Zoe Yates

Folic acid fortification: a double-edged sword.

Purpose of reviewTo examine the impact of folic acid fortification, including its use as a functional food component, on human health. Recent findingsThere is a consensus view that folic acid supplementation has numerous health benefits, many of which are significant in their impact. However, emerging evidence suggests that increased population exposure to folic acid may also have a negative impact with respect to certain developmental and degenerative disorders. As examples, presently much attention is focused on the role of folic acid fortification augmenting colon cancer risk, whereas earlier in the life cycle, the vitamin may additionally influence insulin resistance. Without question, conditions that are influenced by folic acid are both diverse and many – from concerns relating to cognitive decline, breast cancer and vascular disease through to preconceptional issues where maternal folate levels might conceivably alter the phenotype of offspring via epimutations. SummaryThe highly complex and critical biological importance of folic acid-related molecular nutrition makes it a difficult micronutrient to deploy as a simple intervention at a population level – it has far too many biochemical spheres of influence to predict effects in a generalized way. Additionally, several gene variants and other nutrients are interactive factors. It is, therefore, hardly surprising that the scientific community does not have a true consensus view on whether mandatory fortification is appropriate as a population measure. This latter point not withstanding, any ultimate decisions on fortification should be well rooted in scientific fact rather than political expediency.

The methylenetetrahydrofolate reductase C677T mutation induces cell-specific changes in genomic DNA methylation and uracil misincorporation: A possible molecular basis for the site-specific cancer risk modification

The C677T polymorphism in the methylenetetrahydrofolate reductase (MTHFR) gene is associated with a decreased risk of colon cancer although it may increase the risk of breast cancer. This polymorphism is associated with changes in intracellular folate cofactors, which may affect DNA methylation and synthesis via altered one‐carbon transfer reactions. We investigated the effect of this mutation on DNA methylation and uracil misincorporation and its interaction with exogenous folate in further modulating these biomarkers of one‐carbon transfer reactions in an in vitro model of the MTHFR 677T mutation in HCT116 colon and MDA‐MB‐435 breast adenocarcinoma cells. In HCT116 cells, the MTHFR 677T mutation was associated with significantly increased genomic DNA methylation when folate supply was adequate or high; however, in the setting of folate insufficiency, this mutation was associated with significantly decreased genomic DNA methylation. In contrast, in MDA‐MB‐435 cells, the MTHFR 677T mutation was associated with significantly decreased genomic DNA methylation when folate supply was adequate or high and with no effect when folate supply was low. The MTHFR 677T mutation was associated with a nonsignificant trend toward decreased and increased uracil misincorporation in HCT116 and MDA‐MB‐435 cells, respectively. Our data demonstrate for the first time a functional consequence of changes in intracellular folate cofactors resulting from the MTHFR 677T mutation in cells derived from the target organs of interest, thus providing a plausible cellular mechanism that may partly explain the site‐specific modification of colon and breast cancer risks associated with the MTHFR C677T mutation.

A critical role for B-vitamin nutrition in human developmental and evolutionary biology

Several molecular mechanisms that underpin genomic integrity and function are sensitive to B-vitamin status, and in particular, are responsive to the interaction between folate nutrition and folate dependent enzyme polymorphisms. Mechanisms that may be affected include maintenance of genomic CpG methylation patterns for regulated gene expression, and proficient synthesis of nucleotides to prevent DNA strand breakage. We review some of these important molecular mechanisms that underpin the role of folate and other B-vitamins in disease processes, and discuss how they explain why this vitamin may be an important factor in human evolution. We look at three particular phenomena that link folate status/genes to evolutionary pressures: 1) The evolution of pigmentation to avert the impact of UV light on labile folates needed for developmental processes. 2) The effect of C677T-MTHFR on foetal survival, and 3) the ability of folate to mask harmful developmental mutations.

Contemporary Issues Surrounding Folic Acid Fortification Initiatives

The impact of folate on health and disease, particularly pregnancy complications and congenital malformations, has been extensively studied. Mandatory folic acid fortification therefore has been implemented in multiple countries, resulting in a reduction in the occurrence of neural tube defects. However, emerging evidence suggests increased folate intake may also be associated with unexpected adverse effects. This literature review focuses on contemporary issues of concern, and possible underlying mechanisms as well as giving consideration the future direction of mandatory folic acid fortification. Folate fortification has been associated with the presence of unmetabolized folic acid (PteGlu) in blood, masking of vitamin B12 deficiency, increased dosage for anti-cancer medication, photo-catalysis of PteGlu leading to potential genotoxicity, and a role in the pathoaetiology of colorectal cancer. Increased folate intake has also been associated with twin birth and insulin resistance in offspring, and altered epigenetic mechanisms of inheritance. Although limited data exists to elucidate potential mechanisms underlying these issues, elevated blood folate level due to the excess use of PteGlu without consideration of an individual’s specific phenotypic traits (e.g. genetic background and undiagnosed disease) may be relevant. Additionally, the accumulation of unmetabolized PteGlu may lead to inhibition of dihydrofolate reductase and other enzymes. Concerns notwithstanding, folic acid fortification has achieved enormous advances in public health. It therefore seems prudent to target and carefully monitor high risk groups, and to conduct well focused further research to better understand and to minimize any risk of mandatory folic acid fortification.

Effects of folylpolyglutamate synthetase modulation on chemosensitivity of colon cancer cells to 5-fluorouracil and methotrexate

Background: Folylpoly-γ-glutamate synthetase (FPGS) converts intracellular folates and antifolates (for example, methotrexate (MTX)) to polyglutamates. Polyglutamylated folates and antifolates are retained in cells longer and are better substrates than their monoglutamate counterparts for enzymes involved in one carbon transfer. Polyglutamylation of intracellular 5,10-methylenetetrahydrofolate may also enhance the cytotoxicity of 5-fluorouracil (5-FU) by allowing more efficient formation and stabilisation of the inhibitory ternary complex involving thymidylate synthase and a 5-FU metabolite. Aim: We investigated the effects of FPGS modulation on the chemosensitivity of colon cancer cells to 5-FU and MTX. Methods: Human HCT116 colon cancer cells were stably transfected with the sense or antisense FPGS cDNA or blank (control). FPGS protein expression and enzyme activity, growth rate, intracellular folate content and composition, and in vitro chemosensitivity to 5-FU and MTX were determined. Results: Compared with cells expressing endogenous FPGS, those overexpressing FPGS had significantly faster growth rates and higher concentrations of total folate and long chain folate polyglutamates while antisense FPGS inhibition produced opposite results. FPGS overexpression significantly enhanced, whereas FPGS inhibition decreased, chemosensitivity to 5-FU. No significant difference in chemosensitivity to MTX was observed. Conclusions: These data provide functional evidence that FPGS overexpression and inhibition modulate chemosensitivity of colon cancer cells to 5-FU by altering intracellular folate polyglutamylation, providing proof of principle. Thus FPGS status may be an important predictor of chemosensitivity of colon cancer cells to 5-FU based chemotherapy, and FPGS gene transfer may increase the sensitivity of colon cancer cells to 5-FU-based chemotherapy.

Vitamin D, folate, and potential early lifecycle environmental origin of significant adult phenotypes

Solar radiation early in pregnancy interacts with light sensitive vitamins to influence an embryos genetic profile. This influences both adult disease risk and may play a role in the evolution of skin colour.

Polycystic ovary syndrome and the single nucleotide polymorphisms of methylenetetrahydrofolate reductase: a pilot observational study

Polycystic ovary syndrome (PCOS), insulin resistance and overall mortality due to diabetes and coronary artery disease are higher in South Asians than in Caucasians. Aims: We compared the prevalence of the C677T and A1298C single nucleotide polymorphisms in the methylenetetrahydrofolate reductase gene in South Asian and Caucasian women, its association with folate and homocysteine (Hcy) metabolism, and its relevance to future atherogenic events. Methods and results: 71 women were recruited for the study: South Asian PCOS (21) plus controls (9) and Caucasian PCOS (25) plus controls (16). Anthropometric and laboratory parameters were compared. South Asian PCOS women were significantly hyperandrogenic and exhibited a greater degree of insulin resistance. Caucasian PCOS women had higher plasma Hcy concentrations with a 1.9 times higher frequency of the T allele than the South Asian PCOS group. In the presence of this variant allele, plasma Hcy levels appear to be higher in both PCOS groups. The South Asians had a 1.8 times higher frequency of the C allele than the Caucasians; however, the overall frequency was comparable in the two PCOS groups. The frequency of homozygosity, i.e. TT677 and CC1298, was 7.2% and 4.9% in the Caucasians and 0% and 16.6% in the South Asian recruits, respectively. Dietary inadequacies in the South Asian women can influence their plasma folate and B12 concentrations resulting in hyperhomocysteinemia which, in combination with dyslipidaemia and insulin resistance, can lead to long-term atherogenic consequences. Conclusions: Current data suggests that the mechanisms of atherothrombosis have separate pathways in the two ethnic groups. Larger studies exploring the current theme need to be carried out in the PCOS groups to obtain adequate insight.

The impact of phenylketonuria on folate metabolism

Several reports indicate that biopterin and folate pathways may interact. We examined folate metabolism in PKU patients where hyperphenylalaninaemia leads to a likely excess of THB. We found an increase in total HPLC determined red cell folate in PKU (p=0.0422): specifically, there was an increase in total formyl-H(4)folate (p=0.0002) and H(4)folate (p< or =0.0001), and decrease in total 5-methyl-H(4)folate in PKU patients. At the level of individual oligo-gamma-glutamyl coenzymes, we found that formyl-H(4)folate polyglutamates were virtually all increased in PKU (p=0.0223, 0.0004, 0.0004, 0.0012, and 0.0008 for di-, tri-, tetra-, penta-, and deca-gamma-glutamyl formyl-H(4)folate coenzymes, respectively). Hcy levels did not differ between clinical groups, indicating that folate dependent-Hcy remethylation is not compromised as a consequence of an altered PKU folate disposition. In nature, pentaglutamyl folates are considered the metabolically favoured coenzymes (optimum K(m) for dependent enzymes). The presented data support this-we found that red cell pentaglutamates gave the best measure of metabolism; pentaglutamyl formyl-H(4)folate increased in PKU (p=0.0012) and related methenyls behaved similarly, while, pentaglutamyl 5-methyl-H(4)folate and pentaglutamyl H(4)folate decreased (p< or =0.0001 and 0.0265, respectively). Furthermore, pentaglutamates showed the best correlations between one-carbon oxidation states of folate, as well as with Hcy (p=0.0003 r=-0.54, 95% CI; -0.724 to -0.272). That PKU might influence folate metabolism in some way is unsurprising: patients with DHPR deficiency accumulate DHB and develop secondary folate deficiency-responsive only to reduced folates, while CSF levels of THB are significantly correlated to monoamines and red cell folate in depression. Further studies to confirm the present findings and to ascertain precisely what mechanism operates in PKU that impacts upon folate homeostasis so profoundly are required.

TAS2R38 bitter taste genetics, dietary vitamin C, and both natural and synthetic dietary folic acid predict folate status, a key micronutrient in the pathoaetiology of adenomatous polyps

Taste perception may influence dietary preferences and nutrient intakes contributing to diet-related disease susceptibility. This study examined bitter taste genetics and whether variation in the TAS2R38 gene at three polymorphic loci (A49P, V262A and I296V) could alter dietary and systemic folate levels and dietary vitamin C intake, and whether a nutrigenetic circuit existed that might link bitter taste, folate/antioxidant status and risk for a colonic adenomatous polyp. TAS2R38 diplotype predicted bitter taste (PROP) phenotype (p value <0.00001) and red cell folate status (p=0.0179) consistent with the diplotype that has the broadest range of bitter perception (AVI/PAV) also possessing the highest average red cell folate value. However, TAS2R38 diplotype did not predict dietary intake of methylfolic acid, pteroylmonoglutamic acid or total folic acid. Neither did it predict dietary intake of vitamin C. Despite this, intake of dietary folate predicts red cell folate with analysis pointing to a key nutrient-nutrient interaction between vitamin C intake and systemic folate status. Analysis of 38 patients with an adenomatous polyp and 164 controls showed that individually, dietary nutrient intake, nutrient status and taste diplotype did not influence polyp risk. However, red cell folate status (in individuals below the population median value) did interact with bitter taste diplotype (AVI/PAV) to predict polyp risk (p=0.0145). Furthermore, synthetic folic acid (below median intake) was statistically associated with adenoma occurrence (p=0.0215); individuals with adenomatous polyps had a 1.77× higher intake than controls. Additionally, stepwise regression taking account of all dietary nutrients showed a tight relationship between methylfolic acid (but not pteroylmonoglutamic acid) intake and red cell folate level in those with a low folate status and occurrence of an adenomatous polyp (p=0.0039). These findings point to a role for folate in the pathoaetiology of adenomatous polyps, with the natural and synthetic vitamers not necessarily having the same biological effect.

Photoperiod at conception predicts C677T‐MTHFR genotype: A novel gene‐environment interaction

Data is presented, which suggest that the day length a woman experiences during the periconceptional period predicts the C677T‐MTHFR genotype of her child. Logistic regression analysis involving 375 neonates born in the same geographical location within a three year period demonstrated that photoperiod (minutes) at conception predicts both genotype (P = 0.0139) and mutant allele carriage (P = 0.0161); the trend clearly showing that the 677T‐MTHFR allele frequency increases as photoperiod increases. We propose a number of explanations, including a hypothesis in which a long photoperiod around conception decreases maternal systemic folate because of UVA induced dermal oxidative degradation of 5‐methyl‐H4folate, leading to a lower cellular 5,10‐methylene‐H4folate status. In this scenario, 5,10‐methylene‐H4folate would be more efficiently used for dTMP and DNA synthesis by 677T‐MTHFR embryos than wildtype embryos giving the 677T‐MTHFR embryos increased viability, and hence increasing mutant T‐allele frequency. Alternate hypotheses include: increased seasonal availability of folate rich foods that genetically buffer any negative effect of 677T‐MTHFR in embryos; seasonal oxidative stress lowering embryo‐toxic homocysteine; an undefined hormonal effect of photoperiod on the neuroendocrine axis, which mediates genotype/embryo selection. The effect of photoperiod on genotype seems clear, but the speculative molecular mechanism underpinning the effect needs careful examination. Am. J. Hum. Biol., 2010.