Association Between Homocysteine and Vascular Calcification Incidence, Prevalence, and Progression in the MESA Cohort.

Background While elevated homocysteine has been associated with calcification in several studies, its importance as a cardiovascular risk factor remains unclear. This study examines the relationship between homocysteine and vascular and valve calcification in the MESA (Multi-ethnic Study of Atherosclerosis) cohort. Methods and Results MESA participants with baseline homocysteine measurements and cardiac computed tomography scans were included (N=6789). Baseline and follow-up assessment of vascular (coronary artery [CAC], descending thoracic aorta [DTAC]) and valve (aortic valve [AVC], mitral annular [MAC]) calcification was performed. Prevalence ratio/relative risk regression was used to assess the relationship of homocysteine with prevalent and incident calcification, and multivariable logistic regression was used to assess associations between homocysteine and calcification progression. Elevated homocysteine was associated with greater relative risk of prevalent and incident CAC and incident DTAC. We also identified a strong association between elevated homocysteine and CAC and DTAC progression. Elevated homocysteine was found to confer a >2-fold increased risk of severe CAC progression (defined as ΔCAC ≥100/year) and an ≈1.5-fold increased risk for severe DTAC progression (defined as ΔDTAC ≥100/year). Conclusions To our knowledge, this is the first study demonstrating an association between elevated homocysteine and both incidence and progression of coronary and extra-coronary vascular calcification. Our findings suggest a potential role for elevated homocysteine as a risk factor for severe vascular calcification progression. Future studies are warranted to further assess the utility of homocysteine as a biomarker for vascular calcification incidence and progression. Clinical Trial Registration https://www.clinicaltrials.gov/. Unique identifier: NCT00005487

Homocysteine treatment alters redox capacity of both endothelial and tumor cells

Homocysteine is a non-proteinogenic amino acid playing key roles in two interconnected metabolic pathways, namely, the activated methyl cycle and the linear trans-sulfuration pathway that allows the conversion of methionine to cysteine. A dysregulation of intracellular homocysteine metabolism could yield an increased export of this amino acid, leading to hyperhomocysteinemia, which has been associated with an increased risk of cardiovascular diseases. In spite of decades of experimental effort, there is no definitive consensus on what could be the molecular mechanisms whereby hyperhomocysteinemia could contribute to cardiovascular disease. The redox active nature of homocysteine has favored the idea of an induction of oxidative stress as the underlying mechanism of homocysteine toxicity. In contrast, homocysteine can also behave as an anti-oxidant. The present work is aimed to further analyze the capacity of homocysteine to modulate the redox capacity of both endothelial and tumor cells. [Our experimental work is supported by grants BIO2014-56092-R (MINECO and FEDER) and P12-CTS-1507 (Andalusian Government and FEDER) and funds from group BIO-267 (Andalusian Government). The "CIBER de Enfermedades Raras" is an initiative from the ISCIII (Spain)].Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Homocysteine levels and treatment effect in the prospective study of pravastatin in the elderly at risk

Objectives: To assess the effect of preventive pravastatin treatment on coronary heart disease (CHD) morbidity and mortality in older persons at risk for cardiovascular disease (CVD), stratified according to plasma levels of homocysteine.<p></p> Design: A post hoc subanalysis in the PROspective Study of Pravastatin in the Elderly at Risk (PROSPER), started in 1997, which is a double-blind, randomized, placebo-controlled trial with a mean follow-up of 3.2 years.<p></p> Setting: Primary care setting in two of the three PROSPER study sites (Netherlands and Scotland).<p></p> Participants: Individuals (n = 3,522, aged 70–82, 1,765 male) with a history of or risk factors for CVD were ranked in three groups depending on baseline homocysteine level, sex, and study site.<p></p> Intervention: Pravastatin (40 mg) versus placebo.<p></p> Measurements: Fatal and nonfatal CHD and mortality.<p></p> Results: In the placebo group, participants with a high homocysteine level (n = 588) had a 1.8 higher risk (95% confidence interval (CI) = 1.2–2.5, P = .001) of fatal and nonfatal CHD than those with a low homocysteine level (n = 597). The absolute risk reduction in fatal and nonfatal CHD with pravastatin treatment was 1.6% (95% CI = −1.6 to 4.7%) in the low homocysteine group and 6.7% (95% CI = 2.7–10.7%) in the high homocysteine group (difference 5.2%, 95% CI = 0.11–10.3, P = .046). Therefore, the number needed to treat (NNT) with pravastatin for 3.2 years for benefit related to fatal and nonfatal CHD events was 14.8 (95% CI = 9.3–36.6) for high homocysteine and 64.5 (95% CI = 21.4–∞) for low homocysteine.<p></p> Conclusion: In older persons at risk of CVD, those with high homocysteine are at highest risk for fatal and nonfatal CHD. With pravastatin treatment, this group has the highest absolute risk reduction and the lowest NNT to prevent fatal and nonfatal CHD.<p></p&gt

Creutzfeldt-Jakob disease and homocysteine levels in plasma and cerebrospinal fluid

Background: There is evidence that homocysteine contributes to various neurodegenerative disorders. Objective: To assess the values of homocysteine in patients with Creutzfeldt-Jakob disease (CJD) in both cerebrospinal fluid (CSF) and plasma. Methods: Study design: Case control study. Total homocysteine was quantified in CSF and plasma samples of CJD patients (n = 13) and healthy controls (n = 13). Results: Mean values in healthy controls: 0.15 mumol/l +/- 0.07 (CSF) and 9.10 mumol/l +/- 2.99 (plasma); mean values in CJD patients: 0.13 mumol/l +/- 0.03 (CSF) and 9.22 mumol/l +/- 1.81 (plasma). No significant differences between CJD patients and controls were observed (Mann-Whitney U, p > 0.05). Conclusions: The results indicate that the CSF and plasma of CJD patients showed no higher endogenous levels of homocysteine as compared to normal healthy controls. These findings provide no evidence for an additional role of homocysteine in the pathogenetic mechanisms underlying CJD neurodegeneration. Copyright (C) 2005 S. Karger AG, Basel

Homocysteine in cerebrovascular disease: An independent risk factor for subcortical vascular encephalopathy

Hyperhomocysteinemia is a risk factor for obstructive large-vessel disease. Here, we studied plasma concentrations of homocysteine and vitamins in patients suffering from subcortical vascular encephalopathy (SVE), a cerebral small-vessel disease leading to dementia. These results were compared to the homocysteine and vitamin plasma concentrations from patients with cerebral large vessel disease and healthy control subjects. Plasma concentrations of homocysteine, vascular risk factors and vitamin status (B-6, B-12, folate) were determined in 82 patients with subcortical vascular encephalopathy, in 144 patients with cerebral large-vessel disease and in 102 control subjects. Patients with SVE, but not those with cerebral large-vessel disease, exhibited pathologically increased homocysteine concentrations in comparison with control subjects without cerebrovascular disease. Patients with SVE also showed lower vitamin B6 values in comparison to subjects without cerebrovascular disease. Logistic regression analysis showed that homocysteine is associated with the highest risk for SVE (odds ratio 5.7; CI 2.5-12.9) in comparison to other vascular risk factors such as hypertension, age and smoking. These observations indicate that hyperhomocysteinemia is a strong independent risk factor for SVE

The concentration of homocysteine-derived disulfides in human coronary artery

*Background* 
Based on previous findings, we have estimated that, in injured coronary artery tissue, the low molecular weight disulfides homocystine and cysteine-homocysteine, otherwise identified as oxidized homocysteine equivalents (OHcyE), may achieve a total concentration that is higher than the aqueous solubility of homocystine at room temperature. In order to verify whether or not OHcyE could reach their saturation limit in the vascular tissue, we have measured the solubility of homocystine in physiological-like condition.

*Materials and methods* 
The solubility of homocystine has been measured in aqueous sodium chloride solutions at 37 °C by differential pulse polarography based on the reduction of homocystine to homocysteine.

*Results* 
We have estimated that the concentration achieved by OHcyE in injured coronary artery tissue is at least near-saturating, because the solubility of homocystine in physiological-like condition, above which deposition of homocystine and/or cysteine-homocysteine as solid phase occurs, almost exactly matches its value. Near-saturation levels of OHcyE within the vascular tissue means that significant leakage of intracellular fluid can promote OHcyE crystallization in tissue fluids, which may serve to initiate inflammation. 

*Conclusions* 
We speculate that deposition of OHcyE crystals could damage blood vessels and act as a primer of homocysteine-triggered inflammation, thus being along the causal pathway that leads to vascular dysfunction

Influence of hyperhomocysteinemia on the cellular redox state - Impact on homocysteine-induced endothelial dysfunction

Hyperhomocysteinemia is an independent risk factor for the development of atherosclerosis. An increasing body of evidence has implicated oxidative stress as being contributory to homocysteines deleterious effects on the vasculature. Elevated levels of homocysteine may lead to increased generation of superoxide by a biochemical mechanism involving nitric oxide synthase, and, to a lesser extent, by an increase in the chemical oxidation of homocysteine and other aminothiols in the circulation. The resultant increase in superoxide levels is further amplified by homocysteinedependent alterations in the function of cellular antioxidant enzymes such as cellular glutathione peroxidase or extracellular superoxide dismutase. One direct clinical consequence of elevated vascular superoxide levels is the inactivation of the vasorelaxant messenger nitric oxide, leading to endothelial dysfunction. Scavenging of superoxide anion by either superoxide dismutase or 4,5-dihydroxybenzene 1,3-disulfonate (Tiron) reverses endothelial dysfunction in hyperhomocysteinemic animal models and in isolated aortic rings incubated with homocysteine. Similarly, homocysteineinduced endothelial dysfunction is also reversed by increasing the concentration of the endogenous antioxidant glutathione or overexpressing cellular glutathione peroxidase in animal models of mild hyperhomocysteinemia. Taken together, these findings strongly suggest that the adverse vascular effects of homocysteine are at least partly mediated by oxidative inactivation of nitric oxide

Evidence Favoring a Positive Feedback Loop for Physiologic Auto Upregulation of hnRNP-E1 during Prolonged Folate Deficiency in Human Placental Cells

Background: Previously, we determined that heterogeneous nuclear ribonucleoprotein E1 (hnRNP-E1) functions as an intracellular physiologic sensor of folate deficiency. In this model, l-homocysteine, which accumulates intracellularly in proportion to the extent of folate deficiency, covalently binds to and thereby activates homocysteinylated hnRNP-E1 to interact with folate receptor-α mRNA; this high-affinity interaction triggers the translational upregulation of cell surface folate receptors, which enables cells to optimize folate uptake from the external milieu. However, integral to this model is the need for ongoing generation of hnRNP-E1 to replenish homocysteinylated hnRNP-E1 that is degraded.Objective: We searched for an interrelated physiologic mechanism that could also maintain the steady-state concentration of hnRNP-E1 during prolonged folate deficiency.Methods: A novel RNA-protein interaction was functionally characterized by using molecular and biochemical approaches in vitro and in vivo.Results: l-homocysteine triggered a dose-dependent high-affinity interaction between hnRNP-E1 and a 25-nucleotide cis element within the 5'-untranslated region of hnRNP-E1 mRNA; this led to a proportionate increase in these RNA-protein complexes, and translation of hnRNP-E1 both in vitro and within placental cells. Targeted perturbation of this RNA-protein interaction either by specific 25-nucleotide antisense oligonucleotides or mutation within this cis element or by small interfering RNA to hnRNP-E1 mRNA significantly reduced cellular biosynthesis of hnRNP-E1. Conversely, transfection of hnRNP-E1 mutant proteins that mimicked homocysteinylated hnRNP-E1 stimulated both cellular hnRNP-E1 and folate receptor biosynthesis. In addition, ferrous sulfate heptahydrate [iron(II)], which also binds hnRNP-E1, significantly perturbed this l-homocysteine-triggered RNA-protein interaction in a dose-dependent manner. Finally, folate deficiency induced dual upregulation of hnRNP-E1 and folate receptors in cultured human cells and tumor xenografts, and more selectively in various fetal tissues of folate-deficient dams.Conclusions: This novel positive feedback loop amplifies hnRNP-E1 during prolonged folate deficiency and thereby maximizes upregulation of folate receptors in order to restore folate homeostasis toward normalcy in placental cells. It will also functionally impact several other mRNAs of the nutrition-sensitive, folate-responsive posttranscriptional RNA operon that is orchestrated by homocysteinylated hnRNP-E1

Effect of Combination Folic Acid, Vitamin B6 , and Vitamin B12 Supplementation on Fracture Risk in Women: A Randomized, Controlled Trial.

Epidemiologic studies have demonstrated an association of elevated plasma homocysteine levels with greater bone resorption and fracture risk. Vitamins B12 , B6 , and folic acid are cofactors in homocysteine metabolism, and supplementation with B vitamins is effective in lowering homocysteine levels in humans. However, randomized trials of supplemental B vitamins for reduction of fracture risk have been limited. Therefore, we performed an ancillary study to the Women's Antioxidant and Folic Acid Cardiovascular Study (WAFACS), a large randomized trial of women with preexisting cardiovascular disease or three or more coronary risk factors, to test whether a daily B vitamin intervention including folic acid (2.5 mg/day), vitamin B6 (50 mg/day), and vitamin B12 (1 mg/day) reduces nonspine fracture risk over 7.3 years of treatment and follow-up. Among 4810 women, we confirmed 349 nonspine fracture cases by centralized review of medical records. In a substudy of 300 women (150 in treatment group and 150 controls) with paired plasma samples at randomization and follow-up (7.3 years later), we measured two bone turnover markers, including C-terminal cross-linking telopeptide of type I collagen (CTX) and intact type I procollagen N-propeptide (P1NP). In Cox proportional hazards models based on intention-to-treat, we found no significant effects of B vitamin supplementation on nonspine fracture risk (relative hazard = 1.08; 95% confidence interval, 0.88 to 1.34). In a nested case-cohort analysis, there were no significant effects of B vitamins on fracture risk among women with elevated plasma homocysteine levels, or low levels of vitamins B12 or B6 , or folate at baseline. Furthermore, treatment with B vitamins had no effect on change in markers of bone turnover. We found no evidence that daily supplementation with B vitamins reduces fracture risk or rates of bone metabolism in middle-aged and older women at high risk of cardiovascular disease. © 2017 American Society for Bone and Mineral Research

Transsulfuration pathway thiols and methylated arginines: the hunter community study

Background: Serum homocysteine, when studied singly, has been reported to be positively associated both with the endogenous nitric oxide synthase inhibitor asymmetric dimethylarginine [ADMA, via inhibition of dimethylarginine dimethylaminohydrolase (DDAH) activity] and with symmetric dimethylarginine (SDMA). We investigated combined associations between transsulfuration pathway thiols, including homocysteine, and serum ADMA and SDMA concentrations at population level. Methods: Data on clinical and demographic characteristics, medication exposure, C-reactive protein, serum ADMA and SDMA (LC-MS/MS), and thiols (homocysteine, cysteine, taurine, glutamylcysteine, total glutathione, and cysteinylglycine; capillary electrophoresis) were collected from a sample of the Hunter Community Study on human ageing [n = 498, median age (IQR) = 64 (60–70) years]. Results: Regression analysis showed that: a) age (P = 0.001), gender (P = 0.03), lower estimated glomerular filtration rate (eGFR, P = 0.08), body mass index (P = 0.008), treatment with beta-blockers (P = 0.03), homocysteine (P = 0.02), and glutamylcysteine (P = 0.003) were independently associated with higher ADMA concentrations; and b) age (P = 0.001), absence of diabetes (P = 0.001), lower body mass index (P = 0.01), lower eGFR (P&lt;0.001), cysteine (P = 0.007), and glutamylcysteine (P&lt;0.001) were independently associated with higher SDMA concentrations. No significant associations were observed between methylated arginines and either glutathione or taurine concentrations. Conclusions: After adjusting for clinical, demographic, biochemical, and pharmacological confounders the combined assessment of transsulfuration pathway thiols shows that glutamylcysteine has the strongest and positive independent associations with ADMA and SDMA. Whether this reflects a direct effect of glutamylcysteine on DDAH activity (for ADMA) and/or cationic amino acid transport requires further investigations.</br