diet differ between subjects who develop organ dysfunction and those who do not. We also investigated whether expression changes were dependent on the presence of the SNPs of interest. DESIGN: Thirty-three subjects aged 20-67 y were fed for 10 d a baseline diet containing the recommended adequate intake of choline. They then were fed a low-choline diet for up to 42 d or until they developed organ dysfunction. Blood was collected at the end of each phase, and peripheral lymphocytes were isolated and used for genotyping and for gene expression profiling with the use of microarray hybridization. RESULTS: Feeding a low-choline diet changed the expression of 259 genes, and the profiles of subjects who developed and those who did not develop signs of organ dysfunction differed. Group clustering and gene ontology analyses found that the diet-induced changes in gene expression profiles were significantly influenced by the SNPs of interest and that the gene expression phenotype of the variant gene carriers differed significantly even with the baseline diet. CONCLUSION: These findings support our hypothesis that a person's susceptibility to organ dysfunction when fed a low-choline diet is modulated by specific SNPs in genes involved in folate and choline metabolism.

Dietary choline deficiency alters global and gene-specific DNA methylation in the developing hippocampus of mouse fetal brains.

The availability of choline during critical periods of fetal development alters hippocampal development and affects memory function throughout life. Choline deficiency during fetal development reduces proliferation and migration of neuronal precursor cells in the mouse fetal hippocampus and these changes are associated with modifications in the protein levels of some cell cycle regulators and early differentiation markers. We fed C57 BL/6 mouse dams diets deficient or normal in choline content from days 12 to 17 of pregnancy, and then collected fetal brains on embryonic day 17. Using laser-capture micro-dissection we harvested cells from the ventricular and subventricular zones of Ammon's horn and from the prime germinal zone of the dentate gyrus (hippocampus). In the ventricular and subventricular zones from the choline-deficient group, we observed increased protein levels for kinase-associated phosphatase (Kap) and for p15(INK4b) (two cell cycle inhibitors). In the dentate gyrus, we observed increased levels of calretinin (an early marker of neuronal differentiation). In fetal brain from mothers fed a choline-deficient diet, DNA global methylation was decreased in the ventricular and subventricular zones of Ammon's horn. We also observed decreased gene-specific DNA methylation of the gene (Cdkn3) that encodes for Kap, correlating with increased expression of this protein. This was not the case for p15(INK4b) or calretinin (Cdkn2b and Calb2, respectively). These data suggest that choline deficiency-induced changes in gene methylation could mediate the expression of a cell cycle regulator and thereby alter brain development.

Diethanolamine alters proliferation and choline metabolism in mouse neural precursor cells.

Diethanolamine (DEA) is a widely used ingredient in many consumer products and in a number of industrial applications. It has been previously reported that dermal administration of DEA to mice diminished hepatic stores of choline and altered brain development in the fetus. The aim of this study was to use mouse neural precursor cells in vitro to assess the mechanism underlying the effects of DEA. Cells exposed to DEA treatment (3mM) proliferated less (by 5-bromo-2-deoxyuridine incorporation) at 48 h (24% of control [CT]), and had increased apoptosis at 72 h (308% of CT). Uptake of choline into cells was reduced by DEA treatment (to 52% of CT), resulting in diminished intracellular concentrations of choline and phosphocholine (55 and 12% of CT, respectively). When choline concentration in the growth medium was increased threefold (to 210 microM), the effects of DEA exposure on cell proliferation and apoptosis were prevented, however, intracellular phosphocholine concentrations remained low. In choline kinase assays, we observed that DEA can be phosphorylated to phospho-DEA at the expense of choline. Thus, the effects of DEA are likely mediated by inhibition of choline transport into neural precursor cells and by altered metabolism of choline. Our study suggests that prenatal exposure to DEA may have a detrimental effect on brain development.

Dietary isoflavones differentially induce gene expression changes in lymphocytes from postmenopausal women who form equol as compared with those who do not.

Human and animal studies suggest that dietary soy isoflavones reduce cancer risk, ameliorate postmenopausal syndrome and decrease bone resorption in postmenopausal women. The capacity to form the metabolite equol from daidzein is suggested as an important modulator of response to isoflavones; this capacity depends on gut colonization with appropriate bacteria. We administered a dietary supplement containing high-dose purified soy isoflavones (genistein, 558 mg/day; daidzein, 296 mg/day; and glycitein, 44 mg/day) to 30 postmenopausal women for 84 days and collected peripheral lymphocytes at timed intervals. Using microarray analysis, we determined whether changes in gene expression associated with this treatment support existing hypotheses as to isoflavones' mechanisms of action. Expression of a large number of genes was altered by isoflavone treatment, including induction of genes associated with cyclic adenosine 3',5'-monophosphate (cAMP) signaling and cell differentiation and decreased expression of genes associated with cyclin-dependent kinase activity and cell division. We report that isoflavone treatment in subjects who have the capacity to produce equol differentially affects gene expression as compared with nonproducers, supporting the plausibility of the importance of equol production. In general, isoflavones had a stronger effect on some putative estrogen-responsive genes in equol producers than in nonproducers. Our study suggests that, in humans, isoflavone changes are related to increased cell differentiation, increased cAMP signaling and G-protein-coupled protein metabolism and increased steroid hormone receptor activity and have some estrogen agonist effects; equol-production status is likely to be an important modulator of responses to isoflavones.

Choline deficiency increases lymphocyte apoptosis and DNA damage in humans.

BACKGROUND: Whereas deficiency of the essential nutrient choline is associated with DNA damage and apoptosis in cell and rodent models, it has not been shown in humans. OBJECTIVE: The objective was to ascertain whether lymphocytes from choline-deficient humans had greater DNA damage and apoptosis than did those from choline-sufficient humans. DESIGN: Fifty-one men and women aged 18-70 y were fed a diet containing the recommended adequate intake of choline (control) for 10 d. They then were fed a choline-deficient diet for up to 42 d before repletion with 138-550 mg choline/d. Blood was collected at the end of each phase, and peripheral lymphocytes were isolated. DNA damage and apoptosis were then assessed by activation of caspase-3, terminal deoxynucleotide transferase-mediated dUTP nick end-labeling, and single-cell gel electrophoresis (COMET) assays. RESULTS: All subjects fed the choline-deficient diet had lymphocyte DNA damage, as assessed by COMET assay, twice that found when they were fed the control diet. The subjects who developed organ dysfunction (liver or muscle) when fed the choline-deficient diet had significantly more apoptotic lymphocytes, as assessed by the activated caspase-3 assay, than when fed the control diet. CONCLUSIONS: A choline-deficient diet increased DNA damage in humans. Subjects in whom these diets induced liver or muscle dysfunction also had higher rates of apoptosis in their peripheral lymphocytes than did subjects who did not develop organ dysfunction. Assessment of DNA damage and apoptosis in lymphocytes appears to be a clinically useful measure in humans (such as those receiving parenteral nutrition) in whom choline deficiency is suspected.

Perinatal choline influences brain structure and function.

Choline is derived not only from the diet, but also from de novo synthesis. It is important for methyl-group metabolism, the formation of membranes, kidney function, and neurotransmission. When deprived of dietary choline, most adult men and postmenopausal women develop signs of organ dysfunction (fatty liver or muscle damage) and have a decreased capacity to convert homocysteine to methionine. Choline is critical during fetal development, when it influences stem cell proliferation and apoptosis, thereby altering brain structure and function (memory is permanently enhanced in rodents exposed to choline during the latter part of gestation).

Gene expression profiling of choline-deprived neural precursor cells isolated from mouse brain.

Choline is an essential nutrient and an important methyl donor. Choline deficiency alters fetal development of the hippocampus in rodents and these changes are associated with decreased memory function lasting throughout life. Also, choline deficiency alters global and gene-specific DNA methylation in several models. This gene expression profiling study describes changes in cortical neural precursor cells from embryonic day 14 mice, after 48 h of exposure to a choline-deficient medium. Using Significance Analysis of Microarrays, we found the expression of 1003 genes to be significantly changed (from a total of 16,000 total genes spotted on the array), with a false discovery rate below 5%. A total of 846 genes were overexpressed while 157 were underexpressed. Classification by gene ontology revealed that 331 of these genes modulate cell proliferation, apoptosis, neuronal and glial differentiation, methyl metabolism, and calcium-binding protein classes. Twenty-seven genes that had changed expression have previously been reported to be regulated by promoter or intron methylation. These findings support our previous work suggesting that choline deficiency decreases the proliferation of neural precursors and possibly increases premature neuronal differentiation and apoptosis.

Genetic and epigenetic transgenerational implications related to omega-3 fatty acids. Part II: maternal FADS2 rs174575 genotype and DNA methylation predict toddler cognitive performance.

Maternal transfer of fatty acids is important to fetal brain development. The prenatal environment may differentially affect the substrates supporting declarative memory abilities, as the level of fatty acids transferred across the placenta may be affected by the maternal fatty acid desaturase 2 (FADS2) rs174575 single nucleotide polymorphism. In this study, we hypothesized that toddler and maternal rs174575 genotype and FADS2 promoter methylation would be related to the toddlers' declarative memory performance. Seventy-one 16-month-old toddlers participated in an imitation paradigm designed to test immediate and long-term declarative memory abilities. FADS2 rs174575 genotype was determined and FADS2 promoter methylation was quantified from blood by bisulfite pyrosequencing for the toddlers and their natural mothers. Toddlers of GG mothers at the FADS2 rs174575 single nucleotide polymorphism did not perform as well on memory assessments as toddlers of CC or CG mothers when controlling for plasma α-linolenic acid and child genotype. Toddler methylation status was related to immediate memory performance, whereas maternal methylation status was related to delayed memory performance. Thus, prenatal experience and maternal FADS2 status have a pervasive, long-lasting influence on the brain development of the offspring, but as the postnatal environment becomes more primary, the offsprings' own biology begins to have an effect.

Genetic and epigenetic transgenerational implications related to omega-3 fatty acids. Part I: maternal FADS2 genotype and DNA methylation correlate with polyunsaturated fatty acid status in toddlers: an exploratory analysis.

Polyunsaturated fatty acid metabolism in toddlers is regulated by a complex network of interacting factors. The contribution of maternal genetic and epigenetic makeup to this milieu is not well understood. In a cohort of mothers and toddlers 16 months of age (n = 65 mother-child pairs), we investigated the association between maternal genetic and epigenetic fatty acid desaturase 2 (FADS2) profiles and toddlers' n-6 and n-3 fatty acid metabolism. FADS2 rs174575 variation and DNA methylation status were interrogated in mothers and toddlers, as well as food intake and plasma fatty acid concentrations in toddlers. A multivariate fit model indicated that maternal rs174575 genotype, combined with DNA methylation, can predict α-linolenic acid plasma concentration in all toddlers and arachidonic acid concentrations in boys. Arachidonic acid intake was predictive for its plasma concentration in girls, whereas intake of 3 major n-3 species (eicosapentaenoic, docosapentaenoic, and docosahexaenoic acids) were predictive for their plasma concentrations in boys. FADS2 genotype and DNA methylation in toddlers were not related to plasma concentrations or food intakes, except for CpG8 methylation. Maternal FADS2 methylation was a predictor for the boys' α-linolenic acid intakes. This exploratory study suggests that maternal FADS2 genetic and epigenetic status could be related to toddlers' polyunsaturated fatty acid metabolism.

Perinatal α-linolenic acid availability alters the expression of genes related to memory and to epigenetic machinery, and the Mecp2 DNA methylation in the whole brain of mouse offspring.

Many animal and human studies indicated that dietary ω-3 fatty acids could have beneficial roles on brain development, memory, and learning. However, the exact mechanisms involved are far from being clearly understood, especially for α-linolenic acid (ALA), which is the precursor for the ω-3 elongation and desaturation pathways. This study investigated the alterations induced by different intakes of flaxseed oil (containing 50% ALA), during gestation and lactation, upon the expression of genes involved in neurogenesis, memory-related molecular processes, and DNA methylation, in the brains of mouse offspring at the end of lactation (postnatal day 19, P19). In addition, DNA methylation status for the same genes was investigated. Maternal flaxseed oil supplementation during lactation increased the expression of Mecp2, Ppp1cc, and Reelin, while decreasing the expression of Ppp1cb and Dnmt3a. Dnmt1 expression was decreased by postnatal flaxseed oil supplementation but this effect was offset by ALA deficiency during gestation. Mecp2 DNA methylation was decreased by maternal ALA deficiency during gestation, with a more robust effect in the lactation-deficient group. In addition, linear regression analysis revealed positive correlations between Mecp2, Reelin, and Ppp1cc, between Gadd45b, Bdnf, and Creb1, and between Egr1 and Dnmt1, respectively. However, there were no correlations, in any gene, between DNA methylation and gene expression. In summary, the interplay between ALA availability during gestation and lactation differentially altered the expression of genes involved in neurogenesis and memory, in the whole brain of the offspring at the end of lactation. The Mecp2 epigenetic status was correlated with ALA availability during gestation. However, the epigenetic status of the genes investigated was not associated with transcript levels, suggesting that either the regulation of these genes is not necessarily under epigenetic control, or that the whole brain model is not adequate for the exploration of epigenetic regulation in the context of this study.

Epigenetic regulation of TNFA expression in periodontal disease.

BACKGROUND: Tumor necrosis factor-α (TNF-α) plays a central role in the molecular pathogenesis of periodontal disease. However, the epigenetic regulation attributable to microbial and inflammatory signals at the biofilm-gingival interface are poorly understood. In this study, the DNA methylation alteration within the TNFA promoter in human gingival biopsies from different stages of periodontal disease is investigated and the regulatory mechanism of TNFA transcription by DNA methylation is explored. METHODS: Gingival biopsies were obtained from 17 patients with chronic periodontitis (CP) and 18 periodontally healthy individuals. Another 11 individuals participated in an experimentally induced gingivitis study, and gingival biopsies were collected at the baseline, induction, and resolution phase. To confirm that TNFA promoter methylation modulated TNFA transcription, THP.1 cells were treated with a DNA methyltransferase inhibitor, 5-Aza-2-deoxycytidine (5-Aza-2dC), and an RAW294.7 cell line transfected with a TNFA promoter-specific luciferase reporter system with or without methylation was used. RESULTS: In gingival biopsies from individuals with severe CP, two individual cytosine-guanine dinucleotides (CpG sites) within the TNFA promoter (at -163 and -161 bp) displayed increased methylation in CP samples compared to those with gingival health (16.1% ± 5.1% versus 11.0% ± 4.6%, P = 0.02 and 19.8% ± 4.1% versus 15.4% ± 3.6%, P = 0.04, respectively). The methylation level at -163 bp was inversely associated with the transcription level of TNFA (P = 0.018). However, no significant difference in the TNFA promoter methylation pattern was observed in samples biopsied during the induction or resolution phase of experimentally induced gingivitis, which represented a reversible periodontal lesion. THP.1 cells treated with 5-Aza-2dC demonstrated a time-dependent increase in TNFA messenger level. It was also found that the luciferase activity decreased 2.6-fold in a construct containing an in vitro methylated TNFA promoter when compared to the unmethylated insert (P = 0.03). CONCLUSION: Although the biopsy samples represented a mixed cell population, the change in promoter methylation status in chronic periodontal disease suggested that DNA methylation may be an important regulatory mechanism in controlling TNFA transcriptional expression in periodontal disease.

Challenges in nutrition-related DNA methylation studies.

Abstract The rapid progress in nutritional epigenetics allowed for a much better understanding of the mechanisms involved in gene-nutrient interactions and the roles that nutrition has in transgenerational inheritance of acquired epigenetic traits. Studies indicated that a considerable number of nutrients or diet types are capable of inducing epimutations. In parallel, the rapid development of genome-wide DNA methylation detection methods allowed for a broader image on how nutrition impacts the epigenetic status in human and animal models. But this increased complexity in the epigenetic field and also brought important challenges that need resolution, or it suggests that some of the initial epigenetic paradigms have to be revisited or reconsidered. The aim of this review is to discuss the inherent challenges that need to be resolved, from both practical and theoretical aspects, stemming from the rapid progress in the field of nutritional epigenetics, with a focus on DNA methylation. Because such challenges are present at every stage of study development, the review systematically discusses the most common issues relevant to DNA methylation in a nutritional context. Various types of challenges and potential bias generators are discussed within study design, sample quality, detection methods, data processing, and statistical and bioinformatic analysis. Additional aspects to be considered include epigenetic heterogeneity of treatment groups, the role of genomic variability in introducing measurement bias and errors in interpretation of changes, and issues related to the final interpretation of results and in assigning functional significance. It is also posited that all these issues will be largely resolved within the next decade.

Nutritional epigenetics.

Within the last two decades, significant progress has been made in understanding the importance of epigenetic mechanisms in the regulation of gene expression as a consequence of gene-environment interactions. Nutrition, among many other environmental factors, is a key player that can induce epigenetic changes not only in the directly exposed organisms but also in subsequent generations through the transgenerational inheritance of epigenetic traits. This article aims to provide insights into the usefulness of the mouse model for epigenetic studies involving nutrition as well as the inherent limitations when compared with epigenetic phenomena in humans. Mice are one of the most versatile models for nutrition and epigenetic studies because of several features, such as short life-span, relative low cost for generating samples, the existence of well-characterized genetically engineered lines, the detailed sequencing of genomes, and the relative similarity of their metabolic processes to human metabolism. However, several limitations have to be acknowledged, such as the different location of genes on the chromosomes (and hence possibly different consequences of some epigenetic alterations), differences in the epigenetic patterns established during late embryogenesis, and possible epigenetic differences associated with cellular senescence caused by the different structure of telomeres when compared with humans. All these aspects have to be carefully analyzed when deciding whether a mouse model should be considered for a study in nutrition and epigenetics. Consequently, the results obtained from mouse studies should be carefully interpreted regarding their relevance to humans.