The relation of plasma homocysteine to radiographic knee osteoarthritis.

OBJECTIVE: Homocysteine has been implicated in multiple diseases that involve changes in structural tissue. In vitro studies have found that it alters the structure of collagen cross-linking thus affecting stability and mineralization such as that occurring in bone tissue. In the present study we considered the possible relationship between plasma homocysteine levels and the development and progression of knee osteoarthritis (OA). METHODS: The study question was posed in 691 men and 966 women from the original and offspring cohorts of the Framingham Osteoarthritis Study. We divided individuals into three groups according to plasma homocysteine levels and compared their risk for the development of new and progression of existing OA. We adjusted for potential confounders including age, body mass index, weight change, and physical activity. RESULTS: In the crude analysis, men in the middle homocysteine tertile were found to be at a greater risk than men in the lowest tertile for incident OA [odds ratios of 1.9 (1.1-3.5)]. This result persisted after adjusting for covariates [odds: 2.0, (1.1-3.8)]. No significant correlation was seen in women for the development of OA. In the evaluation of progression no significant trends were seen for both men and women. CONCLUSIONS: Although cellular and molecular studies of homocysteine-related pathophysiology suggest a possible correlation between plasma homocysteine levels and OA, the present clinical study did not conclusively demonstrate such an association. However, further research is needed to explore the role of homocysteine in specific aspects of OA etiopathogenesis.

Inhibition of folate enzymes by sulfasalazine.

Sulfasalazine (salicylazosulfapyridine), an agent widely used for the treatment of ileitis and colitis, is also a competitive inhibitor of intestinal folate transport (1, 2). The mechanism of action of sulfasalazine remains uncertain. To further explore the mechanism of sulfasalazine action, the interaction of the drug with the folate recognition site was tested with three enzymes: dihydrofolate reductase, methylenetetrahydrofolate reductase, and serine transhydroxymethylase, each catalyzing a reaction involving a different folate derivative. Each of these enzymes was inhibited by sulfasalazine in the same concentration range as that previously observed to inhibit intestinal folate transport; the kinetic data are consistent with a competitive mode of inhibition. Specificity of inhibition was demonstrated by the finding that the reduction of the pteridine ring of pteroylheptaglutamic acid by dihydrofolate reductase was subject to inhibition, whereas the hydrolysis of the gamma-glutamyl peptide side chain by chicken pancreas conjugase was not affected. These results are interpreted to indicate that sulfasalazine interferes with a folate recognition site which is common to these enzymes and to the intestinal transport system. Sulfasalazine, therefore, has certain properties of an antifolate drug.

An overview of folate status in a population-based study from São Paulo, Brazil and the potential impact of 10 years of national folic acid fortification policy.

BACKGROUND/OBJECTIVES: Food fortification is an important strategy in public health policy for controlling micronutrient malnutrition and a major contributing factor in the eradication of micronutrients' deficiencies. Approximately 50 countries worldwide have adopted food fortification with folic acid (FA). FA fortification of wheat and maize flours has been mandatory in Brazil since 2004. To assess the effect of 10 years of FA food fortification policy on folate status of residents of São Palo, Brazil using a population-based survey. SUBJECTS/METHODS: Data were from 750 individuals aged ⩾12 years who participated in a cross-sectional population-based survey in São Paulo city, Brazil. Fasting blood samples were collected, and folate was assayed by affinity-high performance liquid chromatografy method with electrochemical detection. The participants provided information about food intake based on two 24 h dietary recall. RESULTS: Only 1.76% of population had folate deficiency (<6.8 nmol/l). The mean folate concentration was 29.5 (95% confidence interval: 27.3-31.7) nmol/l for all sex-age groups. The mean folate intake for the population was 375.8 (s.e.m.=6.4) µg/day of dietary folate equivalents (DFEs). When comparing folate intake in DFE from food folate and FA from fortified foods, FA contributed 50% or more of the DFE in almost all sex-age groups. The major contributors of folate intake are processed foods made from wheat flour fortified with FA, especially among subjects younger than 20 years old. CONCLUSIONS: The deficiency of folate is very low, and food fortification contributed to folate intake and had a notable influence on rankings of food contributors of folate.

B-Vitamin Therapy for Kidney Transplant Recipients Lowers Homocysteine and Improves Selective Cognitive Outcomes in the Randomized FAVORIT Ancillary Cognitive Trial.

BACKGROUND: Objectives: Elevated plasma total homocysteine (tHcy) is associated with increased risk of cardiovascular disease, stroke and dementia. Results of clinical trials using B-vitamins to reduce the cognitive risks attributed to tHcy have been inconsistent. The high prevalence of both hyperhomocysteinemia and cognitive impairment among kidney transplant recipients makes them an important population in which to evaluate the effect of lowering homocysteine on cognitive function. We therefore evaluated whether B-vitamin therapy to lower tHcy would prevent cognitive-decline in a cohort of stable kidney transplant recipients. DESIGN: The study was a longitudinal ancillary of the FAVORIT trial, a randomized, placebo-controlled multi-site trial of high-dose B vitamins to reduce cardiovascular and cerebrovascular events in clinically stable kidney transplant recipients with elevated tHcy. PARTICIPANTS: 584 participants from 18 sites across North America. INTERVENTION: The intervention consisted of a daily multivitamin containing high-doses of folate (5.0 mg), vitamin B12 (1.0 mg) and vitamin B6 (50 mg). The placebo consisted of a daily multi-vitamin containing no folate and recommended daily allowances of vitamins B12 and B6 (0 mg folate; 2.0 µg vitamin B12; 1.4 mg vitamin B6). MEASUREMENTS: Annual neuropsychological assessment for up to 5 years (mean 3.3 years) using a standardized test battery. Efficacy was analyzed on an intention-to-treat basis using end-of-trial data. Subgroup analyses included stratification for baseline plasma B-vitamin and tHcy concentrations. RESULTS: At baseline, cognitive impairment was common with 61% of participants falling more than one standard deviation below published norms for at least one cognitive test. Fewer than 1% of participants had insufficient plasma folate < 5 ng/ml or vitamin B12 < 148 pmol/L. However, 44.6% had plasma B6 concentrations < 30 nmol/L. At follow-up, processing speed and memory scores were modestly but significantly better in the B-vitamin supplement group than in controls (p≤0.05). There was no interaction between baseline tHcy, B-vitamin status and treatment on the cognitive outcomes. CONCLUSIONS: High-dose B-vitamin supplementation provided modest cognitive benefit for kidney transplant recipients with elevated baseline tHcy. Since nearly all participants were folate and vitamin B12 sufficient at baseline, the potential cognitive benefits of folate and B12 supplementation in individuals with poor B-vitamin status remains to be determined.

Effects of choline deficiency and methotrexate treatment upon liver folate content and distribution.

We examined the effects of feeding rats a choline deficient diet, of treating rats with low doses of methotrexate (MTX, 0.1 mg/kg, daily), and of combined choline deficiency and MTX treatment upon the content and distribution of folates in liver. We used a newly devised technique for analysis of folates which utilized affinity chromatography followed by high pressure liquid chromatography. Compared to control rats, total hepatic folate content decreased by 31% in the choline deficient rats, by 48% in the MTX treated rats, and by 60% in rats which were both choline deficient and treated with MTX. In extracts of livers from control rats, folates were present predominantly as penta (35%) and hexaglutamyl (52%) derivatives. The pteridine ring structure distribution of these folates was as follows: 48% 5-methyltetrahydrofolate, 14% formylated tetrahydrofolate, and 39% tetrahydrofolate. In choline deficient animals, there was a decrease in the relative concentration of pentaglutamyl folates and an increase in the relative concentration of heptaglutamyl folates. In livers from MTX treated animals, MTX-polyglutamates with 2-5 glutamate residues accumulated. The consequences of MTX treatment were: a) an elongation of the glutamate chains of the folates as the proportion of hepta- and octaglutamyl derivatives was increased relative to penta- and hexaglutamyl folates; b) the occurrence of unreduced folic acid; c) a decrease in the relative concentration of 5-methyltetrahydrofolate and an increase in the relative concentration of formylated tetrahydrofolate, and d) no change in the relative concentrations of tetrahydrofolate. In livers from animals that were both choline deficient and treated with MTX, the tetrahydrofolate concentrations were 50% of control while formylated tetrahydrofolate concentrations increased 3-fold. These data are discussed from the standpoint of the current understanding of mechanisms that regulate the elongation of the glutamic acid chains of folates and those that regulate folate dependent synthesis and utilization of one carbon unit.

Folate deficiency enhances the development of colonic neoplasia in dimethylhydrazine-treated rats.

In patients with ulcerative colitis, epidemiological work has suggested an association between low folate status and an increased risk of colonic neoplasia. The aim of the present study was to determine if experimental folate deficiency increases the likelihood of developing neoplasia in rats treated with the carcinogen dimethylhydrazine. Weanling male Sprague-Dawley rats were fed with an amino acid-defined diet containing either 8 or 0 mg/kg folic acid. After 5 weeks of defined diet, weekly s.c. injections of dimethylhydrazine (20 mg/kg) were administered to both groups. Serum, whole blood, liver, and colonic folate concentrations at the time of sacrifice were significantly lower in folate-depleted animals (P less than 0.001). There were significant differences in the incidence of colonic neoplasia between the two groups after 20 weeks of dimethylhydrazine exposure: folate-deficient rats had a greater incidence of dysplasia (6 of 7 versus 2 of 7 animals; P less than 0.05) and carcinoma (6 of 7 versus 1 of 7 animals; P less than 0.01). Furthermore, a significantly greater proportion of folate-replete rats than folate-deficient rats were free of neoplastic lesions (5 of 7 versus 0 of 7 animals; P less than 0.05). These results suggest that, in this animal model, folate deficiency increases the risk of malignancy when there is an underlying predisposition to colorectal cancer.

Methylenetetrahydrofolate reductase polymorphism, dietary interactions, and risk of colorectal cancer.

Folate derivatives are important in experimental colorectal carcinogenesis; low folate intake, particularly with substantial alcohol intake, is associated with increased risk. The enzyme 5,10-methylenetetrahydrofolate reductase (MTHFR) catalyzes the conversion of 5,10-methylenetetrahydrofolate, required for purine and thymidine syntheses, to 5-methyltetrahydrofolate, the primary circulatory form of folate necessary for methionine synthesis. A common mutation (677C-->T) in MTHFR reduces enzyme activity, leading to lower levels of 5-methyltetrahydrofolate. To evaluate the role of folate metabolism in human carcinogenesis, we examined the associations of MTHFR mutation, plasma folate levels, and their interaction with risk of colon cancer. We also examined the interaction between genotype and alcohol intake. We used a nested case-control design within the Physicians' Health Study. Participants were ages 40-84 at baseline when alcohol intake was ascertained and blood samples were drawn. During 12 years of follow-up, we identified 202 colorectal cancer cases and matched them to 326 cancer-free controls by age and smoking status. We genotyped for the MTHFR polymorphism and measured plasma folate levels. Men with the homozygous mutation (15% in controls) had half the risk of colorectal cancer [odds ratio (OR), 0.49; 95% confidence interval (CI), 0.27-0.87] compared with the homozygous normal or heterozygous genotypes. Overall, we observed a marginal significant increased risk of colorectal cancer (OR, 1.78; 95% CI, 0.93-3.42) among those whose plasma folate levels indicated deficiency (<3 ng/ml) compared with men with adequate folate levels. Among men with adequate folate levels, we observed a 3-fold decrease in risk (OR, 0.32; 95% CI, 0.15-0.68) among men with the homozygous mutation compared with those with the homozygous normal or heterozygous genotypes. However, the protection due to the mutation was absent in men with folate deficiency. In men with the homozygous normal genotype who drank little or no alcohol as reference, those with the homozygous mutation who drank little or no alcohol had an 8-fold decrease in risk (OR, 0.12; 95% CI, 0.03-0.57), and for moderate drinkers, a 2-fold decrease in risk (OR, 0.42; 95% CI, 0.15-1.20); no decrease in risk was seen in those drinking 1 or more drinks/day. Our findings provide support for an important role of folate metabolism in colon carcinogenesis. In particular, these results suggest that the 677C-->IT mutation in MTHFR reduces colon cancer risk, perhaps by increasing 5,10-methylenetetrahydrofolate levels for DNA synthesis, but that low folate intake or high alcohol consumption may negate some of the protective effect.

Disruption of thymidylate synthesis and glycine-serine interconversion by L-methionine and L-homocystine in Raji cells.

Excessive concentrations of L-methionine inhibited the folate-dependent de novo synthesis of thymidylic acid (TMP) in Raji cells, demonstrating the usefulness of this cell line for the study of methionine-folate antagonism. The effect was also produced by L-homocystine but not by other amino acids including D-methionine and L-ethionine, suggesting that this effect is exerted by a common intermediate of methionine and homocystine metabolism. L-Methionine, L-homocysteine, S-adenosylmethionine (SAM), and S-adenosylhomocysteine (SAH) are not inhibitors of thymidylate synthase activity. On the other hand the capacity of the cells to incorporate serine 3-carbon and glycine 2-carbon into DNA is impaired by the presence of L-methionine or L-homocystine. Studies with cell-free extracts demonstrated that the glycine cleavage enzyme is inhibited by 45% by L-methionine, L-homocysteine, SAM or SAH. Serine hydroxymethylase on the other hand was slightly stimulated by these sulfur-containing compounds and this stimulation was shown to occur in the intact cell as well. These findings suggest that when levels of L-methionine metabolites are elevated, there is an increase in the use of glycine to maintain the intracellular concentration of serine, which is required for homocysteine detoxification by conversion to cystathionine. The reduction in TMP synthesis caused by excess L-methionine or L-homocystine may result from increased utilization of one-carbon units for serine synthesis.

Effect of anions on folate binding by isolated brush border membranes from rat kidney.

The characteristics of folate binding by brush border membranes from rat kidney homogenates were investigated. At pH 7.4, binding of [3',5',9-3H]-pteroylglutamic acid to membranes containing endogenous folate is inhibited by anions, with chloride being most effective followed by bromide, thiocyanate, iodide, phosphate and sulfate. A maximum inhibition of 70-75% is attained at a concentration of 0.1 M chloride and an incubation time of 30 min. The inhibition diminishes with increased incubation time and at 24 h is negligible. The binding of [3',5',9-3H]pteroylglutamic acid to brush border membranes stripped of endogenous folate by acid treatment is not inhibited by anions. Anion sensitivity can be restored to these treated membranes by reconstitution with membrane-derived folate, particularly 5-methyltetrahydropteroyl-glutamic acid, or by preincubation with synthetic 5-methyltetrahydropteroyl-glutamic acid. Inhibition of [3',5',9-3H]pteroylglutamic acid binding by anions in membranes with endogenous folate is best explained by an anion-induced stabilization of endogenous folate-binding protein complex resulting in a decreased rate of exchange with exogenous [3',5',9-3H]pteroylglutamic acid.

Carrier affinity as a mechanism for the pH-dependence of folate transport in the small intestine.

A mildly acidic pH in the lumen of the small intestine markedly enhances the transport of folate. This study investigated the relationship between pH and the affinity between folic acid and the apical membrane transporter using brush border membrane vesicles from rat jejunum and differentiated monolayer cultures of the colon carcinoma cell line, CaCo-2. Uptake studies with BBMV were conducted at folic acid concentrations of 0.1 to 50 mumol/l, conditions which were suitable for analyzing uptake data based on the Michaelis-Menten equation modified to include a nonsaturable component. These analyses yielded apparent Km values of 0.6 and 12.3 microM at pH 5.5 and pH 7.4, respectively (P less than 0.05). Values for Vmax were lower at pH 5.5 than at pH 7.4 (0.8 vs. 1.6 pmol/mg protein per 10 s, P less than 0.05). The studies with CaCo-2 cells employed folic acid concentrations of 0.1 to 5 mumol/l. Under these conditions the apparent Km for folic uptake was lowest at pH 6.0, where the Km was 0.7 mumol/l. The apparent Km increased sharply as a neutral pH was approached; reaching a value of 13.9 mumol/l at pH 7.1. These data suggest that the prominent pH effect on intestinal folate transport is, in part, explained by an increased affinity of the folate substrate for its membrane transporter.

Low circulating vitamin B(6) is associated with elevation of the inflammation marker C-reactive protein independently of plasma homocysteine levels.

BACKGROUND: Lower vitamin B(6) concentrations are reported to confer an increased and independent risk for cardiovascular disease (CVD). The mechanism underlying this relationship, however, remains to be defined. Other diseases, such as rheumatoid arthritis, are associated with reduced vitamin B(6) levels. Despite a clear distinction in pathophysiology, inflammatory reaction may be the major link between these diseases. We hypothesized a relationship between pyridoxal 5'-phosphate (PLP), the active form of vitamin B(6), and the marker of inflammation C-reactive protein (CRP). We also evaluated whether total plasma homocysteine (tHcy), a well-defined risk factor for CVD and a major determinant of plasma PLP levels, had a possible role as a mediator of this hypothesized relationship. METHODS AND RESULTS: Data from 891 participants from the population-based Framingham Heart Study cohort were analyzed. Subjects were divided into 2 groups according to normal or elevated CRP values: group 1, CRP <6 mg/L; group 2, CRP >/=6 mg/L. Plasma PLP levels were substantially lower in group 2 than in group 1 (mean values in group 2, 36.5 nmol/L versus 55.8 nmol/L in group 1, P<0.001). In a multiple logistic regression model adjusted for tHcy, the association of PLP with CRP remained highly significant (P=0.003). CONCLUSIONS: Low plasma PLP is associated with higher CRP levels independently of tHcy. This observation may reflect a vitamin B(6) utilization in the presence of an underlying inflammatory process and represent a possible mechanism to explain the decreased vitamin B(6) levels in CVD.

Plasma homocysteine and parental myocardial infarction in young adults in Jerusalem.

BACKGROUND: A causal role for mildly elevated plasma homocysteine (tHcy) in cardiovascular disease remains undetermined. To address the unresolved issue of the antecedent-consequent directionality of the relationship, we assessed the familial association of tHcy with parental myocardial infarction (MI) in young Israeli men and women. We also compared tHcy concentrations in Jerusalem, where rates of coronary heart disease (CHD) are high, with the United States Third National Health and Examination Survey (NHANES III). METHODS AND RESULTS: A total of 8646 17-year-olds and 6952 parents were examined from 1976 to 1979 in Jerusalem. At ages 28 to 32 years, offspring of parents who experienced a documented MI during a 10-year follow-up (n=133 men, 62 women; 72% response) and offspring of CHD-free parents (n=389 men, 208 women; 71% response) were reexamined. tHcy levels were determined by the same laboratory for the NHANES non-Hispanic white population aged 25 to 34 years (n=379) and the Jerusalem population sample (n=858). Men from Jerusalem, but not women, had clearly higher tHcy levels than the sample from the United States (90th percentile, 23 versus 14 micromol/L). This difference was largely attributable to lower plasma vitamin B12 levels in the Israeli population. Male case offspring had higher adjusted tHcy than did controls (1.9 micromol/L, P=0.002). Logistic modeling revealed a graded increase in risk of parental MI across quintiles of offspring tHcy, with an adjusted odds ratio of 2.7 in the 5th quintile (P=0.0026 for trend). CONCLUSIONS: The higher tHcy in young male offspring of parents with CHD suggests that elevated tHcy precedes manifestation of CHD. The elevated population tHcy in men may contribute to the high incidence of CHD in Israel.

Controlled comparison of L-5-methyltetrahydrofolate versus folic acid for the treatment of hyperhomocysteinemia in hemodialysis patients.

BACKGROUND: The hyperhomocysteinemia regularly found in hemodialysis patients is largely refractory to combined oral B-vitamin supplementation featuring supraphysiological doses of folic acid. We evaluated whether a high-dose L-5-methyltetrahydrofolate-based regimen provided improved total homocysteine (tHcy)-lowering efficacy in chronic hemodialysis patients. METHODS AND RESULTS: We block-randomized 50 chronic, stable hemodialysis patients on the basis of their screening predialysis tHcy levels, sex, and dialysis center into 2 groups of 25 subjects treated for 12 weeks with oral folic acid at 15 mg/d (FA group) or an equimolar amount (17 mg/d) of oral L-5-methyltetrahydrofolate (MTHF group). All 50 subjects also received 50 mg/d of oral vitamin B(6) and 1.0 mg/d of oral vitamin B(12). The mean percent reductions (+/-95% CIs) in predialysis tHcy were not significantly different: MTHF, 17.0% (12.0% to 22.0%); FA, 14.8% (9.6% to 20.1%); P=0.444 by matched ANCOVA adjusted for pretreatment tHcy. Final on-treatment values (mean with 95% CI) were MTHF, 20.0 micromol/L (18.8 to 21.2 micromol/L); FA, 19.5 micromol/L (18.3 to 20.7 micromol/L). Moreover, neither treatment resulted in "normalization" of tHcy levels (ie, final on-treatment values <12 micromol/L) among a significantly different or clinically meaningful number of patients: MTHF, 2 of 25 (8%); FA, 0 of 25 (0%); Fisher's exact test of between-groups difference, P=0.490. CONCLUSIONS: Relative to high-dose folic acid, high-dose oral L-5-methyltetrahydrofolate-based supplementation does not afford improved tHcy-lowering efficacy in hemodialysis patients. The preponderance of hemodialysis patients (ie, >90%) exhibit mild hyperhomocysteinemia refractory to treatment with either regimen. This treatment refractoriness is not related to defects in folate absorption or circulating plasma and tissue distribution.

Effect of dietary patterns on serum homocysteine: results of a randomized, controlled feeding study.

BACKGROUND: Elevated blood levels of homocysteine are associated with an increased risk of atherosclerotic cardiovascular disease. Although numerous studies have assessed the impact of vitamin supplements on homocysteine, the effect of dietary patterns on homocysteine has not been well studied. METHODS AND RESULTS: During a 3-week run-in, 118 participants were fed a control diet, low in fruits, vegetables, and dairy products, with a fat content typical of US consumption. During an 8-week intervention phase, participants were then fed 1 of 3 randomly assigned diets: the control diet, a diet rich in fruits and vegetables but otherwise similar to control, or a combination diet rich in fruits, vegetables, and low-fat dairy products and reduced in saturated and total fat. Between the end of run-in and intervention periods, mean change in homocysteine was +0.46 micromol/L in the control diet, +0.21 micromol/L in the fruits and vegetables diet (P=0.47 compared with control), and -0.34 micromol/L in the combination diet (P=0.03 compared with control, P=0.12 compared with the fruits and vegetables diet). In multivariable regression models, change in homocysteine was significantly and inversely associated with change in serum folate (P=0.03) but not with change in serum vitamin B(12) (P=0.64) or pyridoxal 5' phosphate, the coenzyme form of vitamin B(6) (P=0.83). CONCLUSIONS: Modification of dietary patterns can have substantial effects on fasting levels of total serum homocysteine. These results provide additional insights into the mechanisms by which diet might influence the occurrence of atherosclerotic cardiovascular disease.

Renal insufficiency, vitamin B(12) status, and population attributable risk for mild hyperhomocysteinemia among coronary artery disease patients in the era of folic acid-fortified cereal grain flour.

Fortification of enriched cereal grain flour products with folic acid has drastically reduced the prevalence of deficient plasma folate status, a major determinant of plasma total homocysteine (tHcy) levels. We hypothesized that even more liberally defined "suboptimal" plasma folate status might no longer contribute importantly to the population attributable risk (PAR) for mild hyperhomocysteinemia, a putative atherothrombotic risk factor. We determined fasting plasma tHcy, folate, vitamin B(12), and pyridoxal 5'-phosphate levels, along with serum creatinine and albumin levels, in 267 consecutive patients (aged 61+/-9 [mean+/-SD] years, 76.4% men and 26.6% women) with stable coronary artery disease (CAD) who were nonusers of vitamin supplements or had abstained from supplement use for at least 6 weeks before examination. Subjects were evaluated a minimum of 3 months after the implementation of flour fortification was largely completed. Relative risk estimates for the calculation of PAR were derived from a multivariable-adjusted logistic regression model with >/=12 micromol/L tHcy as the dependent variable and with age, sex, pyridoxal 5'-phosphate (continuous), albumin (continuous), <5 ng/mL folate, <250 pg/mL vitamin B(12), and >/=1.3 mg/dL creatinine as the independent variables. The prevalence of >/=12 micromol/L plasma tHcy was 11.2% (30 of 267 patients). PAR estimates (percentage) for >/=12 micromol/L tHcy were as follows: <5 ng/mL folate (<1%), <250 pg/mL vitamin B(12) (24.5%), and >/=1.3 mg/dL creatinine (37.5%). In the era of folic acid-fortified cereal grain flour, renal insufficiency and suboptimal vitamin B(12) status (but not folate status) contribute importantly to the PAR for mild hyperhomocysteinemia among patients with stable CAD.

Nonfasting plasma total homocysteine levels and all-cause and cardiovascular disease mortality in elderly Framingham men and women.

BACKGROUND: Elevated fasting total homocysteine (tHcy) levels were recently shown to confer an independent risk for all-cause and cardiovascular disease (CVD) mortality among selected Norwegian patients with confirmed coronary heart disease. We examined whether elevated fasting plasma tHcy levels were predictive of all-cause and CVD mortality in a large, population-based sample of elderly US women and men. METHODS: Nonfasting plasma tHcy levels were determined in 1933 elderly participants (mean age, 70 +/- 7 years; 58.9% women) from the original Framingham Study cohort, examined between 1979 and 1982, with follow-up through 1992. Unadjusted and adjusted (ie, for age, sex, diabetes, smoking, systolic blood pressure, total and high-density lipoprotein cholesterol, and creatinine) relative risk estimates (with 95% confidence intervals [CIs]) for total and CVD mortality were generated by proportional hazards modeling, with tHcy levels (quartiles) as the independent variable. RESULTS: There were 653 total deaths and 244 CVD deaths during a median follow-up of 10.0 years. Proportional hazards modeling revealed that tHcy levels of 14.26 micromol/L or greater (the upper quartile), vs less than 14.26 micromol/L (the lower three quartiles), were associated with relative risk estimates of 2.18 (95% CI, 1.86-2.56) and 2.17 (95% CI, 1.68-2.82) for all-cause and CVD mortality, respectively. The relative risk estimates after adjustment for age, sex, systolic blood pressure, diabetes, smoking, and total and high-density lipoprotein cholesterol levels attenuated these associations, but they remained significant: 1.54 (95% CI, 1.31-1.82) for all-cause mortality; 1.52 (95% CI, 1.16-1.98) for CVD mortality. CONCLUSION: Elevated nonfasting plasma tHcy levels are independently associated with increased rates of all-cause and CVD mortality in the elderly.

Fasting plasma homocysteine levels in the insulin resistance syndrome: the Framingham offspring study.

OBJECTIVE: Insulin resistance, associated metabolic abnormalities, and elevated homocysteine levels are risk factors for cardiovascular disease (CVD). We examined relationships between homocysteine levels and features of insulin resistance syndrome (IRS). RESEARCH DESIGN AND METHODS: We measured clinical characteristics, plasma levels of fasting homocysteine, folate, B vitamins, creatinine, and fasting and 2-h insulin and glucose levels after a 75-g oral glucose tolerance test in 2,214 subjects without CVD at the fifth examination (1991-1995) of the Framingham Offspring Study. After excluding 203 subjects with diabetes, the remaining 2,011 subjects were categorized as having none, one, two, or all three of the phenotypes of IRS: impaired glucose tolerance, hypertension, and/or a central metabolic syndrome (two or more traits: obesity, dyslipidemia, or hyperinsulinemia). In addition, in 1,592 subjects attending the sixth examination (1995-1998), we measured the urine albumin/creatinine ratio (UACR). Age-, sex-, creatinine-, vitamin-, and UACR-adjusted mean homocysteine levels or proportions with homocysteine >14 micromol/l in each phenotypic category and differences between categories were assessed with regression models. RESULTS: The mean age of the subjects was 54 years (range 28-82); 55% were women, 12.3% had hyperinsulinemia, and 15.9% had two or more of the IRS phenotypes. Adjusted mean homocysteine levels were higher comparing those with hyperinsulinemia (9.8 micromol/l) and those without (9.4 micromol/l, P = 0.04) and were higher among subjects with two or more IRS phenotypes (9.9 micromol/l) compared with those with 1 or no phenotype (9.3 micromol/l, P = 0.003). Mean UACR levels were also higher among subjects with two or more IRS phenotypes (7.2 mg/g) compared with those with 1 or no phenotype (5.5 mg/g, P = 0.007). CONCLUSIONS: Hyperhomocysteinemia and abnormal urinary albumin excretion are both associated with hyperinsulinemia and may partially account for increased risk of CVD associated with insulin resistance. Because hyperhomocysteinemia and microalbuminuria also reflect endothelial injury, these observations also support the hypothesis that endothelial dysfunction is associated with expression of the IRS.

Oxidative damage caused by free radicals produced during catecholamine autoxidation: protective effects of O-methylation and melatonin.

Catecholamine autoxidation produces reactive oxygen species that have been implicated in the loss of dopaminergic neurons in the nigrostriatal region of the brain that occurs during normal aging and in Parkinson's disease. In the present study, the potential protective effects of catecholamine O-methylation and of melatonin against catecholamine autoxidation-induced protein damage were assessed in vitro using the oxygen radical absorbance capacity (ORAC) assay. The rate of oxidation of the fluorescent protein porphyridium cruentum beta-phycoerythrin (beta-PE) caused by the oxidizing agent CuSO4 was shown to be accelerated by addition of the catecholamines dopamine and L-dopa. Replacement of dopamine and L-dopa in the assay with their O-methylated metabolites 3-O-methyldopamine and 3-O-methyldopa significantly decreased the rate of beta-PE oxidation. When melatonin was added to the ORAC assay in combination with dopamine or L-dopa, the rate of beta-PE oxidation was decreased as well. These findings were consistent with the following interpretations: (1) O-methylated catecholamines are less susceptible to autoxidation than their nonmethylated precursors, and (2) melatonin, which has recently been shown to be a powerful antioxidant, is capable of scavenging free radicals produced during catecholamine autoxidation. These findings suggest that O-methylation and melatonin may be important components of the brain's antioxidant defenses against catecholamine autoxidation and may protect against consequent dopaminergic neurodegeneration.

The pathogenesis of homocysteinemia: interruption of the coordinate regulation by S-adenosylmethionine of the remethylation and transsulfuration of homocysteine.

A unified, biochemical hypothesis is proposed to explain the pathogenesis of homocysteinemia. This hypothesis is based on the existence of coordinate regulation by S-adenosylmethionine (SAM) of the partitioning of homocysteine between de novo methionine synthesis and catabolism through cystathionine synthesis. This coordination, which serves to modulate the cellular concentration of homocysteine based on the requirements for methionine, is impaired in homocysteinemia. This hypothesis is evaluated in the context of the conditions known to be associated with homocysteinemia, including enzymatic defects and vitamin deficiencies. The novelty of the hypothesis is the assertion that impairment of one homocysteine metabolic pathway must lead to the impairment of the other homocysteine metabolic pathway to cause homocysteinemia. This extends the simplistic view that a block of only one of the pathways is sufficient to cause homocysteinemia.

Folate-binding protein and the absorption of folic acid in the small intestine of the suckling rat.

The folate in milk is largely bound to high-affinity folate-binding protein (FBP). With an in vivo intestinal loop technique, we examined the absorption of folic acid bound to FBP (FA-FBP) in the small intestine of the suckling rat. In contrast to unbound folic acid (FA), FA-FBP is absorbed more avidly in the ileum than in the jejunum (p less than 0.025) and its absorption is not inhibited by 1 mmol sulfasalazine/L. Folate-binding activities in the mucosa of the proximal (duodenum and jejunum combined) and distal (ileum) small intestine were also examined and found to be 0.32 and 1.31 pmol/mg protein, respectively (p less than 0.001). A 6-h fast produced a 42% decrease in folate-binding activity in the distal small intestine (p less than 0.01) but did not change activity in the proximal portion. Collectively, these observations suggest that FA-FBP is absorbed by a mechanism that is distinct from that responsible for the absorption of FA and that absorption does not require prior dissociation of the vitamin-binding protein complex.