Severe Hyperhomocysteinemia Research Articles

Molecular and biochemical investigations of patients with intermediate or severe hyperhomocysteinemia.

A discrepancy has been identified between numbers of expected and identified patients with homocystinuria due to cystathionine beta-synthase (CBS) deficiency. Patients homozygous for the frequent c.833T>C (p.I278T) are most often responsive to vitamin B6, and can present with a total-homocysteine (tHcy) <100 μM on a normal diet. In Denmark, patients with tHcy <100 μM are not routinely sequenced for CBS(2) mutations. This study investigated the prevalence of CBS mutations and the common methylenetetrahydrofolate reductase (MTHFR) c.677C>T polymorphism in patients with tHcy ≥ 50 μM and the association with clinical manifestations. We studied a cohort of patients with intermediate and severe hyperhomocysteinemia (tHcy ≥ 50 μM) determined between 1996 and 2011. Among the 413 eligible patients, 184 (45%) patients agreed to participate in the present follow-up study. A MTHFR(3)c.677TT genotype was found in 49% of the patients. Eight patients were found to have mutations in CBS(2). Of those, two were homozygous for c.833T>C (p.I278T), and four were compound heterozygous for c.833T>C. One c.833T>C (p.I278T) compound heterozygote was identified by lowering the threshold for sequencing from tHcy at 100 μM to 50 μM. The most prominent clinical presentation among patients with a CBS(2) mutation was thrombosis presenting at a median age of 25 years. In case of arterial or venous thrombosis without any explanation in individuals below 40 years, tHcy should be part of the thrombophilia screening. When tHcy is between 50 and 100 μM genotyping for the MTHFR(3) c.677TT is relevant, and when tHcy >100 μM CBS should be genotyped.

Severe Hyperhomocysteinemia Decreases Respiratory Enzyme and Na(+)-K(+) ATPase Activities, and Leads to Mitochondrial Alterations in Rat Amygdala.

Severe hyperhomocysteinemia is caused by increased plasma levels of homocysteine (Hcy), a methionine derivative, and is associated with cerebral disorders. Creatine supplementation has emerged as an adjuvant to protect against neurodegenerative diseases, due to its potential antioxidant role. Here, we examined the effects of severe hyperhomocysteinemia on brain metabolism, and evaluated a possible neuroprotective role of creatine in hyperhomocysteinemia, by concomitant treatment with Hcy and creatine (50mg/Kg body weight). Hyperhomocysteinemia was induced in young rats (6-day-old) by treatment with homocysteine (0.3-0.6µmol/g body weight) for 23days, and then the following parameters of rat amygdala were evaluated: (1) the activity of the respiratory chain complexes succinate dehydrogenase, complex II and cytochrome c oxidase; (2) mitochondrial mass and membrane potential; (3) the levels of necrosis and apoptosis; and (4) the activity and immunocontent of Na(+),K(+)-ATPase. Hcy treatment decreased the activities of succinate dehydrogenase and cytochrome c oxidase, but did not alter complex II activity. Hcy treatment also increased the number of cells with high mitochondrial mass, high mitochondrial membrane potential, and in late apoptosis. Importantly, creatine administration prevented some of the key effects of Hcy administration on the amygdala. We also observed a decrease in the activity and immunocontent of the α1 subunit of the Na(+),K(+)-ATPase in amygdala after Hcy- treatment. Our findings support the notion that Hcy modulates mitochondrial function and bioenergetics in the brain, as well as Na(+),K(+)-ATPase activity, and suggest that creatine might represent an effective adjuvant to protect against the effects of high Hcy plasma levels.

Homocysteine, hyperhomocysteinemia and vascular contributions to cognitive impairment and dementia (VCID)

Homocysteine is produced physiologically in all cells, and is present in plasma of healthy individuals (plasma [HCy]: 3–10μM). While rare genetic mutations (CBS, MTHFR) cause severe hyperhomocysteinemia ([HCy]: 100–200μM), mild-moderate hyperhomocysteinemia ([HCy]: 10–100μM) is common in older people, and is an independent risk factor for stroke and cognitive impairment. As B-vitamin supplementation (B6, B12 and folate) has well-validated homocysteine-lowering efficacy, this may be a readily-modifiable risk factor in vascular contributions to cognitive impairment and dementia (VCID).Here we review the biochemical and cellular actions of HCy related to VCID. Neuronal actions of HCy were at concentrations above the clinically-relevant range. Effects of HCy <100μM were primarily vascular, including myocyte proliferation, vessel wall fibrosis, impaired nitric oxide signalling, superoxide generation and pro-coagulant actions. HCy-lowering clinical trials relevant to VCID are discussed. Extensive clinical and preclinical data support HCy as a mediator for VCID. In our view further trials of combined B-vitamin supplementation are called for, incorporating lessons from previous trials and from recent experimental work. To maximise likelihood of treatment effect, a future trial should: supply a high-dose, combination supplement (B6, B12 and folate); target the at-risk age range; and target cohorts with low baseline B-vitamin status. This article is part of a Special Issue entitled: Vascular Contributions to Cognitive Impairment and Dementia edited by M. Paul Murphy, Roderick A. Corriveau and Donna M. Wilcock.

Sulfur amino acids and atherosclerosis: a role for excess dietary methionine.

The homocysteine theory of arteriosclerosis received credence when it was shown that after a methionine load, circulating homocysteine-cysteine concentrations were higher in cardiovascular disease patients than in healthy controls. Subsequent studies showing associations between homocysteine and coronary artery disease, stroke and cognitive impairment, relied on small increases in homocysteine concentration unlike the very high homocysteine seen in the rare genetic disorders that lead to homocystinuria and much higher homocysteine levels. Subsequent studies in cell culture, animals, and humans showed that a variety of cardiovascular adverse effects of "high homocysteine" introduced either as a nonphysiological bolus or as a methionine load led to high homocysteine. We fed apolipoprotein E-deficient mice diets designed to achieve three conditions: (1) high methionine intake with normal blood homocysteine, (2) high methionine intake with B vitamin deficiency and hyperhomocysteinemia, and (3) normal methionine intake with both B vitamin deficiency and hyperhomocysteinemia. We found that the mice fed methionine-rich diets had significant atheromatous pathology in the aortic arch even with normal plasma homocysteine levels. Mice fed B vitamin-deficient diets developed severe hyperhomocysteinemia but without any increase in vascular pathology. Our findings suggest that even moderate increases in methionine intake are atherogenic in susceptible mice while high plasma homocysteine is not.

Organotypic tissue culture investigation of homocysteine thiolactone cardiotoxic effect.

Homocysteine thiolactone was demonstrated to inhibit the growth of 10-12-day-old chicken embryo cardiac tissue explants at 7 × 10⁻⁹ -1 × 10⁻³ M concentrations in a dose-dependent manner. The maximal cardiotoxic effect of homocysteine thiolactone was detected at 1 × 10⁻³ M, which corresponds to severe hyperhomocysteinemia. The results of experiments on culturing of cardiac tissue explants in the medium containing homocysteine thiolactone (1 × 10⁻³ M) and ouabain at concentrations regulating the signal-transducing (1 × 10⁻¹⁰ M) and pumping (1 × 10⁻⁸ M) functions of Na⁺,K⁺ -ATPase indicate that the cardiotoxic effect of homocysteine thiolactone is supposed to result from inhibition of the Na⁺,K⁺ -ATPase pumping function.

Insights into severe 5,10-methylenetetrahydrofolate reductase deficiency: molecular genetic and enzymatic characterization of 76 patients.

5,10-Methylenetetrahydrofolate reductase (MTHFR) deficiency is the most common inherited disorder of folate metabolism and causes severe hyperhomocysteinaemia. To better understand the relationship between mutation and function, we performed molecular genetic analysis of 76 MTHFR deficient patients, followed by extensive enzymatic characterization of fibroblasts from 72 of these. A deleterious mutation was detected on each of the 152 patient alleles, with one allele harboring two mutations. Sixty five different mutations (42 novel) were detected, including a common splicing mutation (c.1542G>A) found in 21 alleles. Using an enzyme assay in the physiological direction, we found residual activity (1.7%-42% of control) in 42 cell lines, of which 28 showed reduced affinity for nicotinamide adenine dinucleotide phosphate (NADPH), one reduced affinity for methylenetetrahydrofolate, five flavin adenine dinucleotide-responsiveness, and 24 abnormal kinetics of S-adenosylmethionine inhibition. Missense mutations causing virtually absent activity were found exclusively in the N-terminal catalytic domain, whereas missense mutations in the C-terminal regulatory domain caused decreased NADPH binding and disturbed inhibition by S-adenosylmethionine. Characterization of patients in this way provides a basis for improved diagnosis using expanded enzymatic criteria, increases understanding of the molecular basis of MTHFR dysfunction, and points to the possible role of cofactor or substrate in the treatment of patients with specific mutations.

Modeling of severe hyperhomocysteinemia in rats

To date it’s proved that the elevated homocysteine levels in the blood is an independent risk factor for a number of diseases. The animal model of rare severe form of hyperhomocysteinemia may be necessary to study aspects of homocysteine participation in damage of organs and tissues in various diseases because the effects of this pathogenetic factor will be most pronounced in such conditions. This article describes a technique for modeling in male Wistar rats a severe form of hyperhomocysteinemia by a three-week intragastric administration of methionine suspension with the addition of this amino acid in the drinking water.

Severe hyperhomocysteinemia due to cystathionine β-synthase deficiency, and Factor V Leiden mutation in a patient with recurrent venous thrombosis.

Homocysteine is an amino acid that is toxic to vascular endothelial cells, and plasma elevations have been associated with venous thromboembolism. Severe hyperhomocysteinemia (>100 μmol/L) may result from mutations in the genes coding for enzymes in the trans-sulfuration or the folate/vitamin B12-dependent re-methylation pathways. Here, we report the case of a young woman with severe, recurrent thrombo-embolic events associated with severe hyperhomocysteinemia (111 μmol/L). We identified a homozygous mutation in the cystathionine β -synthase gene (p.I278T) and the presence of the Factor V Leiden mutation. Family study shows segregation of elevated homocysteine in heterozygous relatives for the mutation in the cystathionine β -synthase gene. Management consisted of anticoagulation with warfarin and supplementation with folate, vitamin B6 (pyridoxine) and vitamin B12. After twelve years of follow-up, plasma homocysteine levels remain in the moderate range (~20 μmol/L, reference range 8-12 μmol/L) and no further thromboembolic events were identified.

Homocysteine: Often Neglected but Common Culprit of Coronary Heart Diseases

Hyperhomocysteimia is an independent risk factors for ischemic heart disease and stroke. We report in absent risk factors the clinical correlation between severe acute coronary syndrome and hyperhomocysteinemia. We report in absent risk factors the clinical correlation severe acute coronary syndrome and hyperhomocysteinemia A 38 years old non-smoker, non-diabetes, normotensive and strict vegetarian with positive family history of ischemic heart diseases had acute myocardial infarction with raised serum homocysteine level. Key words: Acute myocardial infarction, family history of ischemic heart disease, homocysteinee

Creatine prevents the imbalance of redox homeostasis caused by homocysteine in skeletal muscle of rats

Homocystinuria is a neurometabolic disease caused by severe deficiency of cystathionine beta-synthase activity, resulting in severe hyperhomocysteinemia. Affected patients present several symptoms including a variable degree of motor dysfunction, being that the pathomechanism is not fully understood. In the present study we investigated the effect of chronic hyperhomocysteinemia on some parameters of oxidative stress, namely 2'7'dichlorofluorescein (DCFH) oxidation, levels of thiobarbituric acid-reactive substances (TBARS), antioxidant enzyme activities (SOD, CAT and GPx), reduced glutathione (GSH), total sulfhydryl and carbonyl content, as well as nitrite levels in soleus skeletal muscle of young rats subjected to model of severe hyperhomocysteinemia. We also evaluated the effect of creatine on biochemical alterations elicited by hyperhomocysteinemia. Wistar rats received daily subcutaneous injection of homocysteine (0.3-0.6 μmol/g body weight), and/or creatine (50mg/kg body weight) from their 6th to the 28th days age. Controls and treated rats were decapitated at 12h after the last injection. Chronic homocysteine administration increased 2'7'dichlorofluorescein (DCFH) oxidation, an index of production of reactive species and TBARS levels, an index of lipoperoxidation. Antioxidant enzyme activities, such as SOD and CAT were also increased, but GPx activity was not altered. The content of GSH, sulfhydril and carbonyl were decreased, as well as levels of nitrite. Creatine concurrent administration prevented some homocysteine effects probably by its antioxidant properties. Our data suggest that the oxidative insult elicited by chronic hyperhomocystenemia may provide insights into the mechanisms by which homocysteine exerts its effects on skeletal muscle function. Creatine prevents some alterations caused by homocysteine.

Blood thiol status and erythrocyte glutathione-S-transferase in chronic kidney disease patients on treatment with frequent (daily) hemodialysis

Background. Chronic kidney disease (CKD) is a condition of impaired homeostasis of blood thiols characterized by a severe hyperhomocysteinemia (HH) and abnormal expression of the red blood cell glutathione (GSH)-consuming enzyme GSH S-transferase (eGST) (Galli et al., Clin Chem 1999). The correlation between plasma Hcy and eGST recently identified by our group (Dessì et al., Amino Acids 2012) suggests a role of this detoxifying enzyme in the impaired thiol status of CKD treated with hemodialysis therapy (HD). This retrospective study is aimed at investigating whether frequent HD can alleviate these biochemical symptoms of CKD. Methods. Laboratory data of a population of 98 HD patients investigated for plasma Hcy and blood thiol status between 1999 and 2004 were examined. A frequent HD method carried out with a 2-h daily schedule (daily HD) (DHD) was compared with standard 4-h × 3/week protocol of HD (SHD) in either cross-sectional (n = 70 SHD vs. n = 28 DHD) and prospective A-B design (n = 18 SHD patients shifted to DHD). Results. The results demonstrate that DHD produces a better correction than SHD of the uremic retention solute Hcy as well as of Cys and Cys-Gly measured in plasma. Such a correction effect of DHD on HH correlates with that on the detoxification enzyme eGST and on pGSH. Conclusions. These findings point to a role of frequent dialysis in the depuration of uremic retention solutes that may interfere with thiol metabolism and redox in HD patients. These solutes may include substrates of eGST that await further investigation for molecular identification and better removal by more efficient dialysis therapies.

Nutriepigenetic regulation by folate–homocysteine–methionine axis: a review

Although normally folic acid is given during pregnancy, presumably to prevent neural tube defects, the mechanisms of this protection are unknown. More importantly it is unclear whether folic acid has other function during development. It is known that folic acid re-methylates homocysteine (Hcy) to methionine by methylene tetrahydrofolate reductase-dependent pathways. Folic acid also generates high-energy phosphates, behaves as an antioxidant and improves nitric oxide (NO) production by endothelial NO synthase. Interestingly, during epigenetic modification, methylation of DNA/RNA generate homocysteine unequivocally. The enhanced overexpression of methyl transferase lead to increased yield of Hcy. The accumulation of Hcy causes vascular dysfunction, reduces perfusion in the muscles thereby causing musculopathy. Another interesting fact is that children with severe hyperhomocysteinaemia (HHcy) have skeletal deformities, and do not live past teenage. HHcy is also associated with the progeria syndrome. Epilepsy is primarily caused by inhibition of gamma-amino-butyric-acid (GABA) receptor, an inhibitory neurotransmitter in the neuronal synapse. Folate deficiency leads to HHcy which then competes with GABA for binding on the GABA receptors. With so many genetic and clinical manifestations associated with folate deficiency, we propose that folate deficiency induces epigenetic alterations in the genes and thereby results in disease.

Evaluation of the prevalence of severe hyperhomocysteinemia in adult patients with thrombosis who underwent screening for thrombophilia

Treatment with B-vitamins and betaine reduces the high risk of thrombosis in patients with homocystinuria, a metabolic syndrome that is characterized by severe hyperhomocysteinemia (HHcy). In contrast, there is no clear demonstration that B-vitamins reduce the risk of thrombosis in patients with mild HHcy: for this reason, many question the clinical utility of measuring total Hcy (tHcy) in patients with thrombosis. However, thrombosis may be the first clinical manifestation of homocystinuria in patients reaching adulthood without signs and symptoms of the syndrome. 1) to measure the prevalence of severe, previously undiagnosed, HHcy among patients with thrombosis 2) to profile these patients on the basis of their characteristics. Six Italian Thrombosis Centers completed a first questionnaire, reporting tHcy levels in patients with thrombosis who underwent thrombophilia screening, and a second questionnaire, reporting the characteristics of patients with severe HHcy (tHcy>100μmol/L). Of 19,678 cross-sectionally collected patients with thrombosis who underwent thrombophilia screening in the last 12.5years (median value, range 6-17), 38 had severe HHcy (0.2%). Their median age at diagnosis was 47years (range 19-83) and the median level of tHcy was 130μmol/L (range 101-262). Venous thromboembolism (71%) was more frequent than arterial thromboembolism (26%); recurrent thrombosis occurred in 42% of cases. Measurement of tHcy in adult patients with thrombosis may reveal the presence of severe HHcy. Since treatment of patients with severe HHcy decreases the risk of thrombosis, measurement of tHcy in patients with thrombosis may prove clinically useful.

A case of homocystinuria due to CBS gene mutations revealed by cerebral venous thrombosis

BackgroundHomocystinuria caused by cystathionine beta synthase (CBS) deficiency is most often diagnosed in childhood and has a variable expressivity. The most frequent abnormalities include intellectual disability, ectopia lentis, myopia, skeletal abnormalities or thromboembolism. ObjectiveTo report a case of homocystinuria unraveled by cerebral venous thrombosis (CVT). ObservationA 17 year old female was admitted in our department of neurology for subacute headache and presented seizures in the emergency room. Cerebral imaging revealed CVT. Severe hyperhomocysteinemia was found and led to the diagnosis of homocystinuria due to composite heterozygous mutations in the CBS gene. Further investigations disclosed lens subluxation in association with myopia, mild scoliosis and osteopenia. The patient was treated by heparin followed by warfarin, vitamin therapy and dietary methionine restriction. Total homocysteine and methionine levels became normal in a few weeks and the patient had a complete recovery. ConclusionIn patients with CVT, plasma total homocysteine measurement as part of the etiologic work up may reveal severe hyperhomocysteinemia due to CBS or remethylation defects that require specific treatment and management including perhaps protein-restricted diet and/or vitamin therapy for life.

Comparative antiadhesive properties of crude extract and phenolic fraction isolated from aerial parts of Tribulus pterocarpus during severe hyperhomocysteinemia

The phenolic fraction and the crude extract from Tribulus pterocarpus have different biological activity, including antiplatelet-antiadhesive properties. Since it is demonstrated that hyperhomocysteinemia may act as stimulator of blood platelet activation (platelet adhesion, aggregation, and secretion), but various antiplatelet compounds are able to reduce hyperactivation of blood platelets induced by hyperhomocysteinemia. The aim of our present experiments was to investigate in vitro one of the step in platelet activation process - platelet adhesion to collagen induced by the model of severe hyperhomocyateinemia in the presence of the phenolic fraction and the crude extract from T. pterocarpus. Severe hyperhomocysteinemia was induced by reduced form of Hcy in the concentrations 0.1mM and 1mM, or using HTL in the concentrations 0.1, 0.5 and 1 μM. Adhesion of blood platelets to collagen was determined according to Tuszynski and Murphy. We observed that the phenolic fraction and the crude extract from T. pterocarpus have the inhibitory effect on platelet adhesion during severe hyperhomocysteinemia. The action of tested phenolic and crude extract was concentration-dependent, but the phenolic fraction was stronger antiadhesive action than the crude extract. We suggest that T. pterocarpus may be good source of antiplatelet compounds during hyperhomocysteinemia.

A case of venous thrombolism with severe hyperhomocysteinemia

A case of venous thrombolism with severe hyperhomocysteinemia

Pulmonary thromboembolism due to severe hyperhomocysteinemia associated with a methyltetrahydrofolate reductase mutation

Pulmonary thromboembolism due to severe hyperhomocysteinemia associated with a methyltetrahydrofolate reductase mutation

Homocysteine induces energy imbalance in rat skeletal muscle: Is creatine a protector?

Homocystinuria is a neurometabolic disease caused by a severe deficiency of cystathionine beta-synthase activity, resulting in severe hyperhomocysteinemia. Affected patients present several symptoms including a variable degree of motor dysfunction. In this study, we investigated the effect of chronic hyperhomocysteinemia on the cell viability of the mitochondrion, as well as on some parameters of energy metabolism, such as glucose oxidation and activities of pyruvate kinase, citrate synthase, isocitrate dehydrogenase, malate dehydrogenase, respiratory chain complexes and creatine kinase in gastrocnemius rat skeletal muscle. We also evaluated the effect of creatine on biochemical alterations elicited by hyperhomocysteinemia. Wistar rats received daily subcutaneous injections of homocysteine (0.3-0.6 µmol/g body weight) and/or creatine (50 mg/kg body weight) from the 6th to the 28th days of age. The animals were decapitated 12 h after the last injection. Homocysteine decreased the cell viability of the mitochondrion and the activities of pyruvate kinase and creatine kinase. Succinate dehydrogenase was increased other evaluated parameters were not changed by this amino acid. Creatine, when combined with homocysteine, prevented or caused a synergistic effect on some changes provoked by this amino acid. Creatine per se or creatine plus homocysteine altered glucose oxidation. These findings provide insights into the mechanisms by which homocysteine exerts its effects on skeletal muscle function, more studies are needed to elucidate them. Although creatine prevents some alterations caused by homocysteine, it should be used with caution, mainly in healthy individuals because it could change the homeostasis of normal physiological functions.

Homocysteine and Cardiovascular Risk: The Perils of Reductionism in a Complex System

Patients with genetic defects causing severe hyperhomocysteinemia have a high rate of mortality from cardiovascular diseases at an early age, irrespective of the specific genetic lesion. Long-term therapy with B vitamins or betaine in patients with homocystinuria due to cystathionine β-synthase deficiency decreases homocysteine and reduces the rate of vascular events by 90%. Kilmer McCully studied the vascular pathology in young patients who died with homocystinuria and in 1969 proposed that a moderate increase in circulating homocysteine could be involved in the pathogenesis of vascular occlusive disease in the general population, i.e., the so-called homocysteine theory of atherosclerosis. This theory was investigated in the early 1990s in many clinical and epidemiologic studies. A metaanalysis of studies of homocysteine concentration and vascular risk concluded that an increase of 0.68 mg/L (5 μmol/L) in plasma homocysteine had an impact on cardiovascular risk that was equivalent to a 19-mg/dL (0.5-mmol/L) increase in total cholesterol. In general, the risk estimates from the prospective studies are weaker than those obtained from case–control and cross-sectional studies (1, 2), yet they clearly support plasma homocysteine as an independent cardiovascular risk factor. The possible role of homocysteine in the development of vascular disease inspired research on the biochemical and molecular effects on the vasculature of homocysteine and the determinants of circulating homocysteine concentrations. Experimental studies revealed numerous homocysteine effects relevant to the pathogenesis of vascular disease, including induction of endothelial dysfunction, inhibition of biological methylation through accumulation of S -adenosylhomocysteine, homocysteinylation and posttranslational modification of proteins, increased oxidant stress, and decreased availability of nitric oxide (1). In addition, plasma homocysteine positively correlates with a variety of biochemical parameters, clinical traits, and lifestyle factors associated with increased cardiovascular risk, including dyslipidemia, increased blood pressure, impaired renal function, a sedentary …

Does phase angle correlate with hyperhomocysteinemia? A study of patients with classical homocystinuria

The role of the phase angle in hyperhomocysteinemia has yet to be assessed. Classical homocystinuria is a rare genetic disease characterized by severe hyperhomocysteinemia, as well as increased levels of methionine and reduced levels of cysteine. The objective of this study was to investigate the potential relationship between phase angle and homocysteine, cysteine, and methionine levels in patients with classical homocystinuria. Eight patients were included in the study. Phase angle was measured with a tetrapolar bioimpedance analyzer. Serum homocysteine, cysteine, and methionine levels were measured by HPLC. Only three patients had adequate metabolic control of their disease. Median phase angle was 5.9° (range=5.4°-8.5°). There was a significant correlation between phase angle and levels of homocysteine (r=-0.807, p=0.015), methionine (r=-0.711, p=0.048), and cysteine (r=0.836, p=0.010). Was also positively correlated with BMI and arm muscle circumference (p<0.05). Two patients had phase angles below the 5th percentile, and only one above the 50th percentile. Our findings suggest that cellular integrity is affected in patients with high homocysteine levels, thus indicating that phase angle could be a valuable indicator of prognosis and classical homocystinuria. It also suggests a role for this indicator in other forms of hyperhomocysteinemia and other inborn errors of metabolism.