Dietary treatment of hyperhomocysteinemia in peripheral arterial disease

Abstracts

A homocisteína está envolvida na gênese da aterosclerose e, assim, é considerada um importante e prevalente fator de risco na doença arterial periférica. O estado nutricional vitamínico deficiente, em especial do folato, é a principal causa de hiper-homocisteinemia nesses casos. Embora ainda não haja consenso sobre a dose exata e a forma de utilização do folato em suplementos e sobre adequação alimentar ou fortificação de cereais para o tratamento da hiper-homocisteinemia, diversos estudos realizados em pacientes com doença vascular periférica mostraram que o folato, isoladamente, pode reduzir as concentrações de homocisteína, bem como a concentração de alguns marcadores biológicos do processo de aterosclerose. No entanto, estudos recentes não comprovaram esse benefício sobre o processo inflamatório associado à hiper-homocisteinemia. Desta forma, embora a utilização isolada do folato seja uma terapêutica custo-efetiva no controle da hiper-homocisteinemia, seu impacto na evolução das doenças arteriais ainda persiste inconclusivo. Esta revisão abordará os efeitos obtidos com as diversas formas de utilização do folato no tratamento da hiper-homocisteinemia.

Homocisteína; aterosclerose; vitamina; ácido fólico

REVIEW ARTICLE

^INutricionista, Doutora em Bases Gerais da Cirurgia, Faculdade de Medicina de Botucatu (FMB), Universidade Estadual Paulista (UNESP), Botucatu, SP, Coordenadora, Curso de Graduação em Nutrição, Universidade Metodista de Piracicaba, Campus Lins, SP

^IIProfessor titular, Departamento de Saúde Pública, e coordenador, Centro de Metabolismo em Exercício e Nutrição, FMB, UNESP, Botucatu, SP

^IIIProfessor adjunto, Departamento de Cirurgia e Ortopedia, FMB, UNESP, Botucatu, SP

Correspondence

ABSTRACT

Homocysteine plays a role in the genesis of atherosclerosis and, thus, it is considered an important and prevalent risk factor for peripheral arterial disease. Impaired vitamin nutritional status, especially regarding folate, may be mainly attributed to hyperhomocysteinemia. Although there is still no consensus as to the exact dose and method of use of folate in supplements, dietary adjustment or cereal fortification for the treatment of hyperhomocysteinemia, several studies conducted in patients with peripheral vascular disease have shown that isolated folate may reduce homocysteine levels, as well as the levels of some biological markers in the atherosclerotic process. However, recent studies have not corroborated this benefit for the inflammatory process associated with hyperhomocysteinemia. Consequently, although the use of folate is a cost-effective therapy for the control of hyperhomocysteinemia, its impact on the evolution of vascular diseases remains inconclusive. This literature review addresses the effects of several forms of folate therapies in the treatment of hyperhomocysteinemia.

Keywords: Homocysteine, atherosclerosis, vitamin, folic acid.

Introduction

Peripheral artery disease (PAD) is defined as an artery disease of the extremities which reduces the blood flow during exercise or, in advanced stages, even during rest.¹ Epidemiological studies show that PAD's prevalence varies between 1.6 and 12%.^2,3

Circulatory system diseases were the main causes of death among Brazilians in 2007 (28.2%)⁴ and in world population (18 million);⁵ among the various etiologies, atherosclerosis was the most common cause. Among patients with symptomatic atherosclerotic disease, 15.9% have symptomatic polyvascular disease (PAD, cardiovascular and cerebrovascular disease).⁶ In addition, in the intermittent claudication phase, around 65% of individuals present important and simultaneous impairment of coronary and cerebrovascular sectors, and 30% of the patients with coronary or cerebral vascular disease present PAD.⁷ Thus, PAD is associated with an increase in morbidity and mortality due to cardiovascular diseases (approximately 30%) and cerebrovascular diseases,⁸ being, thus, an important predictor for both.

Some risk factors for the development of atherosclerotic disease are very well-known, such as age, sex (male), dyslipidemia, smoking, systemic hypertension, diabetes mellitus, obesity, sedentarianism and genetic factors or family history of atherosclerotic disease.⁶ Last years, other risk factors were identified, such as hyperhomocysteinemia, whose study might broaden the understanding on physiopathological mechanisms of atherosclerosis and enable the development of new preventive or therapeutic measures.

Homocysteine and hyperhomocysteinemia metabolism in PAD

Homocysteine is a non-protein sulfur amino acid, it is not a constituent of the diet and is not formed of proteins, but it is exclusively an intermediary product of intracellular metabolism of the essential amino acid methionine.^9,10 (Figure 1)

Homocysteine's intracellular metabolism occurs by means of two remethylation pathways, responsible for the conversion of homocysteine in methionine, and a transsulfuration pathway, which converts homocysteine in cysteine (Figure 2). These pathways depend upon vitamins B6, B12 and folate, which act as coenzymes or co-substrates. In one of the remethylation pathways, the donor of the methyl group is the 5-methylenetetrahydrofolate (5-MTHF), main form of the folate in plasma, produced from the reduction of 5,10-methylenetetrahydrofolate by the enzyme methylenetetrahydrofolate reductase (MTHFR). Transference of the methyl group to homocysteine is made by means of the enzyme methionine-synthase (MS), which has as co-factor the methylcobalamin (MCOB), a coenzyme derived from cobalamin (vitamin B12). Subsequently, a new molecule of 5-MTHF is synthesized by the transference of a carbon atom (from a source of carbon as serine) for a molecule of tetrahydrofolate, producing methylenetetrahydrofolate and glycine.¹³

In the other pathway of remethylation (demethylation), which exists mainly in the liver, the donor of the methyl group for homocysteine conversion into methionine is betaine. Once formed, methionine is activated by adenosine triphosphate (ATP), forming S-adenosylmethionine, which works as donor of methyl group to a diversity of receptors. The product of these reactions of methylation is S-adenosylmethionine, which is hydrolyzed regenerating homocysteine, which becomes available to initiate a new cycle of transference of methyl groups.¹⁴

In the transsulfuration pathway, initially homocysteine is condensed with a serine molecule to form cystathionine in an irreversible reaction which is catalyzed by the enzyme cystathionine-β-synthase (CBS) and has as co-factor pyridoxal-5’-phosphate, a derivate of pyridoxine or vitamin B6. Afterward, cystathionine is hydrolyzed in cysteine and α-ketobutyrate acid; this reaction also requires vitamin B6 and is catalyzed by the enzyme γ-cystathionase. Therefore, in addition to the production of cysteine, the transsulfuration pathway is responsible for the catabolism of toxic homocysteine, not necessary in the methyl groups transference cycle. Thus, this metabolization pathway aims at homocysteine's renal excretion, decreasing the seric concentration of this amino acid. In normal metabolic conditions, homocysteine metabolization is distributed equally between these two pathways, which are coordinated and compete for the use of available homocysteine.¹⁴

The normal homocysteine concentration in plasma when analyzed at fasting is approximately 10 µmol/L due to the cellular exportation mechanism, varying between 5 and 15 µmol/L; above these values hyperhomocysteine is characterized.¹⁵ Kang et al.¹⁶ classify arbitrarily hyperhomocysteine in severe form, concentrations over 100 µmol/L; moderate, concentrations between 31 and 100 µmol/L; and mild, concentrations between 15 and 30 µmol/L.

Since the discovery of homocysteine, various and important prospective and case-control studies have shown that hyperhomocysteinemia is a risk factor for vascular diseases and is prevalent in patients with PAD (28 to 60%^17,18, compared with 1% in general population).⁷ Mean concentrations of homocysteine in patients with various manifestation of PAD, such as intermittent claudication,¹⁹ ileo-femoral lesions,²⁰ Leriche syndrome,²¹ carotid stenosis,²² and abdominal aorta aneurysm,²³ showed to be significantly higher than those found in controls. Approximately 30% of young patients carrying PAD present hyperhomocysteinemia. National studies in patients with coronary artery disease^24-26 and peripheral artery disease^18,27,28 have confirmed the high prevalence of hyperhomocysteinemia (20 and 60%) and the high concentration of homocysteine in patients in relation to controls. A meta-analysis study with a sample of approximately 4,000 persons concluded that an increase of 5 µmol/L in homocysteine plasma concentrations is associated with the development of cardiovascular disease in risk ratio (odds ratio) of 1.6 for men and 1.8 for women, compared to 6.8 for patients with PAD.¹⁷ According to current existing evidence, hyperhomocysteine presents risk ratio of 3.0 for patients with symptomatic PAD.⁷ In addition, prospective studies have evidenced that hyperhomocysteinemia is associated to the increase in early death risk due to cardiovascular disease²⁹ and PAD progression^30,31 and non-lethal coronary arterial disease in patients with symptomatic PAD.³²

The most common causes of hyperhomocysteine in general population are related to genetic defects in codification of enzymes or nutritional deficiency of vitamins involved in the homocysteine metabolism. Plasma concentration of homocysteine is inversely associated to folate blood concentrations, vitamin B6 and B12 and ingestion of these vitamins, mainly in old people.³³

Genetic hyperhomocysteine is frequently the result of molecular alterations associated with MTHFR heterozygosis or CBS. It is estimated that 5 to 10% of severe hyperhomocysteine cases are caused by defects in remethylation pathways, homozygotic deficiency of MTHFR (677CT) more common and less severe when compared with that caused by CBS deficiency, but with a worse prognosis, because it does not respond well to available treatments. Individuals who are monozygotic mutants for MTHFR enzyme present only 30% of the activity observed in individuals with normal genotype, with the value of up to 65% for heterozygotic genotype.³⁵ Homozygosis for MTHFR enzyme is present in 5 to 20% of the general population and was observed in 16.7% of PAD patients, who had a moderately increased homocysteine.³⁶ In Brazil, it was observed that 19% of 191 individuals with coronary artery disease were homozygotic for the MTHFR enzyme's thermolabile variant, but no correlation with homocysteine was studied.³⁷ Moderate elevations in homocysteine concentrations are not found in all the patients with this polymorphism, which suggests that the phenotype might be influenced by other factors, such as vitamin nutritional state, hormons, some diseases and drugs.³⁸ There are great influences of folate seric concentrations in the MTHFR enzyme's polymorphism, and hyperhomocysteine in homozygotic individuals is observed only when there is folate deficiency (values inferior to 15.4 µmol/L).³⁹

Hyperhomocysteine's pathogenic mechanisms

Despite the great amount of epidemiological data establishing a correlation between hyperhomocysteine and increase in vascular diseases risk, the mechanisms through which hyperhomocysteine contributes to estrogenesis and thrombogenesis are hypothetical and controversial. The aggression to the endothelium seems to be one of the mechanisms through which homocysteine leads to vascular lesion. Groundbreaking works^40,41 with non-human primates (baboons) have shown that L-homocysteine intravenous injection during 5 days provoked endothelial lesion characterized by endothelial desquamation , proliferation of smooth muscle cells and vascular intimal layer, mediated by reduction of plaquetary half-life, with a rapid formation of vascular lesions, similar to early atherosclerotic lesions in humans. The degree of endothelial lesion provoked by hyperhomocysteine was similar to the one observed in association with other risk factors, as in hypercholesterolemia and systemic hypertension.

National and current experimental researches reinforced the role of hyperhomocysteine induced by methionine overload in the formation of aortic atherosclerotic plaque in swines⁴² and iliac artery in rabbits.⁴³ After a period of 30 or 60 days of methionine-rich diet, there was a significant increase in homocysteine concentrations and in the formation of atherosclerotic plaques by foamy cells, but no smooth muscle cells were observed, neither cholesterol crystals nor inflammatory cells.

One of the main mechanisms of endothelial dysfunction induced by hyperhomocysteine would be related with the diminution of endothelial relaxing factor's bioavailability, nitric oxide, synthesized from L-arginine by the action of the enzyme nitric oxide-synthase. Nitric oxide is a strong endogenous vasodilator which inhibits plaquetary aggregation, leukocytes migration and the proliferation and migration of the smooth muscle cell and restricts the activation and expression of adhesion molecules and the production of superoxide anions.⁴⁴ Previous studies in hyperhomocysteinemic animals have shown that homocysteine reduced the nitric oxide's bioavailability in a culture of endothelial cells, probably by induction of oxidative stress, which deactivates nitric oxide or by inhibition of nitric oxide-synthase, or even by increasing dimethylarginine concentrations, which contributes to the reduction of nitric oxide's bioavailability.⁴⁵

It is also speculated that hyperhomocysteine played an important role in endothelial lesion through oxidative and inflammatory mechanisms. The sulfhidryl group (SH) present in homocysteine is highly reactive due to being an electrons donor in the oxidation systems, rapidly oxidized in disulphide (SS). Homocysteine is readily oxidized when it reaches plasma, mainly in consequence of its autoxidation to form SS, including homocysteine, homocysteine-derived SS and thiolactone homocysteine, the latter being the most reactive form of homocysteine. The SH group of SS and thiolactone homocysteine would react with oxygen, producing hydrogen peroxide and superoxide, which would initiate lipid peroxidation both in endothelial surface and in low density lipoproteins (LDL).⁴⁶ LDL oxidation, favored mainly by thiolactone homocysteine, would form aggregates to be captured by macrophages in the arteries' intimal layer, which would stimulate a vascular pro-inflammatory response by means of the expression of adhesion molecules, chemotactic proteins and growth factors to form foamy cells and, consequently, atheromatous lesions. Furthermore, autoxidation of homocysteine in the plasma could favor reduction in the expression and activity of glutathione peroxidase and, therefore, inhibit endothelial cells' antioxidant potential.⁴⁷ In cultures of endothelial cells, smooth muscle cells and human monocytes, hyperhomocysteinemia induced the expression of the monocyte chemoattractant protein (1-MCP-1) and that of interleukin-8 (IL-8) by activating nuclear factor kappa B (NF-kB) and inducing positive acute phase C-reactive protein (CRP). These substances would broaden vascular inflammatory response and contribute to the initiation and progress of atherosclerosis.⁴⁸

Other homocysteine effects would be alterations in antithrombotic properties of vascular endothelium. In vitro studies in cells exposed to homocysteine have demonstrated an increase in activity of coagulant factors XII and V, reduction of C-protein activation, inhibition of tissue plasminogen activator, reduction of nitric oxide and prostacyclin bioavailability, stimulation of plaquetary aggregation, increase in von Willebrand factor activity, inhibition of thombomodulin expression, induction of tissue factor expression and suppression of heparan-sulphate expression in the vascular wall.⁴⁹ All these alterations would generate a vascular thrombogenic setting, with the activation of coagulation cascade and vascular tonus modification.

The variability of homocysteine actions pointed by these studies shows that there is still not a single explanatory hypothesis for homocysteine's atherothrombogenic effects.

Hyperhomocysteine nutritional treatment

Nutritional factors, particularly consumption and seric concentration of folate, vitamin B12 and B6, seem to be the major parameters in homocysteine metabolization. Deficiencies, isolated or combined, of vitamins involved in the various pathways of homocysteine metabolism would be important markers for hyperhomocysteine.⁵⁰ In North-American elders, two thirds (67%) of hyperhomocysteine cases were attributed to improper dietary habits in relation to one or more complex B vitamins.⁵¹ In individuals carrying PAD in our milieu, the frequency of hyperhomocysteine with insufficient ingestion of 2 or 3 complex B vitamins (52.5%) was significantly higher in relation to those with insufficient ingestion of 1 vitamin (7.5%), which confirms the relation between hyperhomocysteinemia and vitamin nutritional state.¹⁸ According to a 12-year prospective follow-up study with 46,036 male participants,⁵² consumption of food folate and supplements containing folic acid would be inversely associated with the occurrence of PAD and could contribute for its prevention.

Dietary deficiency of folate would provoke insufficient formation of 5-MTHF, which is necessary as a methyl radical donor group in homocysteine remethylation into methionine. Vitamin B12 deficiency, on its turn, would lead to impairment in the transference of the 5-MTHF's methyl radical for homocysteine in remethylation pathway through methionine-synthase. Therefore, vitamin B6 deficiency would hinder the conversion of homcysteine into cysteine by CBS enzymes and γ-cystathionase, which are activated by vitamin B6 in the transsulfuration pathway. These conditions would favor the accumulation of intracellular homocysteine, which would be transported for the extracellular compartment and, consequently, provoke increase in circulating concentrations of homocysteine. This process would restrain intracellular toxicity, but, on the other hand, could expose vascular setting to hazarding effects of the excess of homocysteine.⁵³ Among these vitamins, folate would be the major dietary determinant of homocysteine concentration, because its deficiency could restrain the homocysteine' remethylation pathway into methionine and favor its accumulation in the extracellular environment.⁵⁰

Folate is a generic term used to name this essential and hydrosoluble vitamin of the complex B, which presents itself in the active form of tetrahydrofolic acid and functions as coenzyme for transference reactions of necessary carbon units in several metabolic pathways, including purines and pirimidines metabolism, as well as amino acids interconversions. Folic acid (pteroyl-L-glutamic acid), an oxidized and stable form of folate, despite rarely found in foods, is the form used in vitamin supplements and fortified food products, representing 20% of folate in the diet. In natural foods, so called food folate is found in the form of pteroyl-L-glutamate, which represents 80% of folate in the diet (Table 1).

Ideal consumption of vitamins B12, B6 and folate is defined by the Recommended Dietary Allowances (RDA), advocated by the Dietary Reference Intakes,⁵⁵ defined as sufficient level of food consumption to satisfy the needs of almost every healthy individual (between 97 and 98%), comprehended in a certain group, age group and life stage (Table 2).

In 1996, Food and Drug Administration (FDA) regulated the fortification of cereals as flour, rice, pastas and corn with 140 µg of folic acid by 100 g of the product, aiming at reducing the risk of neural tube defects in newborns. After this regulation, a significant increase was observed in the seric folates concentrations and significant reduction in homocysteine concentrations in 350 adults and elders participating in the Framingham Offspring Study.⁵⁶ Fortification with 499 and 665 µg of folic acid for 30 g of matinal cereals was equally efficient in reducing homocysteine and increasing seric folate in patients with coronary artery disease. However, fortification with 127 µg, a quantity close to that proposed by the FDA, was not sufficient to significantly reduce homocysteine and induce a moderate increase in seric folate. Therefore, homocysteine reduced and seric folate increased linearly with the increase in folic acid contained in the cereal. In face of these results, the authors suggested that a fortification level over (approximately 350 µg) the one proposed by the FDA should be secured.⁵⁷ Brazil does not have recent nationally representative information on micronutrients deficiency in adults. Even so, there was a governmental decision on universal fortification of wheat and corn flours produced in the country with folic acid,⁵⁸ based on locally-limited studies performed by different institutions in several geographic regions. The resolution made mandatory the fortification of wheat and corn flour cereals with 150 µg of folic acid for 100 g of cereal, representing 37% of RDA for adults, aiming at preventing neural tube defects in newborns and folate deficiency in children and adults.⁵⁹

Considering that hyperhomocysteinemia is common in individuals presenting nutritional impairment of vitamins B6, B12 and especially folate, and given the significant participation these vitamins have in homocysteinemia, various studies successfully used these vitamins to reduce blood concentrations of this amino acid. Folate, isolated or in combination with vitamins B6 and B12, may reduce plasma homocysteine, including when there are no deficiencies. There is still no consensus on the exact dose of these vitamins for the treatment of hyperhomcysteinemia, and different doses were used in many studies. The contribution of vitamin therapy, especially that of folic acid, in hyperhomocysteinemia is widely studied in healthy individuals and in coronary arterial disease, but still rare in PAD (Table 3).

A meta-analysis of 11 studies carried out by Boushey et al.¹⁷ in 1995 have shown groundbreaking results of the effect of the treatment with folic acid on homocysteinemia in coronary, cerebral and peripheral vascular diseases. Nine intervention studies have shown reduction of homocysteine and normalization of seric folate after isolated supplementation of folic acid (more than 400 µg/day) and concluded that the consumption of approximately 200 µg of dietetic folate (3 or more portions of fruits and vegetables a day) would reduce in 4µmol/L the concentrations of homocysteine in cardiovascular, cerebrovascular and peripheral artery disease. However, the authors have highlighted that not all the vascular diseases associated with hyperhomocysteinemia would be prevented with folic acid supplementation and that the fortification or increase in foods possessing folate could have a greater preventive effect.

In another meta-analysis with 12 randomized studies and 1,114 individuals,⁶⁰ the reducing effects of folic acid on homocysteine in vascular disease were also studied. Daily use of 50 to 500 µg of folic acid between 3 to 12 weeks have reduced in 25% the concentration of homocysteine, and the reduction was higher in hyperhomocysteinemic individuals (> 12 µmol/L) and folate seric deficiency (< 12 µmol/L) before treatment with folic acid. The authors have reported yet that the supplementation with vitamin B12 potentialized the reduction of homocysteine in 7% and that vitamin B6 had no additional effect on homocysteine reduction.

Studies performed in our milieu have confirmed these findings. A case-control study has investigated the supplementation of folic acid (500 µg) associated with vitamin B12 in 40 patients with symptomatic PAD and showed efficiency in the normalization of homocysteinemia in 4 weeks of treatment (before, 18.807.73 µmol/L; after, 11.34 µmol/L; p < 0.001), contrarily to supplementation with vitamin B6 (250 mg), which did not alter hyperhomocysteinemia.⁶⁴ Equally, another cross-study with adult hypertensive individuals practicing supervised physical exercise has shown significant reduction of mean homocysteine concentrations after 2 months of treatment with folic acid (500 µg) when diuretic use was absent.⁶⁵

A comparative study on the effects of three forms of supplementation with folic acid on homocysteinemia showed that: 1) daily consumption of fortified matinal cereals and 2) the use of supplements seemed to be the most effective methods for increasing the levels of folate and reducing blood homocysteine; 3) increase in folate-rich foods would not be exactly an effective strategy to reduce homocysteinemia, probably due to the variations in bioavailability of folate in fruits and vegetables and to the infrequent ingestion of these foods.⁶² Other authors have suggested that it was little likely that the diet alone was sufficient to increase circulating folate concentrations and reduce homocysteinemia, once food folate (pteroyl-L-glutamate) would present half the bioavailability of folic acid originated from supplements (pteroyl-L-glutamate).⁶⁶ Most folates present in foods, even if very stable to light, are labile and thermosensitive. These characteristics have led to considerable losses of folate in processing foods at high temperatures. So, 50 to 90% of the content of this vitamin might be destroyed by coction or other processes as wrapping and refinement.⁵⁵

The consumption of fortified foods or supplements with vitamin B12 is also recommended in cases of hypercysteinemia in elders, because 10-30% of these patients may present malabsorption of this vitamin due to the reduction of intrinsic factor, and the supplements guarantee an adequate absorption by passive diffusion, therefore without the presence of intrinsic factor.⁶

Despite the existence of studies demonstrating that folate-rich foods were not sufficient to reduce plasma homocysteine, other studies have shown that, after ingestion of approximately 600 µg of food folate present in vegetable foods (greens, legumes and fruits), there was a reduction of homocysteinemia, besides additional benefits of reduction in saturated fats consumption and increase in fibers, iron, thiamine, vitamin C and B6 consumption. It is worth reporting that, contrarily to supplements and fortified foods, quantification of food folate consumption is imprecise due to the variation of composition tables and, therefore, it needs various observation periods.^61,63

Other therapeutic effects of using vitamins, especially folate, were investigated in hyperhomocysteinemia, mainly in some biological markers of the atherosclerosis process possibly associated to hyperhomocysteinemia, such as endothelial dysfunction, coagulability, muscle cells proliferation, activity of reactive species of the oxygen metabolism, pro-inflammatory response^67,68 and oxidation of the LDL fraction.⁴⁶ In this sense, some studies carried out in the cardiology area^69-71 in healthy individuals,^72-74 adults and elders, hyperhomocysteinemic or not, have shown a reduction of endothelial dysfunction due to a significant increase in vasodilating response, dependent or independent on the brachial artery endothelium with high doses of folic acid (500 to 1,000 µg) in 6 to 12 weeks of treatment. In these studies, assessment of vasodilating response dependent on the endothelium was performed by comparison in response to a period of ischemia (tourniquet), and the vasodilating response independent on the endothelium, which is, by direct action on smooth muscle cells, was evaluated after administration of vasoactive agents as sublingual nitroglycerin. Using these techniques, it was possible to analyze the diameter and the changes in blood flow in brachial artery through Doppler ultrasound and plethysmography. The mechanism through which folic acid improves vasodilating response is not known, but it is speculated that with the reduction of circulating homocysteine there is a greater bioavailability of nitric oxide due to the stimulus to endothelial nitric oxide-synthase, which favors vasodilation.⁷³

The use of folic acid for 2 months in experimental model of hyperhomocysteinemic rats submitted to carotid endarterectomy was effective in reducing homocysteine concentration and intimal hyperplasia, assessed through luminal stenosis percentage. The findings of this study support the theory that hyperhomocysteinemia increases the mitogenic stimulus of smooth muscle cells, producing a broad intimal hyperplasia in response to the carotid endarterectomy model. Based on these results, the authors suggested the analysis of plasma homocysteine at endarterectomy post-operative: if hyperhomocysteinemia is verified, patients could initiate therapy with folic acid and avoid post-endarterectomy restenosis.⁷⁵ In other experimental study with rats, hyperhomocysteinemia induction increased the percentage of endothelial lesion in cerebral vases observed through electronic microscopy; according to the authors, the lesion was provoked by alterations in intracellular oxidative metabolism. Endothelial lesions found in hyperhomocysteinemic rats were similar to those typically found in degenerative cerebral diseases as Alzheimer and Parkinson. On the other hand, the inclusion of moderate doses of folic acid in the diet for 8 weeks resulted in morphological changes and, consequently, reduced significantly these endothelial lesions.⁷⁶ In 250 patients submitted to coronary angioplasty, combined therapy of folic acid (1,000 µg), vitamin B12 (400 µg) and B6 (10 mg), used for 6 months, could be used as adjuvant therapy in coronary angioplasty, because it promoted a significant reduction of homocysteinemia and of the post-angioplasty restenosis rate.⁷⁷

The influence of acid folic therapy on systemic inflammatory markers of atherosclerosis, such as PCR and LDL-oxidized, is still controversial. Results observed in a recent study with healthy men and women who had received 800 µg of folic acid (n = 264) or placebo (n = 266) daily by 12 months indicated that there was an increase in 400% in seric folate concentration, significant reduction in homocysteine concentrations (28%) and maintenance of ultrasensitive PCR (PCR-us) and LDL-oxidized.⁷⁸ In a double-blind controlled study similar to the previous one, with 65 hyperhomocysteinemic patients carrying symptomatic PAD, receiving placebo or complex B (B1, B6, and B12) vitamins or folic acid (500 µg) for 6 months, it was observed that the treatment for homocysteine reduction did not alter homeostatic markers (tissue factor, TF, and tissue factor pathway inhibitor, TFPI), of chronic inflammation (such as PCR-us), interleukins (6, 8 and 18) and monocyte chemotactic protein (MCP-1).⁷⁹ The authors concluded that the treatment for homocysteine reduction in symptomatic PAD patients is questionable in relation to the control of chronic inflammatory process inherent to atherosclerosis and examined on the effect of hyperhomocysteinemia in chronic inflammation and coagulation of symptomatic PAD patients. The use of antioxidant vitamins as alpha-tocopherol, ascorbic acid, beta-carotene, associated to folic acid and vitamin B12 for 15 days, was more effective in significantly reducing in vitro LDL oxidation in patients with coronary artery disease.⁸⁰

Having in mind the whole of these works, to fight mild forms of hyperhomocysteinemia, dietary adequacy or use of fortified foods with folic acid would be the more advisable choice; in cases of moderate hyperhomocysteinemia, drug supplementation with folic acid would be more recommended, due to being more effective in reducing homocysteinemia.

Effects of hyperhomcysteinemia treatment on morbidity and mortality due to vascular diseases

Currently, various prospective studies were preformed to evaluate the effects of supplementation with vitamins involved in homocysteine metabolism on the prevalence and/or progression of atherosclerotic lesions in various vascular sectors.

In a groundbreaking study, 232 adult patients carrying symptomatic PAD, 30% with hyperhomocysteinemia (17.812.7 µmol/L), were treated with 500 µg of folic acid associated with vitamin B6 (250 mg) for 2 years. There was a significant reduction in homocysteine concentration (17.812.7 to 8.22.2 µmol/L) and that of frequency of PAD progression, but not of the incidence of new cerebral or coronary events in hyperhomocysteinemic patients after vitamin treatment in comparison to not treated normo-homocysteinemic patients. In this study, progression diagnoses were: 1) symptomatology along with reduction of ankle-arm pressure index and/or by angiography for PAD; 2) presence of transitory ischemic attack or stroke for cerebral disease; 3) presence of acute myocardial infarct or progression of angina pectoris to coronary artery disease.⁸¹ In another study, performed with patients with peripheral, coronary or cerebral vascular disease, supplementation with folic acid (500 µg) associated with vitamin B6 (250 mg), when compared with placebo, showed only a reduction of abnormal rate of electrocardiographic test in exercise, but no effect on ankle-arm pressure index or regression of atherosclerotic lesions in carotid and femoral artery.⁸²

In accordance with the previous study, supplementation with 500 µg of folic acid for 2 years did not reduce the recurrence of cardiovascular events in 300 patients.⁸³

Aiming at observing the recurrence of stroke, new coronary events and death, a controlled randomized double-blind multicenter study called VISP (The Vitamin Intervention for Stroke Prevention) was performed, evaluating 3,680 adults and elders with previous diagnosis of stroke. Subjects who presented plasma homocysteine below 10 µmol/L were distributed in two distinct groups, one receiving high doses of folic acid, vitamin B6 and B12 (2,500 µg, 25 mg and 400 µg, respectively) and other receiving low doses (20 µg, 0.2 mg, and 6 µg, respectively). After a 2-year follow-up, supplementation with low vitamins doses was more effective in reducing homocysteine, the risk of stroke (10%, p = 0.05), of coronary events (26% p < 0.001) and death (16%, p = 0.001). A probable explanation for the ineffectiveness of the high doses treatment could be associated with homocysteinemia in pre-treatment, once response to vitamin treatment is knowingly more effective in hyperhomocysteinemic individuals.⁸⁴

Cereals fortification with folic acid and the use of supplements reduced the relative risk of coronary disease in 8 to 23% in women over 55 years old and in 11 to 13% in men over 45 years old;^85,86 according to the authors, this may be a beneficial and not onerous strategy only in cases of hyperhomocysteinemia.

In face of conflicting results of these prospective studies, various recent reviews have concluded that well-designed studies should be urgently performed to assess the impact of hyperhomocysteinemia treatment on morbidity and mortality^87,88 and progression of vascular diseases, including PAD.⁸⁹ The authors of this review emphasized that homocysteine is easy to be assessed and, more importantly, easy and cheap to be treated, although some clinical studies question the beneficial effects of this therapeutics.⁹⁰ On the other hand, the decline in cardiovascular mortality in the United States could be attributed, partially, to the fortification with folate since 1998, presumably due to the increase of blood folate and reduction of blood concentrations of homocysteine.⁹¹

Conclusion

Information currently available evidence that combined supplementation of folate, vitamin B12 and vitamin B6 might be used in treating hyperhomocysteinemia. Indeed, supplementation or fortification, or diet adequacy isolated from folate, promotes an effective reduction of plasma homocysteine, configuring a cost-effective treatment which presents efficacy, with no adverse reactions, characterizing, therefore, a therapeutic coadjuvant in relation to risk-benefit, very favorable in controlling homocysteinemia. In relation to the impact of supplementation on morbidity and mortality due to vascular diseases, the studies have shown to be divergent and inconclusive.

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Dietary treatment of hyperhomocysteinemia in peripheral arterial disease

Luciene de Souza Venâncio^I; Roberto Carlos Burini^II; Winston Bonetti Yoshida^III

Publication Dates

• Publication in this collection
  01 Sept 2010
• Date of issue
  2010

History

• Received
  26 Apr 2009
• Accepted
  03 Feb 2010