Dietary Modulation of the Epigenome.

Epigenetics is the study of heritable mechanisms that can modify gene activity and phenotype without modifying the genetic code. The basis for the concept of epigenetics originated more than 2,000 yr ago as a theory to explain organismal development. However, the definition of epigenetics continues to evolve as we identify more of the components that make up the epigenome and dissect the complex manner by which they regulate and are regulated by cellular functions. A substantial and growing body of research shows that nutrition plays a significant role in regulating the epigenome. Here, we critically assess this diverse body of evidence elucidating the role of nutrition in modulating the epigenome and summarize the impact such changes have on molecular and physiological outcomes with regards to human health.

The origin of betaine in mouse oocytes and preimplantation embryos†.

Betaine has important roles in preimplantation mouse embryos, including as an organic osmolyte that functions in cell volume regulation in the early preimplantation stages and as a donor to the methyl pool in blastocysts. The origin of betaine in oocytes and embryos was largely unknown. Here, we found that betaine was present from the earliest stage of growing oocytes. Neither growing oocytes nor early preantral follicles could take up betaine, but antral follicles were able to transport betaine and supply the enclosed oocyte. Betaine is synthesized by choline dehydrogenase, and female mice lacking Chdh did not have detectable betaine in their oocytes or early embryos. Supplementing betaine in their drinking water restored betaine in the oocyte only when supplied during the final stages of antral follicle development but not earlier in folliculogenesis. Together with the transport results, this implies that betaine can only be exogenously supplied during the final stages of oocyte growth. Previous work showed that the amount of betaine in the oocyte increases sharply during meiotic maturation due to upregulated activity of choline dehydrogenase within the oocyte. This betaine present in mature eggs was retained after fertilization until the morula stage. There was no apparent role for betaine uptake via the SIT1 (SLC6A20) betaine transporter that is active at the 1- and 2-cell stages. Instead, betaine was apparently retained because its major route of efflux, the volume-sensitive organic osmolyte - anion channel, remained inactive, even though it is expressed and capable of being activated by a cell volume increase.

Prenatal choline, cannabis, and infection, and their association with offspring development of attention and social problems through 4 years of age.

BACKGROUND: Prenatal choline is a key nutrient, like folic acid and vitamin D, for fetal brain development and subsequent mental function. We sought to determine whether effects of higher maternal plasma choline concentrations on childhood attention and social problems, found in an initial clinical trial of choline supplementation, are observed in a second cohort. METHODS: Of 183 mothers enrolled from an urban safety net hospital clinic, 162 complied with gestational assessments and brought their newborns for study at 1 month of age; 83 continued assessments through 4 years of age. Effects of maternal 16 weeks of gestation plasma choline concentrations ⩾7.07 µM, 1 s.d. below the mean level obtained with supplementation in the previous trial, were compared to lower levels. The Attention Problems and Withdrawn Syndrome scales on Child Behavior Checklist 1½-5 were the principal outcomes. RESULTS: Higher maternal plasma choline was associated with lower mean Attention Problems percentiles in children, and for male children, with lower Withdrawn percentiles. Higher plasma choline concentrations also reduced Attention Problems percentiles for children of mothers who used cannabis during gestation as well as children of mothers who had gestational infection. CONCLUSIONS: Prenatal choline's positive associations with early childhood behaviors are found in a second, more diverse cohort. Increases in attention problems and social withdrawal in early childhood are associated with later mental illnesses including attention deficit disorder and schizophrenia. Choline concentrations in the pregnant women in this study replicate other research findings suggesting that most pregnant women do not have adequate choline in their diets.

Effects of a Novel High-Quality Protein Infant Formula on Energetic Efficiency and Tolerance: A Randomized Trial.

OBJECTIVES: Protein overfeeding in infants can have negative effects, such as diabetes and childhood obesity; key to reducing protein intake from formula is improving protein quality. The impact of a new infant formula [study formula (SF)] containing alpha-lactalbumin, lactoferrin, partially hydrolyzed whey, and whole milk on growth and tolerance compared to a commercial formula (CF) and a human milk reference arm was evaluated. METHODS: This randomized, double-blind trial included healthy, singleton, term infants, enrollment age ≤14 days. Primary outcome was mean daily weight gain. Secondary outcomes were anthropometrics, formula intake, serum amino acids, adverse events, gastrointestinal characteristics, and general disposition. RESULTS: Non-inferiority was demonstrated. There were no differences between the formula groups for z scores over time. Formula intake [-0.33 oz/kg/day, 95% confidence interval (CI): -0.66 to -0.01, P = 0.05] and mean protein intake (-0.13 g/kg/day, 95% CI: -0.26 to 0.00, P = 0.05) were lower in the SF infants, with higher serum essential amino acid concentrations (including tryptophan) compared to the CF infants. Energetic efficiency was 14.0% (95% CI: 8.3%, 19.7%), 13.0% (95% CI: 6.0%, 20.0%), and 18.1% (95% CI: 9.4%, 26.8%) higher for weight, length, and head circumference, respectively, in SF infants compared to the CF infants. SF infants had significantly fewer spit-ups and softer stool consistency than CF infants. CONCLUSIONS: The SF resulted in improved parent-reported gastrointestinal tolerance and more efficient growth with less daily formula and protein intake supporting that this novel formula may potentially reduce the metabolic burden of protein overfeeding associated with infant formula.

MicroRNA-129-5p is regulated by choline availability and controls EGF receptor synthesis and neurogenesis in the cerebral cortex.

Choline availability modulates neurogenesis and cerebral cortex development through the regulation of neural progenitor cell (NPC) proliferative and differentiation capacity. In this study, we demonstrated that cortical NPC self-renewal is controlled by choline via the expression of a microRNA (miR-129-5p), whose role in the developing brain has not been examined, and which, in turn, inhibits synthesis of the epidermal growth factor receptor (EGFR) protein. Specifically, we found that low choline (LC) availability led to the upregulation of miR-129-5p expression in cortical NPCs in vitro and in vivo, causing the downregulation of EGFR and thereby disrupting NPC self-renewal and cortical neurogenesis. Furthermore, in response to LC availability, methylation potential (the S-adenosylmethionine: S-adenosylhomocysteine ratio) in the developing brain was reduced. Restoring methylation potential in LC cortical NPCs led to the re-establishment of normal miR-129-5p expression. We concluded that inhibiting miR-129-5p function and restoring EGFR protein levels in vivo is sufficient to reverse LC-induced defects in cortical NPC self-renewal. For the first time, to our knowledge, we have identified the molecular links that explain how a change in the availability of the diet metabolite choline impacts the essential cellular processes underlying brain development.-Trujillo-Gonzalez, I., Wang, Y., Friday, W. B., Vickers, K. C., Toth, C. L., Molina-Torres, L., Surzenko, N., Zeisel, S. H. MicroRNA-129-5p is regulated by choline availability and controls EGF receptor synthesis and neurogenesis in the cerebral cortex.

Low availability of choline in utero disrupts development and function of the retina.

Adequate supply of choline, an essential nutrient, is necessary to support proper brain development. Whether prenatal choline availability plays a role in development of the visual system is currently unknown. In this study, we addressed the role of in utero choline supply for the development and later function of the retina in a mouse model. We lowered choline availability in the maternal diet during pregnancy and assessed proliferative and differentiation properties of retinal progenitor cells (RPCs) in the developing prenatal retina, as well as visual function in adult offspring. We report that low choline availability during retinogenesis leads to persistent retinal cytoarchitectural defects, ranging from focal lesions with displacement of retinal neurons into subretinal space to severe hypocellularity and ultrastructural defects in photoreceptor organization. We further show that low choline availability impairs timely differentiation of retinal neuronal cells, such that the densities of early-born retinal ganglion cells, amacrine and horizontal cells, as well as cone photoreceptor precursors, are reduced in low choline embryonic d 17.5 retinas. Maintenance of higher proportions of RPCs that fail to exit the cell cycle underlies aberrant neuronal differentiation in low choline embryos. Increased RPC cell cycle length, and associated reduction in neurofibromin 2/Merlin protein, an upstream regulator of the Hippo signaling pathway, at least in part, explain aberrant neurogenesis in low choline retinas. Furthermore, we find that animals exposed to low choline diet in utero exhibit a significant degree of intraindividual variation in vision, characterized by marked functional discrepancy between the 2 eyes in individual animals. Together, our findings demonstrate, for the first time, that choline availability plays an essential role in the regulation of temporal progression of retinogenesis and provide evidence for the importance of adequate supply of choline for proper development of the visual system.-Trujillo-Gonzalez, I., Friday, W. B., Munson, C. A., Bachleda, A., Weiss, E. R., Alam, N. M., Sha, W., Zeisel, S. H., Surzenko, N. Low availability of choline in utero disrupts development and function of the retina.

Betaine-homocysteine S-methyltransferase deficiency causes increased susceptibility to noise-induced hearing loss associated with plasma hyperhomocysteinemia.

Betaine-homocysteine S-methyltransferases (BHMTs) are methionine cycle enzymes that remethylate homocysteine; hence, their malfunction leads to hyperhomocysteinemia. Epidemiologic and experimental studies have revealed a correlation between hyperhomocysteinemia and hearing loss. Here, we have studied the expression of methionine cycle genes in the mouse cochlea and the impact of knocking out the Bhmt gene in the auditory receptor. We evaluated age-related changes in mouse hearing by recording auditory brainstem responses before and following exposure to noise. Also, we measured cochlear cytoarchitecture, gene expression by RNA-arrays and quantitative RT-PCR, and metabolite levels in liver and plasma by HPLC. Our results indicate that there is an age-dependent strain-specific expression of methionine cycle genes in the mouse cochlea and a further regulation during the response to noise damage. Loss of Bhmt did not cause an evident impact in the hearing acuity of young mice, but it produced higher threshold shifts and poorer recovery following noise challenge. Hearing loss was associated with increased cochlear injury, outer hair cell loss, altered expression of cochlear methionine cycle genes, and hyperhomocysteinemia. Our results suggest that BHMT plays a central role in the homeostasis of cochlear methionine metabolism and that Bhmt2 up-regulation could carry out a compensatory role in cochlear protection against noise injury in the absence of BHMT.-Partearroyo, T., Murillo-Cuesta, S., Vallecillo, N., Bermúdez-Muñoz, J. M., Rodríguez-de la Rosa, L., Mandruzzato, G., Celaya, A. M., Zeisel, S. H., Pajares, M. A., Varela-Moreiras, G., Varela-Nieto, I. Betaine-homocysteine S-methyltransferase deficiency causes increased susceptibility to noise-induced hearing loss associated with plasma hyperhomocysteinemia.

Dietary choline and betaine intakes and risk of total and lethal prostate cancer in the Atherosclerosis Risk in Communities (ARIC) Study.

PURPOSE: Two prior cohort studies suggested that choline, but not betaine intake, is associated with an increased risk of advanced prostate cancer (PCa). Given that evidence remains limited, we evaluated whether intakes of choline and derivative betaine are associated with total and lethal PCa risk and PCa death in men with PCa. METHODS: We included 6,528 men (24.4% African American) without a cancer diagnosis at baseline (1987-1989) followed through 2012. Dietary intake was assessed using a food frequency questionnaire coupled with a nutrient database. We used Cox proportional hazards regression to estimate hazards ratios (HRs) and 95% confidence intervals (CIs) of total and lethal PCa risk overall and by race. RESULTS: Choline intake was not associated with total (n = 811) or lethal (n = 95) PCa risk overall or by race. Betaine intake was inversely associated with lethal (tertile 3 vs 1, HR 0.59, 95% CI 0.35-1.00, p trend = 0.04), but not total PCa risk; patterns for lethal PCa were similar by race. Neither nutrient was associated with PCa death in men with PCa. CONCLUSIONS: Choline intake was not associated with total or lethal PCa or with PCa death in men with PCa. Betaine intake was inversely associated with lethal, but not total PCa risk or with PCa death in men with PCa. Our results do not support the hypothesis that higher choline intake increases lethal PCa risk, but do suggest that higher betaine intake may be associated with lower lethal PCa risk. Further investigation with a larger number of lethal cases is needed.

Betaine is accumulated via transient choline dehydrogenase activation during mouse oocyte meiotic maturation.

Betaine (N,N,N-trimethylglycine) plays key roles in mouse eggs and preimplantation embryos first in a novel mechanism of cell volume regulation and second as a major methyl donor in blastocysts, but its origin is unknown. Here, we determined that endogenous betaine was present at low levels in germinal vesicle (GV) stage mouse oocytes before ovulation and reached high levels in the mature, ovulated egg. However, no betaine transport into oocytes was detected during meiotic maturation. Because betaine can be synthesized in mammalian cells via choline dehydrogenase (CHDH; EC 1.1.99.1), we assessed whether this enzyme was expressed and active. Chdh transcripts and CHDH protein were expressed in oocytes. No CHDH enzyme activity was detected in GV oocyte lysate, but CHDH became highly active during oocyte meiotic maturation. It was again inactive after fertilization. We then determined whether oocytes synthesized betaine and whether CHDH was required. Isolated maturing oocytes autonomously synthesized betaine in vitro in the presence of choline, whereas this failed to occur in Chdh-/- oocytes, directly demonstrating a requirement for CHDH for betaine accumulation in oocytes. Overall, betaine accumulation is a previously unsuspected physiological process during mouse oocyte meiotic maturation whose underlying mechanism is the transient activation of CHDH.

Trimethylamine N-Oxide, the Microbiome, and Heart and Kidney Disease.

Trimethylamine N-oxide (TMAO) is a biologically active molecule and is a putative promoter of chronic diseases including atherosclerosis in humans. Host intestinal bacteria produce its precursor trimethylamine (TMA) from carnitine, choline, or choline-containing compounds. Most of the TMA produced is passively absorbed into portal circulation, and hepatic flavin-dependent monooxygenases (FMOs) efficiently oxidize TMA to TMAO. Both observational and experimental studies suggest a strong positive correlation between increased plasma TMAO concentrations and adverse cardiovascular events, such as myocardial infarction, stroke, and death. However, a clear mechanistic link between TMAO and such diseases is not yet validated. Therefore, it is debated whether increased TMAO concentrations are the cause or result of these diseases. Here, we have tried to review the current understanding of the properties and physiological functions of TMAO, its dietary sources, and its effects on human metabolism. Studies that describe the potential role of TMAO in the etiology of cardiovascular and other diseases are also discussed.

Altered methylation of specific DNA loci in the liver of Bhmt-null mice results in repression of Iqgap2 and F2rl2 and is associated with development of preneoplastic foci.

Folate B12-dependent remethylation of homocysteine is important, but less is understood about the importance of the alternative betaine-dependent methylation pathway-catalyzed by betaine-homocysteine methyltransferase (BHMT)-for establishing and maintaining adequate DNA methylation across the genome. We studied C57Bl/6J Bhmt (betaine-homocysteine methyltransferase)-null mice at age 4, 12, 24, and 52 wk (N = 8) and observed elevation of S-adenosylhomocysteine concentrations and development of preneoplastic foci in the liver (increased placental glutathione S-transferase and cytokeratin 8-18 activity; starting at 12 wk). At 4 wk, we identified 63 differentially methylated CpGs (DMCs; false discovery rate < 5%) proximal to 81 genes (across 14 chromosomes), of which 18 were differentially expressed. Of these DMCs, 52% were located in one 15.5-Mb locus on chromosome 13, which encompassed the Bhmt gene and defined a potentially sensitive region with mostly decreased methylation. Analyzing Hybrid Mouse Diversity Panel data, which consisted of 100 inbred strains of mice, we identified 97 DMCs that were affected by Bhmt genetic variation in the same region, with 7 overlapping those found in Bhmt-null mice (P < 0.001). At all time points, we found a hypomethylated region mapping to Iqgap2 (IQ motif-containing GTPase activating protein 2) and F2rl2 (proteinase-activated receptor-3), 2 genes that were also silenced and underexpressed, respectively.-Lupu, D. S., Orozco, L. D., Wang, Y., Cullen, J. M., Pellegrini, M., Zeisel, S. H. Altered methylation of specific DNA loci in the liver of Bhmt-null mice results in repression of Iqgap2 and F2rl2 and is associated with development of preneoplastic foci.

Astronaut ophthalmic syndrome.

During and after missions on the International Space Station, some astronauts experience ophthalmic changes, including choroidal folds, optic disc edema, cotton-wool spots, globe flattening, and refraction changes. Astronauts with ophthalmic issues had significantly higher plasma concentrations of metabolites that are associated with the 1-carbon metabolic pathway than those without ophthalmic issues. We hypothesized that genetic differences might explain the metabolite differences. Indeed, genetics and B vitamin status were significant predictors of ophthalmic issues. We now have developed a hypothesis regarding the mechanisms that link 1-carbon pathway genetics and the condition that we suggest calling, "astronaut ophthalmic syndrome." We maintain that this condition is genetically predisposed and is associated with endothelial dysfunction that is induced by oxidative stress. Subsequent edema can hinder cerebrospinal fluid efflux and can lead to locally increased pressures in the subarachnoid space within the orbit, which impinges on the optic nerve and/or eye in affected individuals. Confirming this hypothesis will help characterize the genetics of 1-carbon pathway metabolism, homocysteine, oxidative stress, endothelial dysfunction, and cardiovascular and potentially other diseases.-Zwart, S. R., Gibson, C. R., Gregory, J. F., Mader, T. H., Stover, P. J., Zeisel, S. H., Smith, S. M. Astronaut ophthalmic syndrome.

Feasibility and Acceptability of Maternal Choline Supplementation in Heavy Drinking Pregnant Women: A Randomized, Double-Blind, Placebo-Controlled Clinical Trial.

BACKGROUND: Choline, an essential nutrient, serves as a methyl-group donor for DNA methylation and is a constituent of the neurotransmitter acetylcholine and a precursor to major components of cell membranes. Findings from animal studies suggest that choline supplementation during pregnancy can mitigate adverse effects of prenatal alcohol exposure on growth and neurocognitive function. We conducted a randomized, double-blind exploratory trial to examine feasibility and acceptability of a choline supplementation intervention during pregnancy. METHODS: Seventy heavy drinkers, recruited in mid-pregnancy, were randomly assigned to receive a daily oral dose of 2 g of choline or a placebo from time of enrollment until delivery. Each dose consisted of an individually wrapped packet of powder that, when mixed with water, produced a sweet tasting grape-flavored drink. Adherence was assessed by collecting used and unused drink packets on a monthly basis and tabulating the number used. Side effects were assessed in monthly interviews. Blood samples obtained at enrollment and at 4 and 12 weeks after randomization were assayed for plasma choline concentration. RESULTS: Adherence was good-to-excellent (median doses taken = 74.0%; interquartile range = 53.9 to 88.7%) and was not related to a range of sociodemographic characteristics or to alcohol consumption ascertained using a timeline follow-back interview. By 4 weeks, plasma choline concentrations were significantly higher in the choline supplementation than the placebo arm, and this group difference continued to be evident at 12 weeks. The only side effect was a small increase in nausea/dyspepsia. No effects were seen for diarrhea, vomiting, muscle stiffness, blood pressure, or body odor changes. CONCLUSIONS: This study demonstrated that a choline supplementation program with very heavy drinkers during pregnancy is feasible even among highly disadvantaged, poorly educated women. The broad acceptability of this intervention is indicated by our finding that adherence was not related to maternal education, intellectual function, depression, nutritional status, or alcohol use.

Efficacy of Maternal Choline Supplementation During Pregnancy in Mitigating Adverse Effects of Prenatal Alcohol Exposure on Growth and Cognitive Function: A Randomized, Double-Blind, Placebo-Controlled Clinical Trial.

BACKGROUND: We recently demonstrated the acceptability and feasibility of a randomized, double-blind choline supplementation intervention for heavy drinking women during pregnancy. In this study, we report our results relating to the efficacy of this intervention in mitigating adverse effects of prenatal alcohol exposure (PAE) on infant growth and cognitive function. METHODS: Sixty-nine Cape Coloured (mixed ancestry) heavy drinkers in Cape Town, South Africa, recruited in mid-pregnancy, were randomly assigned to receive a daily oral dose of either 2 g of choline or placebo from time of enrollment until delivery. Each dose consisted of an individually wrapped packet of powder that, when mixed with water, produced a sweet tasting grape-flavored drink. The primary outcome, eyeblink conditioning (EBC), was assessed at 6.5 months. Somatic growth was measured at birth, 6.5, and 12 months, recognition memory and processing speed on the Fagan Test of Infant Intelligence, at 6.5 and 12 months. RESULTS: Infants born to choline-treated mothers were more likely to meet criterion for conditioning on EBC than the placebo group. Moreover, within the choline arm, degree of maternal adherence to the supplementation protocol strongly predicted EBC performance. Both groups were small at birth, but choline-treated infants showed considerable catch-up growth in weight and head circumference at 6.5 and 12 months. At 12 months, the infants in the choline treatment arm had higher novelty preference scores, indicating better visual recognition memory. CONCLUSIONS: This exploratory study is the first to provide evidence that a high dose of choline administered early in pregnancy can mitigate adverse effects of heavy PAE on EBC, postnatal growth, and cognition in human infants. These findings are consistent with studies of alcohol-exposed animals that have demonstrated beneficial effects of choline supplementation on classical conditioning, learning, and memory.

CASAnova: a multiclass support vector machine model for the classification of human sperm motility patterns.

The ability to accurately monitor alterations in sperm motility is paramount to understanding multiple genetic and biochemical perturbations impacting normal fertilization. Computer-aided sperm analysis (CASA) of human sperm typically reports motile percentage and kinematic parameters at the population level, and uses kinematic gating methods to identify subpopulations such as progressive or hyperactivated sperm. The goal of this study was to develop an automated method that classifies all patterns of human sperm motility during in vitro capacitation following the removal of seminal plasma. We visually classified CASA tracks of 2817 sperm from 18 individuals and used a support vector machine-based decision tree to compute four hyperplanes that separate five classes based on their kinematic parameters. We then developed a web-based program, CASAnova, which applies these equations sequentially to assign a single classification to each motile sperm. Vigorous sperm are classified as progressive, intermediate, or hyperactivated, and nonvigorous sperm as slow or weakly motile. This program correctly classifies sperm motility into one of five classes with an overall accuracy of 89.9%. Application of CASAnova to capacitating sperm populations showed a shift from predominantly linear patterns of motility at initial time points to more vigorous patterns, including hyperactivated motility, as capacitation proceeds. Both intermediate and hyperactivated motility patterns were largely eliminated when sperm were incubated in noncapacitating medium, demonstrating the sensitivity of this method. The five CASAnova classifications are distinctive and reflect kinetic parameters of washed human sperm, providing an accurate, quantitative, and high-throughput method for monitoring alterations in motility.

Maternal dietary intake of choline in mice regulates development of the cerebral cortex in the offspring.

Maternal diets low in choline, an essential nutrient, increase the risk of neural tube defects and lead to low performance on cognitive tests in children. However, the consequences of maternal dietary choline deficiency for the development and structural organization of the cerebral cortex remain unknown. In this study, we fed mouse dams either control (CT) or low-choline (LC) diets and investigated the effects of choline on cortical development in the offspring. As a result of a low choline supply between embryonic day (E)11 and E17 of gestation, the number of 2 types of cortical neural progenitor cells (NPCs)-radial glial cells and intermediate progenitor cells-was reduced in fetal brains (P< 0.01). Furthermore, the number of upper layer cortical neurons was decreased in the offspring of dams fed an LC diet at both E17 (P< 0.001) and 4 mo of age (P< 0.001). These effects of LC maternal diet were mediated by a decrease in epidermal growth factor receptor (EGFR) signaling in NPCs related to the disruption of EGFR posttranscriptional regulation. Our findings describe a novel mechanism whereby low maternal dietary intake of choline alters brain development.-Wang, Y., Surzenko, N., Friday, W. B., Zeisel, S. H. Maternal dietary intake of choline in mice regulates development of the cerebral cortex in the offspring.

Genotype, B-vitamin status, and androgens affect spaceflight-induced ophthalmic changes.

Ophthalmic changes have occurred in a subset of astronauts on International Space Station missions. Visual deterioration is considered the greatest human health risk of spaceflight. Affected astronauts exhibit higher concentrations of 1-carbon metabolites (e.g., homocysteine) before flight. We hypothesized that genetic variations in 1-carbon metabolism genes contribute to susceptibility to ophthalmic changes in astronauts. We investigated 5 polymorphisms in the methionine synthase reductase (MTRR), methylenetetrahydrofolate reductase (MTHFR), serine hydroxymethyltransferase (SHMT), and cystathionine ß-synthase (CBS) genes and their association with ophthalmic changes after flight in 49 astronauts. The number of G alleles of MTRR 66 and C alleles of SHMT1 1420 both contributed to the odds of visual disturbances. Preflight dehydroepiandrosterone was positively associated with cotton wool spots, and serum testosterone response during flight was associated with refractive change. Block regression showed that B-vitamin status and genetics were significant predictors of many of the ophthalmic outcomes that we observed. In one example, genetics trended toward improving (P = 0.10) and B-vitamin status significantly improved (P < 0.001) the predictive model for refractive change after flight. We document an association between MTRR 66 and SHMT1 1420 polymorphisms and spaceflight-induced vision changes. This line of research could lead to therapeutic options for both space travelers and terrestrial patients.

Interactions Between Nuclear Receptor SHP and FOXA1 Maintain Oscillatory Homocysteine Homeostasis in Mice.

BACKGROUND & AIMS: Hyperhomocysteinemia is often associated with liver and metabolic diseases. We studied nuclear receptors that mediate oscillatory control of homocysteine homeostasis in mice. METHODS: We studied mice with disruptions in Nr0b2 (called small heterodimer partner [SHP]-null mice), betaine-homocysteine S-methyltransferase (Bhmt), or both genes (BHMT-null/SHP-null mice), along with mice with wild-type copies of these genes (controls). Hyperhomocysteinemia was induced by feeding mice alcohol (National Institute on Alcohol Abuse and Alcoholism binge model) or chow diets along with water containing 0.18% DL-homocysteine. Some mice were placed on diets containing cholic acid (1%) or cholestyramine (2%) or high-fat diets (60%). Serum and livers were collected during a 24-hour light-dark cycle and analyzed by RNA-seq, metabolomic, and quantitative polymerase chain reaction, immunoblot, and chromatin immunoprecipitation assays. RESULTS: SHP-null mice had altered timing in expression of genes that regulate homocysteine metabolism compared with control mice. Oscillatory production of S-adenosylmethionine, betaine, choline, phosphocholine, glyceophosphocholine, cystathionine, cysteine, hydrogen sulfide, glutathione disulfide, and glutathione, differed between SHP-null mice and control mice. SHP inhibited transcriptional activation of Bhmt and cystathionine γ-lyase by FOXA1. Expression of Bhmt and cystathionine γ-lyase was decreased when mice were fed cholic acid but increased when they were placed on diets containing cholestyramine or high-fat content. Diets containing ethanol or homocysteine induced hyperhomocysteinemia and glucose intolerance in control, but not SHP-null, mice. In BHMT-null and BHMT-null/SHP-null mice fed a control liquid, lipid vacuoles were observed in livers. Ethanol feeding induced accumulation of macrovesicular lipid vacuoles to the greatest extent in BHMT-null and BHMT-null/SHP-null mice. CONCLUSIONS: Disruption of Shp in mice alters timing of expression of genes that regulate homocysteine metabolism and the liver responses to ethanol and homocysteine. SHP inhibits the transcriptional activation of Bhmt and cystathionine γ-lyase by FOXA1.

Folic acid deficiency induces premature hearing loss through mechanisms involving cochlear oxidative stress and impairment of homocysteine metabolism.

Nutritional imbalance is emerging as a causative factor of hearing loss. Epidemiologic studies have linked hearing loss to elevated plasma total homocysteine (tHcy) and folate deficiency, and have shown that folate supplementation lowers tHcy levels potentially ameliorating age-related hearing loss. The purpose of this study was to address the impact of folate deficiency on hearing loss and to examine the underlying mechanisms. For this purpose, 2-mo-old C57BL/6J mice (Animalia Chordata Mus musculus) were randomly divided into 2 groups (n = 65 each) that were fed folate-deficient (FD) or standard diets for 8 wk. HPLC analysis demonstrated a 7-fold decline in serum folate and a 3-fold increase in tHcy levels. FD mice exhibited severe hearing loss measured by auditory brainstem recordings and TUNEL-positive-apoptotic cochlear cells. RT-quantitative PCR and Western blotting showed reduced levels of enzymes catalyzing homocysteine (Hcy) production and recycling, together with a 30% increase in protein homocysteinylation. Redox stress was demonstrated by decreased expression of catalase, glutathione peroxidase 4, and glutathione synthetase genes, increased levels of manganese superoxide dismutase, and NADPH oxidase-complex adaptor cytochrome b-245, α-polypeptide (p22phox) proteins, and elevated concentrations of glutathione species. Altogether, our findings demonstrate, for the first time, that the relationship between hyperhomocysteinemia induced by folate deficiency and premature hearing loss involves impairment of cochlear Hcy metabolism and associated oxidative stress.