Nucleus accumbens shell moderates preference bias during voluntary choice behavior.

The nucleus accumbens (NAc) shell lies anatomically at a critical intersection within the brain's reward system circuitry, however, its role in voluntary choice behavior remains unclear. Rats with electrolytic lesions in the NAc shell were tested in a novel foraging paradigm. Over a continuous two-week period they freely chose among four nutritionally identical but differently flavored food pellets by pressing corresponding levers. We examined the lesion's effects on three behavioral dynamics components: motivation (when to eat), preference bias (what to choose) and persistence (how long to repeat the same choice). The lesion led to a marked increase in the preference bias: i.e., increased selection of the most-preferred choice option, and decreased selection of the others. We found no effects on any other behavioral measures, suggesting no effect on motivation or choice persistence. The results implicate the NAc shell in moderating the instrumental valuation process by inhibiting excessive bias toward preferred choice options.

Characterization of the newly isolated ω-oxidizing yeast Candida sorbophila DS02 and its potential applications in long-chain dicarboxylic acid production.

α, ω-Dicarboxylic acids (DCAs) are multipurpose chemicals widely used in polymers, perfumes, plasticizers, lubricants, and adhesives. The biotransformation of DCAs from alkanes and fatty acids by microorganisms has attracted recent interest, since synthesis via chemical oxidation causes problems in terms of the environment and safety. We isolated an ω-oxidizing yeast from a wastewater disposal facility of a petrochemical factory by chemostat enrichment culture. The haploid strain identified as Candida sorbophila DS02 grew on glucose and dodecane, exhibiting greater cell shrinkage on the latter. In flask cultures with mixed alkanes (C10-16) and fatty acid methyl esters (C10-16), DS02 used mixed alkanes simultaneously unlike Candida tropicalis and Yarrowia lipolytica and showed high substrate resistance. In flask cultures with acrylic acid-a known inhibitor of ß-oxidation-DS02 produced 0.28 g/l dodecanedioic acid (DDDA) from dodecane, similar to wild-type C. tropicalis ATCC 20336. In fed-batch fermentation, DS02 produced 9.87 g/l DDDA, which was 5.7-fold higher than wild-type C. tropicalis. These results suggest that C. sorbophila strain DS02 has potential applications for the large-scale production of DCA.

The Effects of a Single Developmentally Entrained Pulse of Testosterone in Female Neonatal Mice on Reproductive and Metabolic Functions in Adult Life.

Early postnatal exposures to sex steroids have been well recognized to modulate predisposition to diseases of adulthood. There is a complex interplay between timing, duration and dose of endocrine exposures through environmental or dietary sources that may alter the sensitivity of target tissues to the exogenous stimuli. In this study, we determined the metabolic and reproductive programming effects of a single developmentally entrained pulse of testosterone (T) given to female mice in early postnatal period. CD-1 female mice pups were injected with either 5 µg of T enanthate (TE) or vehicle (control [CON] group) within 24 hours after birth and followed to adult age. A total of 66% of T-treated mice exhibited irregular cycling, anovulatory phenotype, and significantly higher ovarian weights than vehicle-treated mice. Longitudinal nuclear magnetic resonance measurements revealed that TE group had greater body weight, whole-body lean, and fat mass than the CON group. Adipose tissue cellularity analysis in TE group revealed a trend toward higher size and number than their littermate CONs. The brown adipose tissue of TE mice exhibited white fat infiltration with down-regulation of several markers, including uncoupling protein 1 (UCP-1), cell death-inducing DNA fragmentation factor, α-subunit-like effector A, bone morphogenetic protein 7 as well as brown adipose tissue differentiation-related transcription regulators. T-injected mice were also more insulin resistant than CON mice. These reproductive and metabolic reprogramming effects were not observed in animals exposed to TE at 3 and 6 weeks of age. Collectively, these data suggest that sustained reproductive and metabolic alterations may result in female mice from a transient exposure to T during a narrow postnatal developmental window.

Bursts and heavy tails in temporal and sequential dynamics of foraging decisions.

A fundamental understanding of behavior requires predicting when and what an individual will choose. However, the actual temporal and sequential dynamics of successive choices made among multiple alternatives remain unclear. In the current study, we tested the hypothesis that there is a general bursting property in both the timing and sequential patterns of foraging decisions. We conducted a foraging experiment in which rats chose among four different foods over a continuous two-week time period. Regarding when choices were made, we found bursts of rapidly occurring actions, separated by time-varying inactive periods, partially based on a circadian rhythm. Regarding what was chosen, we found sequential dynamics in affective choices characterized by two key features: (a) a highly biased choice distribution; and (b) preferential attachment, in which the animals were more likely to choose what they had previously chosen. To capture the temporal dynamics, we propose a dual-state model consisting of active and inactive states. We also introduce a satiation-attainment process for bursty activity, and a non-homogeneous Poisson process for longer inactivity between bursts. For the sequential dynamics, we propose a dual-control model consisting of goal-directed and habit systems, based on outcome valuation and choice history, respectively. This study provides insights into how the bursty nature of behavior emerges from the interaction of different underlying systems, leading to heavy tails in the distribution of behavior over time and choices.

Rats exhibit reference-dependent choice behavior.

Human preferences depend on whether a chosen outcome appears to be a loss or a gain compared with what had been expected, i.e., in comparison to a reference point. Because reference dependence has such a strong influence on human decision-making, it is important to uncover its origins, which will in turn help delineate the underlying mechanisms. It remains unknown whether rats use reference points in decision-making, and yet, the study of rats could help address the question of whether reference dependence is evolutionarily conserved among mammals and could provide a nonhuman animal model to investigate the neural mechanisms underlying this important cognitive process. The aim of the current study was to determine whether rats show reference-dependent choice behavior. We developed a novel paradigm by modifying the "T" maze by installing "pockets" to the left and right of the "T" stem that held reward pellets so rats would potentially develop reference values for each option prior to choice. We found that the rats were indeed sensitive to the way alternatives were presented. That is, they exhibited reference-dependent choice behavior by avoiding the choice option framed as a loss (e.g., having four reward pellets in the pocket, but receiving only one), at least under conditions with certain outcomes and clear differences between the reference and outcome quantities. Despite the small number of rats in this study, this species-level capacity suggests that reference dependence in general and loss aversion in particular may be conserved traits that evolved at or before the emergence of mammals.

Nutrition, epigenetics, and diseases.

Increasing epidemiological evidence suggests that maternal nutrition and environmental exposure early in development play an important role in susceptibility to disease in later life. In addition, these disease outcomes seem to pass through subsequent generations. Epigenetic modifications provide a potential link between the nutrition status during critical periods in development and changes in gene expression that may lead to disease phenotypes. An increasing body of evidence from experimental animal studies supports the role of epigenetics in disease susceptibility during critical developmental periods, including periconceptional period, gestation, and early postnatal period. The rapid improvements in genetic and epigenetic technologies will allow comprehensive investigations of the relevance of these epigenetic phenomena in human diseases.

Combined administration of testosterone plus an ornithine decarboxylase inhibitor as a selective prostate-sparing anabolic therapy.

Because of its anabolic effects on muscle, testosterone is being explored as a function-promoting anabolic therapy for functional limitations associated with aging; however, concerns about testosterone's adverse effects on prostate have inspired efforts to develop strategies that selectively increase muscle mass while sparing the prostate. Testosterone's promyogenic effects are mediated through upregulation of follistatin. We show here that the administration of recombinant follistatin (rFst) increased muscle mass in mice, but had no effect on prostate mass. Consistent with the results of rFst administration, follistatin transgenic mice with constitutively elevated follistatin levels displayed greater muscle mass than controls, but had similar prostate weights. To elucidate signaling pathways regulated differentially by testosterone and rFst in prostate and muscle, we performed microarray analysis of mRNAs from prostate and levator ani of castrated male mice treated with vehicle, testosterone, or rFst. Testosterone and rFst shared the regulation of many transcripts in levator ani; however, in prostate, 593 transcripts in several growth-promoting pathways were differentially expressed after testosterone treatment, while rFst showed a negligible effect with only 9 transcripts differentially expressed. Among pathways that were differentially responsive to testosterone in prostate, we identified ornithine decarboxylase (Odc1), an enzyme in polyamine biosynthesis, as a testosterone-responsive gene that is unresponsive to rFst. Accordingly, we administered testosterone with and without α-difluoromethylornithine (DFMO), an Odc1 inhibitor, to castrated mice. DFMO selectively blocked testosterone's effects on prostate, but did not affect testosterone's anabolic effects on muscle. Co-administration of testosterone and Odc1 inhibitor presents a novel therapeutic strategy for prostate-sparing anabolic therapy.

The effects of testosterone deprivation and supplementation on proteasomal and autophagy activity in the skeletal muscle of the male mouse: differential effects on high-androgen responder and low-androgen responder muscle groups.

Men with prostate cancer who receive androgen deprivation therapy show profound skeletal muscle loss. We hypothesized that the androgen deficiency activates not only the ubiquitin-proteasome systems but also the autophagy and affects key aspects of the molecular cross talk between protein synthesis and degradation. Here, 2-month-old male mice were castrated and treated with either testosterone (T) propionate or vehicle for 7 days (short term) or 43 days (long term), and with and without hydroxyflutamide. Castrated mice showed rapid and profound atrophy of the levator ani muscle (high androgen responder) at short term and lesser atrophy of the triceps muscle (low androgen responder) at long term. Levator ani and triceps muscles of castrated mice showed increased level of autophagy markers and lysosome enzymatic activity; only the levator ani showed increased proteasomal enzymatic activity. The levator ani muscle of the castrated mice showed increased level and activation of forkhead box protein O3A, the inhibition of mechanistic target of rapamicyn, and the activation of tuberous sclerosis complex protein 2 and 5'-AMP-activated protein kinase. Similar results were obtained in the triceps muscle of castrated mice. T rescued the loss of muscle mass after orchiectomy and inhibited lysosome and proteasome pathways dose dependently and in a seemingly IGF-I-dependent manner. Hydroxyflutamide attenuated the effect of T in the levator ani muscle of castrated mice. In conclusion, androgen deprivation in adult mice induces muscle atrophy associated with proteasomal and lysosomal activity. T optimizes muscle protein balance by modulating the equilibrium between mechanistic target of rapamicyn and 5'-AMP-activated protein kinase pathways.

Current trends in the development and application of molecular technologies for cancer epigenetics.

Current progress in epigenetic research supports the view that diet and dietary components are important in cancer etiology by enhancing or inhibiting carcinogenesis. Since diet and dietary factors may significantly contribute to the causation and progression of many cancers, it is important to find the molecular mechanisms of action of such dietary factors for cancer prevention and treatment. Recently, the role of epigenetic mechanisms in the cancer development and progression has attracted more attention as additional evidence along with traditional DNA sequence based mechanisms such as mutations and structural re-arrangements. Such an increasing interest in cancer epigenetics has also accelerated the development and application of molecular assays and tools for DNA methylation detection and histone modification enrichment analysis. In this paper, key assays and methods for epigenetic research are reviewed and discussed in terms of their utility and usability. In addition, more advanced methods for genome-wide analysis are introduced as part of upcoming research trends and directions.

GIT1 is associated with ADHD in humans and ADHD-like behaviors in mice.

Attention deficit hyperactivity disorder (ADHD) is a psychiatric disorder that affects ~5% of school-aged children; however, the mechanisms underlying ADHD remain largely unclear. Here we report a previously unidentified association between G protein-coupled receptor kinase-interacting protein-1 (GIT1) and ADHD in humans. An intronic single-nucleotide polymorphism in GIT1, the minor allele of which causes reduced GIT1 expression, shows a strong association with ADHD susceptibility in humans. Git1-deficient mice show ADHD-like phenotypes, with traits including hyperactivity, enhanced electroencephalogram theta rhythms and impaired learning and memory. Hyperactivity in Git1(-/-) mice is reversed by amphetamine and methylphenidate, psychostimulants commonly used to treat ADHD. In addition, amphetamine normalizes enhanced theta rhythms and impaired memory. GIT1 deficiency in mice leads to decreases in ras-related C3 botulinum toxin substrate-1 (RAC1) signaling and inhibitory presynaptic input; furthermore, it shifts the neuronal excitation-inhibition balance in postsynaptic neurons toward excitation. Our study identifies a previously unknown involvement of GIT1 in human ADHD and shows that GIT1 deficiency in mice causes psychostimulant-responsive ADHD-like phenotypes.

Folate supplementation differently affects uracil content in DNA in the mouse colon and liver.

High folate intake may increase the risk of cancer, especially in the elderly. The present study examined the effects of ageing and dietary folate on uracil misincorporation into DNA, which has a mutagenic effect, in the mouse colon and liver. Old (18 months; n 42) and young (4 months; n 42) male C57BL/6 mice were pair-fed with four different amino acid-defined diets for 20 weeks: folate deplete (0 mg/kg diet); folate replete (2 mg/kg diet); folate supplemented (8 mg/kg diet); folate deplete (0 mg/kg diet) with thymidine supplementation (1·8 g/kg diet). Thymidylate synthesis from uracil requires folate, but synthesis from thymidine is folate independent. Liver folate concentrations were determined by the Lactobacillus casei assay. Uracil misincorporation into DNA was measured by a GC/MS method. Liver folate concentrations demonstrated a stepwise increase across the spectrum of dietary folate levels in both old (P = 0·003) and young (P < 0·001) mice. Uracil content in colonic DNA was paradoxically increased in parallel with increasing dietary folate among the young mice (P trend = 0·033), but differences were not observed in the old mice. The mean values of uracil in liver DNA, in contrast, decreased with increasing dietary folate among the old mice, but it did not reach a statistically significant level (P < 0·1). Compared with the folate-deplete group, thymidine supplementation reduced uracil misincorporation into the liver DNA of aged mice (P = 0·026). The present study suggests that the effects of folate and thymidine supplementation on uracil misincorporation into DNA differ depending on age and tissue. Further studies are needed to clarify the significance of increased uracil misincorporation into colonic DNA of folate-supplemented young mice.

The dynamic structure of the estrogen receptor.

The estrogen receptor (ER) mediates most of the biological effects of estrogens at the level of gene regulation by interacting through its site-specific DNA and with other coregulatory proteins. In recent years, new information regarding the dynamic structural nature of ER has emerged. The physiological effects of estrogen are manifested through ER's two isoforms, ER(α) and ER(ß). These two isoforms (ER(α) and ER(ß)) display distinct regions of sequence homology. The three-dimensional structures of the DNA-binding domain (DBD) and ligand-binding domain (LBD) have been solved, whereas no three-dimensional natively folded structure for the ER N-terminal domain (NTD) is available to date. However, insights about the structural and functional correlations regarding the ER NTD have recently emerged. In this paper, we discuss the knowledge about the structural characteristics of the ER in general and how the structural features of the two isoforms differ, and its subsequent role in gene regulation.

Ageing, chronic alcohol consumption and folate are determinants of genomic DNA methylation, p16 promoter methylation and the expression of p16 in the mouse colon.

Older age, dietary folate and chronic alcohol consumption are important risk factors for the development of colon cancer. The present study examined the effects of ageing, folate and alcohol on genomic and p16-specific DNA methylation, and p16 expression in the murine colon. Old (aged 18 months; n 70) and young (aged 4 months; n 70) male C57BL/6 mice were pair-fed either a Lieber-DeCarli liquid diet with alcohol (18 % of energy), a Lieber-DeCarli diet with alcohol (18 %) and reduced folate (0.25 mg folate/l) or an isoenergetic control diet (0.5 mg folate/l) for 5 or 10 weeks. Genomic DNA methylation, p16 promoter methylation and p16 gene expression were analysed by liquid chromatography-MS, methylation-specific PCR and real-time RT-PCR, respectively. Genomic DNA methylation was lower in the colon of old mice compared with young mice (P < 0.02) at 10 weeks. Alcohol consumption did not alter genomic DNA methylation in the old mouse colon, whereas it tended to decrease genomic DNA methylation in young mice (P = 0.08). p16 Promoter methylation and expression were higher in the old mouse colon compared with the corresponding young groups. There was a positive correlation between p16 promoter methylation and p16 expression in the old mouse colon (P < 0.02). In young mice the combination of alcohol and reduced dietary folate led to significantly decreased p16 expression compared with the control group (P < 0.02). In conclusion, ageing and chronic alcohol consumption alter genomic DNA methylation, p16 promoter methylation and p16 gene expression in the mouse colon, and dietary folate availability can further modify the relationship with alcohol in the young mouse.

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.

Moderate folate depletion modulates the expression of selected genes involved in cell cycle, intracellular signaling and folate uptake in human colonic epithelial cell lines.

Folate deficiency may affect gene expression by disrupting DNA methylation patterns or by inducing base substitution, DNA breaks, gene deletions and gene amplification. Changes in expression may explain the inverse relationship observed between folate status and risk of colorectal cancer. Three cell lines derived from the normal human colon, HCEC, NCM356 and NCM460, were grown for 32-34 days in media containing 25, 50, 75 or 150 nM folic acid, and the expression of genes involved in cell-cycle checkpoints, intracellular signaling, folate uptake and cell adhesion and migration was determined. Expression of Folate Receptor 1 was increased with decreasing media folate in all cell lines, as was p53, p21, p16 and beta-catenin. With decreasing folate, the expression of both E-cadherin and SMAD-4 was decreased in NCM356. APC was elevated in NCM356 but unchanged in the other lines. No changes in global methylation were detected. A significant increase in p53 exon 7-8 strand breaks was observed with decreasing folate in NCM460 cells. The changes observed are consistent with DNA damage-induced activation of cell-cycle checkpoints and cellular adaptation to folate depletion. Folate-depletion-induced changes in the Wnt/APC pathway as well as in genes involved in cell adhesion, migration and invasion may underlie observed relationships between folate status and cancer risk.

Mild depletion of dietary folate combined with other B vitamins alters multiple components of the Wnt pathway in mouse colon.

Preclinical and clinical studies suggest that diminished folate status increases the risk of colorectal carcinogenesis. However, many biochemical functions of folate are dependent on the adequate availability of other 1-carbon nutrients, including riboflavin, vitamin B-6, and vitamin B-12. Aberrations in the Wnt pathway are thought to play an important role in human colorectal cancers. This study therefore investigated if mild depletion of folate combined with depletion of riboflavin, vitamin B-6, and vitamin B-12 could induce alterations in the Wnt pathway in the colonic mucosa. Ninety-six mice were pair-fed diets with different combinations of B vitamin depletion for 10 wk. Genomic DNA methylation and uracil misincorporation were measured by LC/MS and GC/MS. Gene-specific methylation, strand breaks, and expressions were measured by real-time PCR and immunoblotting. Proliferation and apoptosis were determined by immunohistochemistry. DNA strand breaks within the Apc mutation cluster region were induced by folate depletion combined with inadequacies of riboflavin, vitamin B-6, and vitamin B-12 (P < 0.05), but such effects were not induced by folate depletion alone. Similarly, minor changes in the expression of Apc, beta-catenin, and cyclin D1 produced by mild folate depletion were significantly magnified by multiple vitamin depletion. Apoptosis, which can be suppressed by increased Wnt-signaling, was attenuated by the combined deficiency state (P < 0.05) but not by singlet or doublet deficiencies. These findings indicate that a mild depletion of folate that is of insufficient magnitude by itself to induce alterations in components of the Wnt pathway may produce such effects when present in conjunction with mild inadequacies of other 1-carbon nutrients.

Older age and dietary folate are determinants of genomic and p16-specific DNA methylation in mouse colon.

Older age and inadequate folate intake are strongly implicated as important risk factors for colon cancer and each is associated with altered DNA methylation. This study was designed to determine the effects of aging and dietary folate on select features of DNA methylation in the colon that are relevant to carcinogenesis. Old (18 mo; n = 34) and young (4 mo; n = 32) male C57BL/6 mice were randomly divided into 3 groups and fed diets containing 0, 4.5, or 18 mumol folate/kg (deplete, replete, and supplemented groups, respectively) for 20 wk. Genomic DNA methylation and p16 promoter methylation in the colonic mucosa were analyzed by liquid chromatography/electrospray ionization/MS and methylation-specific PCR, respectively. p16 gene expression was determined by real-time RT-PCR. Old mice had significantly lower genomic DNA methylation compared with young mice at each level of dietary folate (4.5 +/- 0.2, 4.8 +/- 0.1, and 4.9 +/- 0.1 vs. 6.0 +/- 0.1, 5.3 +/- 0.2, and 5.9 +/- 0.2%, in folate-deplete, -replete, and -supplemented groups, respectively, P < 0.05) and markedly higher p16 promoter methylation (61.0 +/- 2.7, 69.7 +/- 6.9, and 87.1 +/- 13.4 vs. 10.8 +/- 3.6, 8.4 +/- 1.8, and 4.9 +/- 1.7%, respectively, P < 0.05). In old mice, genomic and p16 promoter DNA methylation each increased in a manner that was directly related to dietary folate (P(trend) = 0.009). Age-related enhancement of p16 expression occurred in folate-replete (P = 0.001) and folate-supplemented groups (P = 0.041), but not in the folate-deplete group. In conclusion, aging decreases genomic DNA methylation and increases promoter methylation and expression of p16 in mouse colons. This effect is dependent on the level of dietary folate.

Oestrogen replacement therapy reduces total plasma homocysteine and enhances genomic DNA methylation in postmenopausal women.

Although oestrogen replacement therapy (ERT), which can affect the risk of major cancers, has been known to reduce total plasma homocysteine concentrations in postmenopausal women, the mechanisms and subsequent molecular changes have not yet been defined. To investigate the effect of ERT on homocysteine metabolism, thirteen healthy postmenopausal women were enrolled in a double-blind, placebo-controlled, randomized, cross-over study consisting of two 8-week long phases, placebo and conjugated equine oestrogen (CEE; 0.625 mg/d). Concentrations of total plasma homocysteine, vitamin B6 and serum folate and vitamin B12 were measured by conventional methods. Genomic DNA methylation was measured by a new liquid chromatography/MS method and promoter methylation status of the oestrogen receptor (ER)alpha, ERbeta and p16 genes was analysed by methylation-specific PCR after bisulfite treatment. The CEE phase demonstrated a significantly decreased mean of total plasma homocysteine concentrations compared with the placebo phase (8.08 micromol/l (6.82-9.39) v. 9.29 (7.53-11.35), P < 0.05) but there was no difference in the blood concentrations of the three B vitamins. The CEE phase also showed a significantly increased genomic DNA methylation in peripheral mononuclear cells compared with the placebo phase (2.85 (SD 0.12) ng methylcytosine/microg DNA v. 2.40 +/- (SD 0.15) P < 0.05). However, there was no difference in promoter methylation in the ERalpha, ERbeta and p16 genes. This study demonstrates that decreased homocysteinaemia by CEE therapy parallels with increased genomic DNA methylation, suggesting a potential new candidate mechanism by which ERT affects the risk of cancers and a possible new candidate biomarker for the oestrogen-related carcinogenesis through folate-related one-carbon metabolism.

Chronic cigarette smoking is associated with diminished folate status, altered folate form distribution, and increased genetic damage in the buccal mucosa of healthy adults.

BACKGROUND: Smoking causes genetic damage in buccal cells and increases the risk of oral cancer. Because folate is instrumental in DNA synthesis and repair, it is a determinant of genetic stability and therefore might attenuate the genotoxic effects of smoking. OBJECTIVE: Our aim was to compare the presence of folate metabolites and select indicators of genetic damage in the mouths of chronic smokers and nonsmokers. DESIGN: Dietary, biochemical, and molecular correlates of folate status were measured in healthy smoker (n = 35) and nonsmoker (n = 21) groups of comparable age, sex, and body mass indexes. RESULTS: After correction for dietary intake, the smokers displayed lower plasma, erythrocyte, and buccal mucosal cell (BMC) folate (20%, 32%, and 50% lower, respectively; P < 0.05) and lower plasma vitamin B-12 and pyridoxal 5-phosphate (P < 0.05) than did nonsmokers. Folate in the BMCs of smokers comprised significantly greater proportions of pteroylmonoglutamate, formyltetrahydrofolate, and 5,10-methenyltetrahyrofolate than did folate in the BMCs of nonsmokers. Although the degree of genomic methylation and uracil incorporation in the buccal cells of the 2 groups were not significantly different, the BMC micronucleus index, a cytologic indicator of genetic damage, in the smokers was 2-fold that of the nonsmokers (9.57 compared with 4.44 micronuclei/1000 cells; P < 0.0001). Neither systemic nor oral folate status was an independent predictor of micronuclei. CONCLUSIONS: Chronic smoking is associated with a lower systemic status of several B vitamins, reduced oral folate, and changes in folate form distribution in the mouth. However, the cytologic damage that is evident in the mouths of smokers does not correlate with oral folate status.

Genetic and epigenetic interactions between folate and aging in carcinogenesis.

Folate is among the most strongly implicated dietary components to convey protection against colon cancer, and diminished folate status is associated with an enhanced risk of colon cancer. Age is also regarded as one of the most important risk factors for colonic carcinogenesis. It is therefore of considerable interest to determine whether the process of aging influences folate metabolism in the colon and whether folate supplementation might prevent the procarcinogenic effects associated with aging. Recent studies in our laboratory demonstrated that the colonic mucosa of elder rats is more susceptible to folate depletion than that of young rats. Depletion of folate results in a shift in the forms of folate in the colon as well as increased uracil incorporation into DNA, a purported mechanism for colonic carcinogenesis. However, modest folate supplementation eliminates evidence of inadequate folate status in the colons of elder rats, suggesting that the relation between age and folate status in the colon might be one mechanism by which aging modulates colorectal cancer risk. Interactions between folate and aging also affect a spectrum of epigenetic and genetic phenomena such as uracil misincorporation, DNA methylation, protein methylation, mitochondrial deletion, and critical gene expression, which could be related to carcinogenesis. Aging and inadequate dietary folate may interact and collectively induce derangements in folate metabolism, thereby provoking subsequent molecular aberrations, which may enhance carcinogenesis. However, folate supplementation appears to reverse these adverse effects of aging, which is potentially of substantial import because the latter is an unmodifiable risk factor.