ABSTRACT
The exact mechanism(s) by which hyperhomocysteinaemia promotes vascular disease remains unclear. Moreover, recent evidence suggests that the beneficial effect of folic acid on endothelial function is independent of homocysteine-lowering. In the present study the effect of a low (400 microg/70 kg/day) and high (5 mg/70 kg/day) dose folic acid supplement on endothelium-dependent relaxation in the isolated perfused mesenteric bed of heterozygous cystathionine beta-synthase deficient mice was investigated. Elevated total plasma homocysteine and impaired relaxation responses to methacholine were observed in heterozygous mice. In the presence of N(G)-nitro-L-arginine methyl ester relaxation responses in wild-type tissues were reduced, but in heterozygous tissues were abolished. Clotrimazole and 18alpha-glycyrrhetinic acid, both inhibitors of non-nitric oxide/non-prostanoid-induced endothelium-dependent relaxation, reduced responses to methacholine in wild-type but not heterozygous tissues. The combination of N(G)-nitro-L-arginine methyl ester and either clotrimazole or 18alpha-glycyrrhetinic acid completely inhibited relaxation responses in wild-type tissues. Both low and high dose folic acid increased plasma folate, reduced total plasma homocysteine and reversed endothelial dysfunction in heterozygous mice. A greater increase in plasma folate in the high dose group was accompanied by a more significant effect on endothelial function. In the presence of N(G)-nitro-L-arginine methyl ester, a significant residual relaxation response was evident in tissues from low and high dose folic acid treated heterozygous mice. These data suggest that the impaired mesenteric relaxation in heterozygous mice is largely due to loss of the non-nitric oxide/non-prostanoid component. While low dose folic acid may restore this response in a homocysteine-dependent manner, the higher dose has an additional effect on nitric oxide-mediated relaxation that would appear to be independent of homocysteine lowering.
Subject(s)
Endothelium, Vascular/drug effects , Folic Acid/pharmacology , Hyperhomocysteinemia/physiopathology , Mesenteric Arteries/drug effects , Vasodilation/drug effects , Animals , Clotrimazole/pharmacology , Cystathionine beta-Synthase/deficiency , Cystathionine beta-Synthase/genetics , Disease Models, Animal , Dose-Response Relationship, Drug , Endothelium, Vascular/physiopathology , Enzyme Inhibitors/pharmacology , Folic Acid/blood , Homocysteine/blood , Hyperhomocysteinemia/blood , Mesenteric Arteries/physiopathology , Methacholine Chloride/pharmacology , Mice , Mice, Inbred C57BL , Mice, Transgenic , NG-Nitroarginine Methyl Ester/pharmacology , Nitric Oxide Synthase/antagonists & inhibitors , Nitroprusside/pharmacology , Vasodilator Agents/pharmacologyABSTRACT
While folic acid has been shown to reverse endothelial dysfunction, the exact underlying mechanism remains elusive. Here, folic acid reversed both the endothelial dysfunction and increased production of superoxide following depletion of rabbit aortic ring tetrahydrobiopterin (BH4) levels with 2,4-diamino-6-hydroxy-pyrimidine (DAHP) and N-acetyl-5-hydroxy-tryptamine (NAS). Incubation with l-nitroarginine methyl ester also attenuated the production of superoxide. DAHP and NAS reduced BH4 concentrations in both aorta and cultured porcine aortic endothelial cells. Folic acid had no effect on BH4 concentrations in either preparation. The superoxide anion scavenger Tiron but not folic acid reversed the endothelial dysfunction produced in aortic rings by inhibition of copper-zinc superoxide dismutase with diethyldithiocarbamic acid. Neither folic acid nor its metabolite 5-methyltetrahydrofolate prevented the in vitro oxidation of BH4. This study demonstrates that folic acid reverses the endothelial dysfunction induced by BH4 depletion independently of either the regeneration or stabilization of BH4 or an antioxidant effect.
Subject(s)
Biopterins/analogs & derivatives , Endothelial Cells/drug effects , Endothelium, Vascular/drug effects , Folic Acid/pharmacology , Animals , Aorta, Thoracic/drug effects , Aorta, Thoracic/physiology , Biopterins/antagonists & inhibitors , Biopterins/metabolism , Cells, Cultured , Ditiocarb/pharmacology , Endothelial Cells/metabolism , Endothelium, Vascular/metabolism , Endothelium, Vascular/physiopathology , In Vitro Techniques , Male , Rabbits , Superoxide Dismutase/antagonists & inhibitors , Superoxides/metabolism , Swine , Vasodilation/drug effectsABSTRACT
Evidence reported from numerous clinical studies over the past decade has revealed an association between increased plasma total homocysteine (tHcy) concentrations and cardiovascular disease (CVD). In addition, epidemiological studies have identified an inverse association between blood folate concentrations, folate intake and cardiovascular endpoints, that are independent of homocysteine. Folic acid supplementation can lower plasma tHcy concentrations safely and inexpensively. Furthermore, folic acid can reverse endothelial dysfunction observed in patients with CVD. This reversal in endothelial dysfunction with folic acid has been shown to be independent of plasma tHcy lowering, suggesting that folate has pleiotropic effects on the vasculature other than homocysteine lowering. In vitro evidence demonstrates that 5-methyltetrahydrofolate (5MeTHF) the main circulating metabolite of folate, can increase nitric oxide production and can directly scavenge superoxide radicals. The potential beneficial role of folic acid supplements on vascular disease are currently being tested in randomized placebo controlled studies.
