RESUMO
A novel microminipig has been recently developed for use in biomedical research. In the present study, age- and sex-related differences, as well as 24-h fluctuations in plasma total homocysteine concentrations (tHcy), were investigated in these microminipigs. tHcy (mean±SD) was 10.2±3.4 µM and significantly correlated with age. By contrast, neither the differences in tHcy between sexes nor the 24-h fluctuations in tHcy after feeding were significant. The kinetics of plasma tHcy after intravenous injection of reduced Hcy showed that its levels peaked within 5 min post-injection, as did the levels of tHcy. These results suggested that reduced Hcy is rapidly oxidized or metabolized. The half-lives of reduced Hcy, tHcy, and reduced cysteine in the blood were 47, 71, and 141 min, respectively. In conclusion, there was a significantly positive correlation between age and plasma tHcy in microminipigs. After intravenous injection of reduced Hcy, plasma tHcy quickly returned to pre-injection levels.
Assuntos
Homocisteína/sangue , Porco Miniatura/sangue , Animais , Cisteína/sangue , Feminino , Homocisteína/metabolismo , Homocisteína/farmacocinética , Masculino , Metionina/sangue , Suínos , Porco Miniatura/metabolismoRESUMO
In the presented work a variation of total aminothiols (cysteine, glutathione, cysteinylglycine and homocysteine) in blood plasma have been shown at modelling hyperhomocysteinemia by daily intraperitoneal (0.6 mkmol/g body weight) and subcutaneous (0.12 mkmol/g body weight) introduction of homocysteine. During two weeks of the intraperitoneal introduction a significant concentration growth (from -40 to 180 mkM) of cysteine was observed. We also observed a moderate change of concentration levels for glutathione (from 10-15 to 30 mkM) and cysteinylglycine (from 1,5 to 4,5 mkM). The homocysteine level has decreased from 300 to 200-250 mkM at second week of experiments. Experimental results with subcutaneous introduction were similar. In this case a stable homocysteine level (-70 mkM) and increase of cysteine level (to 60 mkM) was observed at second week. These data reflect dose-depended processes of organism adaptation to hyperhomocysteinemia, i.e. reinforced capability for homocysteine metabolism and at the same time retention low glutathione level which correlates with hyperhomocysteinemia degree and duration.
Assuntos
Cisteína/sangue , Dipeptídeos/sangue , Glutationa/sangue , Homocisteína/farmacocinética , Animais , Homocisteína/administração & dosagem , Homocisteína/sangue , Injeções Intraperitoneais , Injeções Subcutâneas , Masculino , Ratos , Ratos WistarRESUMO
Interest in radiolabeled amino acids for metabolic imaging of cancer and limitations with [(11)C]methionine has prompted the development of a new (18)F-labeled methionine derivative S-(3-[(18)F]fluoropropyl)homocysteine ([(18)F]FPHCys). The L and D enantiomers of [(18)F]FPHCys were prepared from their respective protected S-(3-tosyloxypropyl)homocysteine precursors 1 by [(18)F]fluoride substitution using K(2.2.2) and potassium oxalate, followed by acid hydrolysis on a Tracerlab FX(FN) synthesis module. [(18)F]-L-FPHCys and [(18)F]-D-FPHCys were isolated in 20 ± 5% radiochemical yield and >98% radiochemical and enantiomeric purity in 65 min. Competitive uptake studies in A375 and HT29 tumor cells suggest that L- and D-[(18)F]FPHCys are taken up by the L-transporter system. [(18)F]-L-FPHCys and [(18)F]-D-FPHCys displayed good stability In Vivo without incorporation into protein at least 2 h postinjection. Biodistribution studies demonstrate good uptake in A375 tumor-bearing rodents with tumor to blood ratios of 3.5 and 5.0 for [(18)F]-L-FPHCys and [(18)F]-D-FPHCys, respectively, at 2 h postinjection.
Assuntos
Homocisteína/análogos & derivados , Neoplasias Experimentais/diagnóstico por imagem , Compostos Radiofarmacêuticos/síntese química , Sistema A de Transporte de Aminoácidos/antagonistas & inibidores , Sistema A de Transporte de Aminoácidos/metabolismo , Sistema L de Transporte de Aminoácidos/antagonistas & inibidores , Sistema L de Transporte de Aminoácidos/metabolismo , Animais , Linhagem Celular Tumoral , Radioisótopos de Flúor , Homocisteína/síntese química , Homocisteína/química , Homocisteína/farmacocinética , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Neoplasias Experimentais/metabolismo , Tomografia por Emissão de Pósitrons , Compostos Radiofarmacêuticos/química , Compostos Radiofarmacêuticos/farmacocinética , Estereoisomerismo , Relação Estrutura-Atividade , Distribuição TecidualRESUMO
The distribution and metabolism of selenohomolanthionine (4,4'-selenobis[2-aminobutanoic acid], SeHLan), a newly identified selenoamino acid in selenized Japanese pungent radish, were evaluated by administering 77Se-labeled SeHLan at a dose of 25 µg/kg body weight in rats. Exogenous 77Se of SeHLan was preferably distributed to the kidneys and remained in the intact form for up to 6 h after dosing. The accumulation in the kidneys is one of the specific characteristics of SeHLan, differing from other selenoamino acids, such as selenomethionine and Se-methylselenocysteine, which preferably accumulate in the pancreas. The intact form of SeHLan was detected in the serum and kidney supernatant but not in the urine, suggesting that the amount of exogenous Se that was distributed to the kidneys was within metabolic capacity. Indeed, the exogenous Se was converted into two urinary metabolites, Se-methylseleno-N-acetyl-galactosamine and trimethylselenonium. Exogenous Se was also detected in several selenoproteins, including selenoprotein P and extracellular glutathione peroxidase. SeHLan is expected to be a potential supplemental source of Se because its distribution differs from that of selenomethionine and Se-methylselenocysteine.
Assuntos
Homocisteína/análogos & derivados , Compostos Organosselênicos/metabolismo , Compostos Organosselênicos/farmacocinética , Selênio/metabolismo , Selênio/farmacocinética , Animais , Homocisteína/análise , Homocisteína/metabolismo , Homocisteína/farmacocinética , Isótopos , Masculino , Compostos Organosselênicos/análise , Ratos , Ratos Wistar , Selênio/análise , Distribuição TecidualRESUMO
Elevated maternal plasma concentrations of homocysteine (Hcy) are associated with pregnancy complications and adverse neonatal outcomes. The postulate that we wish to advance here is that placental transport of Hcy, by competing with endogenous amino acids for transporter activity, may account for some of the damaging impacts of Hcy on placental metabolism and function as well as fetal development. In this article, we provide an overview of some recent studies characterising the transport mechanisms for Hcy across the microvillous plasma membrane (MVM) of the syncytiotrophoblast, the transporting epithelium of human placenta. Three Hcy transport systems have been identified, systems L, A and y(+)L. This was accomplished using a strategy of competitive inhibition to investigate the effects of Hcy on the uptake of well-characterised radiolabelled substrates for each transport system into isolated MVM vesicles. The reverse experiments were also performed, examining the effects of model substrates on [³5S]L-Hcy uptake. This article describes the evidence for systems L, A and y(+)L involvement in placental Hcy transport and discusses the physiological implications of these findings with respect to placental function and fetal development.
Assuntos
Membrana Celular/metabolismo , Homocisteína/metabolismo , Microvilosidades/metabolismo , Placenta/metabolismo , Sistema A de Transporte de Aminoácidos/metabolismo , Sistema A de Transporte de Aminoácidos/fisiologia , Sistema L de Transporte de Aminoácidos/metabolismo , Sistema L de Transporte de Aminoácidos/fisiologia , Sistema y+L de Transporte de Aminoácidos/metabolismo , Sistema y+L de Transporte de Aminoácidos/fisiologia , Transporte Biológico , Feminino , Homocisteína/farmacocinética , Humanos , Microvilosidades/ultraestrutura , Modelos Biológicos , Placenta/ultraestrutura , Gravidez , Radioisótopos de Enxofre/farmacocinéticaRESUMO
Betaine homocysteine S-methyltransferase (BHMT) catalyzes the transfer of a methyl group from betaine to homocysteine (Hcy), forming dimethylglycine and methionine. We previously showed that inhibiting BHMT in mice by intraperitoneal injection of S-(alpha-carboxybutyl)-DL-homocysteine (CBHcy) results in hyperhomocysteinemia. In the present study, CBHcy was fed to rats to determine whether it could be absorbed and cause hyperhomocysteinemia as observed in the intraperitoneal administration of the compound in mice. We hypothesized that dietary administered CBHcy will be absorbed and will result in the inhibition of BHMT and cause hyperhomocysteinemia. Rats were meal-fed every 8 hours an L-amino acid-defined diet either containing or devoid of CBHcy (5 mg per meal) for 3 days. The treatment decreased liver BHMT activity by 90% and had no effect on methionine synthase, methylenetetrahydrofolate reductase, phosphatidylethanolamine N-methyltransferase, and CTP:phosphocholine cytidylyltransferase activities. In contrast, cystathionine beta-synthase activity and immunodetectable protein decreased (56% and 26%, respectively) and glycine N-methyltransferase activity increased (52%) in CBHcy-treated rats. Liver S-adenosylmethionine levels decreased by 25% in CBHcy-treated rats, and S-adenosylhomocysteine levels did not change. Furthermore, plasma choline decreased (22%) and plasma betaine increased (15-fold) in CBHcy-treated rats. The treatment had no effect on global DNA and CpG island methylation, liver histology, and plasma markers of liver damage. We conclude that CBHcy-mediated BHMT inhibition causes an elevation in total plasma Hcy that is not normalized by the folate-dependent conversion of Hcy to methionine. Furthermore, metabolic changes caused by BHMT inhibition affect cystathionine beta-synthase and glycine N-methyltransferase activities, which further deteriorate plasma Hcy levels.
Assuntos
Dieta , Homocisteína/análogos & derivados , Hiper-Homocisteinemia/induzido quimicamente , Aminoácidos/análise , Aminoácidos/sangue , Animais , Betaína/sangue , Betaína-Homocisteína S-Metiltransferase/antagonistas & inibidores , Colina/sangue , Cistationina beta-Sintase/análise , Cistationina beta-Sintase/metabolismo , Inibidores Enzimáticos/administração & dosagem , Glicina N-Metiltransferase/metabolismo , Homocisteína/administração & dosagem , Homocisteína/farmacocinética , Hiper-Homocisteinemia/patologia , Fígado/química , Fígado/enzimologia , Fígado/patologia , Masculino , Ratos , Ratos Endogâmicos F344 , S-Adenosil-Homocisteína/análise , S-Adenosilmetionina/análiseRESUMO
BACKGROUND: Hyperhomocysteinemia in humans is a risk factor for adverse pregnancy outcome, especially congenital malformations. This review summarizes the studies directed on the teratogenicity of homocysteine carried out in animal studies, and elaborates on the underlying mechanisms. METHODS: Literature was searched in Pubmed (NCBI) through January 2010 and selected manually. Keywords comprised homocysteine, congenital abnormalities and animals. RESULTS: Increased frequencies of a wide range of congenital malformations are reported especially in the chicken embryo after exposure to homocysteine (Hcy) in various dosages and forms. Reduced embryonic growth and abnormalities of the vascularization of the yolk sac are described in mouse studies. A study in rats revealed a reduced development of blastocysts. The congenital malformations observed in the chicken embryo model share the mutual involvement of Hcy sensitive neural crest cells. Derangements in the behavior of these cells by interactions between Hcy and pathways involved in vascularization, growth, metabolism, signaling, and DNA synthesis and methylation may explain the wide range of effects on embryonic organs, the yolk sac and placental tissues. CONCLUSIONS: The associations between human hyperhomocysteinemia and congenital malformations are substantiated by chicken and rodent studies. Moreover, derangements of several pathways induced by Hcy are demonstrated with adverse effects on both reproduction and long term health. Because of the high prevalence of hyperhomocysteinemia in both the reproductive and general population, research on underlying epigenetic mechanisms is warranted.
Assuntos
Anormalidades Induzidas por Medicamentos/metabolismo , Ácido Fólico/metabolismo , Homocisteína/farmacocinética , Homocisteína/toxicidade , Teratogênicos/farmacocinética , Teratogênicos/toxicidade , Animais , Galinhas , Anormalidades Congênitas/epidemiologia , Anormalidades Congênitas/metabolismo , Desenvolvimento Embrionário/efeitos dos fármacos , Feminino , Desenvolvimento Fetal/efeitos dos fármacos , Hiper-Homocisteinemia/induzido quimicamente , Hiper-Homocisteinemia/metabolismo , Hiper-Homocisteinemia/fisiopatologia , Camundongos , Crista Neural/efeitos dos fármacos , Gravidez , Complicações na Gravidez/induzido quimicamente , Complicações na Gravidez/metabolismo , Complicações na Gravidez/fisiopatologia , Resultado da Gravidez , RatosRESUMO
PURPOSE: This study investigated the role of an ABC transporter, Mrp3/Abcc3 in intestinal folate absorption. METHODS: Plasma concentrations of folic acid and leucovorin, given orally, were determined in wild-type and Mrp3 ( -/- ) mice. Mucosal-to-serosal transport was determined in the everted intestinal sacs. The plasma concentrations of endogenous 5-methyltetrahydrofolic acid, homocysteine and vitamin B(12), and mRNA levels of hepatic and intestinal folate metabolizing enzymes were compared between wild-type and Mrp3 ( -/- ) mice. RESULTS: C ( max ) and area-under plasma concentration-time curve of folic acid were 3.0- and 2.3-fold lower in Mrp3 ( -/- ) mice compared with wild-type mice, whereas the total body clearance was unchanged. Absorption of leucovorin was significantly delayed in Mrp3 ( -/- ) mice. Mucosal-to-serosal transport of folic acid and leucovorin was significantly decreased in the duodenum of Mrp3 ( -/- ) mice, where their PS ( serosal ) was decreased to 6.3 and 22% of that in wild-type mice, respectively. PS ( serosal ) of 5-methyltetrahydrofolic acid was moderately decreased in Mrp3 ( -/- ) mice. There was no obvious abnormality in folate homeostasis in Mrp3 ( -/- ) mice. CONCLUSIONS: Mrp3 accounts for the serosal efflux of folic acid and leucovorin, while it makes a moderate contribution to the serosal efflux of 5-methyltetrahydrofolic acid in mice. Mrp3 dysfunction does not disrupt folate homeostasis in mouse.
Assuntos
Ácido Fólico/farmacocinética , Absorção Intestinal , Proteínas Associadas à Resistência a Múltiplos Medicamentos/metabolismo , Animais , Ácido Fólico/sangue , Deleção de Genes , Homocisteína/sangue , Homocisteína/farmacocinética , Leucovorina/sangue , Leucovorina/farmacocinética , Masculino , Camundongos , Proteínas Associadas à Resistência a Múltiplos Medicamentos/genética , Tetra-Hidrofolatos/sangue , Tetra-Hidrofolatos/farmacocinética , Vitamina B 12/sangue , Vitamina B 12/farmacocinéticaRESUMO
We recently showed that the developing gut is a significant site of methionine transmethylation to homocysteine and transsulfuration to cysteine. We hypothesized that sulfur amino acid (SAA) deficiency would preferentially reduce mucosal growth and antioxidant function in neonatal pigs. Neonatal pigs were enterally fed a control or an SAA-free diet for 7 days, and then whole body methionine and cysteine kinetics were measured using an intravenous infusion of [1-(13)C;methyl-(2)H(3)]methionine and [(15)N]cysteine. Body weight gain and plasma methionine, cysteine, homocysteine, and taurine and total erythrocyte glutathione concentrations were markedly decreased (-46% to -85%) in SAA-free compared with control pigs. Whole body methionine and cysteine fluxes were reduced, yet methionine utilization for protein synthesis and methionine remethylation were relatively preserved at the expense of methionine transsulfuration, in response to SAA deficiency. Intestinal tissue concentrations of methionine and cysteine were markedly reduced and hepatic levels were maintained in SAA-free compared with control pigs. SAA deficiency increased the activity of methionine metabolic enzymes, i.e., methionine adenosyltransferase, methionine synthase, and cystathionine beta-synthase, and S-adenosylmethionine concentration in the jejunum, whereas methionine synthase activity increased and S-adenosylmethionine level decreased in the liver. Small intestine weight and protein and DNA mass were lower, whereas liver weight and DNA mass were unchanged, in SAA-free compared with control pigs. Dietary SAA deficiency induced small intestinal villus atrophy, lower goblet cell numbers, and Ki-67-positive proliferative crypt cells in association with lower tissue glutathione, especially in the jejunum. We conclude that SAA deficiency upregulates intestinal methionine cycle activity and suppresses epithelial growth in neonatal pigs.
Assuntos
Aminoácidos Sulfúricos/sangue , Aminoácidos Sulfúricos/deficiência , Células Epiteliais/metabolismo , Mucosa Intestinal/crescimento & desenvolvimento , Mucosa Intestinal/metabolismo , Metionina/metabolismo , Animais , Animais Recém-Nascidos , Divisão Celular/fisiologia , Cisteína/metabolismo , Cisteína/farmacocinética , Nutrição Enteral , Células Epiteliais/citologia , Feminino , Glutationa/metabolismo , Células Caliciformes/citologia , Células Caliciformes/metabolismo , Homocisteína/metabolismo , Homocisteína/farmacocinética , Mucosa Intestinal/citologia , Radioisótopos do Iodo , Metionina/farmacocinética , Metionina Adenosiltransferase/metabolismo , Metilação , Modelos Biológicos , Isótopos de Nitrogênio , Tamanho do Órgão , Oxirredução , Sus scrofa , Trítio , Regulação para Cima/fisiologiaRESUMO
CONTEXT: Abnormal homocysteine metabolism may contribute to increased cardiovascular death in type 1 diabetes (T1DM). Amino acid metabolism is altered in T1DM. In vitro, insulin reduces hepatic catabolism of homocysteine by inhibiting liver transsulfuration. It remains to be determined whether methionine-homocysteine metabolism is altered in T1DM. OBJECTIVE: We sought to determine whether insulin deficiency during insulin deprivation or high plasma insulin concentration after insulin treatment alters homocysteine metabolism in T1DM. DESIGN: This was an acute interventional study with paired and comparative controls. SETTING: The study was conducted at a general clinical research center. PATIENTS AND INTERVENTION: We used stable isotope tracers to measure methionine-homocysteine kinetics in six patients with T1DM during insulin deprivation (I-) and also during insulin treatment (I+) and compared them with nondiabetic controls (n = 6). MAIN OUTCOME MEASURES: Homocysteine kinetics (transmethylation, transsulfuration, and remethylation) were from plasma isotopic enrichment of methionine and homocysteine and (13)CO(2). RESULTS: T1DM (I-) had lower rates of homocysteine-methionine remethylation (P < 0.05 vs. control and I+). In contrast, transsulfuration rates were higher in I- than controls and I+ (P < 0.05). Insulin treatment normalized transsulfuration and remethylation (P < 0.05 vs. I- and P > 0.8 vs. control). Plasma homocysteine concentrations were lower in T1DM (P < 0.05 vs. control during both I- and I+), which may be explained by increased homocysteine transsulfuration. Thus, significant alterations of methionine-homocysteine metabolism occur during insulin deprivation in humans with T1DM. CONCLUSIONS: Insulin plays a key role in the regulation of methionine-homocysteine metabolism in humans, and altered homocysteine may occur during insulin deficiency in type 1 diabetic patients.
Assuntos
Diabetes Mellitus Tipo 1/metabolismo , Homocisteína/metabolismo , Insulina/deficiência , Metionina/metabolismo , Adulto , Aminoácidos de Cadeia Ramificada/sangue , Glicemia/metabolismo , Radioisótopos de Carbono , Diabetes Mellitus Tipo 1/sangue , Feminino , Homocisteína/sangue , Homocisteína/farmacocinética , Humanos , Insulina/sangue , Insulina/farmacologia , Masculino , Metionina/sangue , Metionina/farmacocinética , Pessoa de Meia-IdadeRESUMO
Hyperhomocysteinemia is an independent risk factor for cardiovascular disease. Transport of L-homocysteine into and out of the human vascular endothelium is poorly understood. We hypothesized that cultured human aortic endothelial cells (HAEC) would import L-homocysteine on one or more of the L-cysteine transport systems. Inhibitors of the transporters were used to characterize the uptake of [35S]L-homocysteine, [35S]L-homocystine, and [35S]L-cysteine. We found that L-homocysteine uptake is mediated by the sodium-dependent cysteine transport systems X(AG), ASC, and A, and the sodium-independent transport system L. Thus, HAEC utilize multiple cysteine transporters (X(AG) > or = L > ASC > A) to import L-homocysteine. Kinetic analysis supported the uptake results. Michaelis-Menten constants (Km) for the four systems yielded values of 19.0, 27.1, 112, and 1000 microM for systems L, X(AG), ASC, and A, respectively. The binding and uptake of [35S]L-homocystine, the disulfide homodimer of L-homocysteine, was mediated by systems X(AG), L, and ASC but not by system A. In contrast to [35S]L-homocysteine, system x(c) was active for [35S]L-homocystine uptake. A similar pattern was observed for [35S]L-cysteine. Thus, L-homocysteine and L-homocystine found in hyperhomocysteinemic subjects can gain entry into the vascular endothelium by way of multiple L-cysteine transporters.
Assuntos
Células Endoteliais/metabolismo , Homocisteína/metabolismo , Sistema A de Transporte de Aminoácidos/fisiologia , Sistema ASC de Transporte de Aminoácidos/fisiologia , Sistema L de Transporte de Aminoácidos/fisiologia , Sistema X-AG de Transporte de Aminoácidos/fisiologia , Aorta Torácica/citologia , Transporte Biológico Ativo , Células Cultivadas , Cisteína/metabolismo , Cisteína/farmacocinética , Endotélio Vascular/citologia , Endotélio Vascular/metabolismo , Homocisteína/farmacocinética , Humanos , Ligação Proteica , Radioisótopos de EnxofreRESUMO
Homocysteine is a precursor of S-adenosylmethionine (AdoMet) and a metabolite of S-adenosylhomocysteine (AdoHcy). The ratio of AdoMet to AdoHcy, defined as the methylation potential (MP), indicates the flow of methyl groups within the cells. Chronic elevations of total homocysteine (tHcy) in plasma correlate with increased AdoHcy concentrations, decreased MP, and impaired DNA methylation. However, the influence of acute hyperhomocysteinemia on MP is unknown. We induced acute hyperhomocysteinemia in 14 healthy volunteers by oral administration of l-homocysteine (65.1 micromol/kg body wt) in an open, randomized, placebo-controlled two-period crossover study. The kinetics of tHcy in blood and urine, MP in blood, and global DNA methylation in lymphocytes were studied systematically during 48 h. Plasma tHcy concentrations reached a peak at 34 +/- 11 min after an oral load with l-homocysteine and decreased with a half-life of 257 +/- 41 min (means +/- SD). Only 2.3% of the homocysteine dose were recovered in urine. AdoHcy concentrations and MP in whole blood and erythrocytes were not affected by the oral homocysteine load. Furthermore, global DNA methylation in lymphocytes did not change under these conditions. We found no difference between the genotypes of 5,10-methylenetetrahydrofolate reductase in response to the homocysteine load. However, AdoMet content in erythrocytes was significantly higher in the C677T carriers (CT; n = 7) compared with the CC genotype (n = 7). Although chronic elevation of tHcy has been shown to affect MP and DNA methylation, acute elevation of plasma tHcy above 20 micromol/l for 8 h is not sufficient to change MP and to induce DNA hypomethylation in lymphocytes.
Assuntos
Metilação de DNA/efeitos dos fármacos , Eritrócitos/metabolismo , Homocisteína/sangue , Linfócitos/metabolismo , Adulto , Estudos Cross-Over , Genótipo , Meia-Vida , Heterozigoto , Homocisteína/farmacocinética , Homocisteína/farmacologia , Humanos , Inulina , Cinética , Masculino , S-Adenosilmetionina/sangueRESUMO
BACKGROUND: The effects of vitamin B-6 status on steady-state kinetics of homocysteine metabolism in humans are unclear. OBJECTIVE: The objective was to determine the effects of dietary vitamin B-6 restriction on the rates of homocysteine remethylation and synthesis in healthy humans. DESIGN: Primed, constant infusions of [(13)C(5)]methionine, [3-(13)C]serine, and [(2)H(3)]leucine were conducted in healthy female (n=5) and male (n=4) volunteers (20-30 y) before and after 4 wk of dietary vitamin B-6 restriction (<0.5 mg vitamin B-6/d) to establish whether vitamin B-6 status affects steady-state kinetics of homocysteine metabolism in the absence of concurrent methionine intake. Effects of dietary vitamin B-6 restriction on vitamin B-6 status, plasma amino acid concentrations, and the rates of reactions of homocysteine metabolism were assessed. RESULTS: Dietary vitamin B-6 restriction significantly reduced plasma pyridoxal 5-phosphate (PLP) concentrations (55.1 +/- 8.3 compared with 22.6 +/- 1.3 nmol/L; P=0.004), significantly increased plasma glycine concentrations (230 +/- 14 compared with 296 +/- 15; P=0.008), and significantly reduced basal (43%; P < 0.001) and PLP-stimulated (35%; P=0.004) lymphocyte serine hydroxymethyltransferase activities in vitro. However, the in vivo fluxes of leucine, methionine, and serine; the rates of homocysteine synthesis and remethylation (total and vitamin B-6-dependent); and the concentrations of homocysteine, methionine, and serine in plasma were not significantly affected by dietary vitamin B-6 restriction. CONCLUSIONS: Moderate vitamin B-6 deficiency does not significantly alter the rates of homocysteine remethylation or synthesis in healthy young adults in the absence of dietary methionine intake.
Assuntos
Homocisteína/metabolismo , Fosfato de Piridoxal/sangue , Vitamina B 6/farmacologia , Adulto , Isótopos de Carbono , Deutério , Feminino , Glicina/sangue , Homocisteína/farmacocinética , Humanos , Masculino , Metionina/administração & dosagem , Metionina/metabolismo , Metionina/farmacocinética , Metilação , Metilenotetra-Hidrofolato Redutase (NADPH2)/genética , Polimorfismo Genético , Serina/administração & dosagem , Serina/metabolismo , Serina/farmacocinética , Vitamina B 6/administração & dosagemRESUMO
Methionine is a sulfur-containing amino acid that is reversibly converted into homocysteine. Homocysteine is an independent cardiovascular risk factor frequently associated with the insulin resistance syndrome. The effects of insulin on methionine and homocysteine kinetics in vivo are not known. Six middle-aged male volunteers were infused with L-[methyl-2H3,1-13C]methionine before (for 3 h) and after (for 3 additional hours) an euglycemic hyperinsulinemic (150 mU/l) clamp. Steady-state methionine and homocysteine kinetics were determined using either plasma (i.e., those of methionine) or intracellular (i.e., those of plasma homocysteine) enrichments. By use of plasma enrichments, insulin decreased methionine rate of appearance (Ra; both methyl- and carbon Ra) by 25% (P < 0.003 vs. basal) and methionine disposal into proteins by 50% (P < 0.0005), whereas it increased homocysteine clearance by approximately 70% (P < 0.025). With intracellular enrichments, insulin increased all kinetic rates, mainly because homocysteine enrichment decreased by approximately 40% (P < 0.001). In particular, transmethylation increased sixfold (P < 0.02), transsulfuration fourfold (P = 0.01), remethylation eightfold (P < 0.025), and clearance eightfold (P < 0.004). In summary, 1) physiological hyperinsulinemia stimulated homocysteine metabolic clearance irrespective of the model used; and 2) divergent changes in plasma methionine and homocysteine enrichments were observed after hyperinsulinemia, resulting in different changes in methionine and homocysteine kinetics. In conclusion, insulin increases homocysteine clearance in vivo and may thus prevent homocysteine accumulation in body fluids. Use of plasma homocysteine as a surrogate of intracellular methionine enrichment, after acute perturbations such as insulin infusion, needs to be critically reassessed.
Assuntos
Homocisteína/sangue , Homocisteína/farmacocinética , Hiperinsulinismo/metabolismo , Metionina/sangue , Metionina/farmacocinética , Glicemia/metabolismo , Testes Respiratórios , Técnica Clamp de Glucose , Humanos , Hiperinsulinismo/sangue , Insulina/sangue , Isoenzimas , Cinética , Masculino , Metilenotetra-Hidrofolato Redutase (NADPH2)/metabolismo , Pessoa de Meia-Idade , Técnica de Diluição de RadioisótoposRESUMO
The epithelial cells that line the renal proximal tubule have been shown to be the primary cellular targets where mercuric ions gain entry, accumulate, and induce pathologic effects in vivo. Recent data have implicated at least one of the organic anion transport systems in the basolateral uptake of inorganic mercury (Hg). With the use of a line of type II MDCK cells transfected stably with the human organic anion transporter 1 (hOAT1), the hypothesis that hOAT1 can transport mercuric conjugates of homocysteine (Hcy) was tested. Indeed, MDCK II cells expressing a functional form of hOAT1 gained the ability to transport the mercuric conjugate 2-amino-4-(3-amino-3-carboxy-propylsulfanylmercuricsulfanyl) butyric acid (Hcy-S-Hg-S-Hcy). In addition, p-aminohippurate and the dicarboxylates adipate and glutarate (but not succinate or malonate) inhibited individually the uptake of Hcy-S-Hg-S-Hcy in a concentration-dependent manner. Furthermore, a direct relationship between the uptake of Hcy-S-Hg-S-Hcy and the induction of cellular injury and death was demonstrated in the hOAT1-expressing MDCK II cells only. These data represent the first line of direct evidence implicating one of the organic anion transporters in the uptake of a mercuric conjugate of Hcy in a mammalian cell. Thus, mercuric conjugates of Hcy are potential transportable substrates of OAT1. More important, the findings from the present study implicate the activity of OAT1 in the uptake and toxicity of Hg (when in the form of Hcy-S-Hg-S-Hcy in the extracellular compartment) in proximal tubular epithelial cells in vivo.
Assuntos
Células Epiteliais/metabolismo , Homocisteína/farmacocinética , Túbulos Renais/citologia , Mercúrio/farmacocinética , Proteína 1 Transportadora de Ânions Orgânicos/metabolismo , Adipatos/farmacologia , Animais , Transporte Biológico , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Ácidos Dicarboxílicos/farmacologia , Células Epiteliais/citologia , Glutaratos/farmacologia , Homocisteína/química , Humanos , Mercúrio/química , Intoxicação do Sistema Nervoso por Mercúrio/metabolismo , Proteína 1 Transportadora de Ânions Orgânicos/genética , Temperatura , Ácido p-Aminoipúrico/farmacocinéticaRESUMO
Homocysteine has recently received a lot of attention as an independent risk factor for atherosclerotic and thrombotic cardiovascular disease. Plasma homocysteine levels tend to rise with age, but are also greatly influenced by nutritional factors. Early reports suggested that there were differences in the metabolism of homocysteine in adult and immature animals. The current work tests the hypothesis that adult and juvenile animals respond differently to chronic administration of homocysteine. We have previously found that adult rabbits given homocysteine parenterally twice daily for seven weeks developed progressive folate deficiency and concurrently developed an impairment of homocysteine metabolism. We now report that juvenile rabbits do not develop folate deficiency with chronic homocysteine loading and do not have progressively higher trough levels of homocysteine, as do the adults. In addition, juvenile rabbits that have been chronically pre-treated with homocysteine exhibit a lower peak homocysteine level after a single dose than do juvenile rabbits that have never received homocysteine. This adaptation did not occur in the adult rabbits. In addition, adult homocysteine-treated rabbits had evidence of oxidative stress as evidenced by higher levels of malondialdehyde in liver tissue than adult controls. The homocysteine-treated juvenile rabbits had the same levels of malondialdehyde as the juvenile control rabbits. We conclude that the plasma elimination kinetics are altered in juvenile rabbits in response to homocysteine pre-treatment. The difference in metabolism of homocysteine may protect the juvenile rabbits from the damaging effects of homocysteine. Future studies are planned to elucidate the mechanism of this adaptive response.
Assuntos
Envelhecimento/fisiologia , Homocisteína/farmacologia , Animais , Modelos Animais de Doenças , Ácidos Graxos/análise , Ácido Fólico/sangue , Glutationa/análise , Homocisteína/sangue , Homocisteína/farmacocinética , Peroxidação de Lipídeos/efeitos dos fármacos , Fígado/química , Taxa de Depuração Metabólica , Coelhos , Substâncias Reativas com Ácido Tiobarbitúrico/análiseRESUMO
OBJECTIVE: The purpose of the present work was to determine the fasting plasma total homocysteine (tHcy) levels and the time-course response of tHcy concentrations after the consumption of urban and rural Mexican model diets in two groups of Mexican women from urban and rural areas. METHODS: Thirty-three adult women (age range = 18-49 y) were studied. Fifteen women were from a rural community in the state of Mexico. The other 18 were from cities and consumed diets that regularly included an important amount of animal foods. The study was designed as a two-period crossover study in which subjects consumed the model urban or rural diet in a 2-wk interval. Seven milliliters of venous blood was drawn before ingestion of experimental diets (basal) to measure total cholesterol, high-density lipoprotein cholesterol, triacylglycerol, tHcy, folate, vitamin B12, and methionine. Blood samples were then obtained 30, 60, 90, 180, and 240 min after the beginning of meal consumption. RESULTS: The rural and urban groups showed similar concentrations of tHcy 4 h after meal consumption and after fasting. However, the urban and rural groups had higher methionine plasma concentrations after the urban diet than after the rural diet. In contrast, there was no significant difference in methionine plasma levels between the rural and urban groups with each diet. Those women with low tHcy concentrations maintained those values over the study period, and those with high tHcy concentrations maintained those values. There was no significant difference in tHcy concentrations due to consumption of the two diets (P = 0.31) or the interaction between population and diet (P = 0.84). However, there was a significant difference in the concentration of tHcy between the rural (8.73 +/- 0.17 microM/L) and the urban (9.27 +/- 0.13 microM/L) populations (P = 0.01). In both groups, average tHcy concentration was in the normal range. In both populations, the nutrition status for folate and vitamin B12 was adequate, although plasma folate concentration was significantly lower in the rural population than in the urban population (P < 0.01). Plasma vitamin B12 concentrations were similar in both groups. No subject had low plasma vitamin B12. CONCLUSIONS: Plasma tHcy concentrations in rural and urban Mexican women were within the range considered adequate; however, urban women showed significant higher concentrations than did rural women independently of the consumed diet and the plasma methionine concentration. These results indicated that there is no short-term variation in plasma tHcy due to the consumption of rural or urban diets.
Assuntos
Dieta , Homocisteína/sangue , Adolescente , Adulto , Estudos Cross-Over , Jejum , Feminino , Ácido Fólico/sangue , Homocisteína/farmacocinética , Humanos , Metionina/sangue , México , Pessoa de Meia-Idade , Estado Nutricional , Valores de Referência , Saúde da População Rural , Saúde da População Urbana , Vitamina B 12/sangueRESUMO
This study was designed to investigate the effects of captopril, an angiotensin-converting enzyme inhibitor, on inhibition of endothelium-dependent relaxation induced by homocysteine in isolated rat aorta. Isometric tension recordings were used to assess inhibitory effects of homocysteine and protective effects of captopril on endothelium-dependent relaxation of aortic rings. Exposure of aortic rings to homocysteine (0.3 approximately 3 mmol/L) for 30 min induced a significant concentration-dependent inhibition of endothelium-dependent relaxation response to acetylcholine (ACh), but did not affect endothelium-independent relaxation response to sodium nitroprusside. Pre-incubation of aortic rings with captopril (3 approximately 30 micromol/L) for 15 min and co-incubation of aortic rings with homocysteine (1 mmol/L) for another 30 min attenuated the inhibition of homocysteine in a dose-dependent manner. Moreover, superoxide dismutase (SOD, 200 U/mL), a scavenger of superoxide anions, reduced homocysteine-induced inhibition. L-Arginine (3 mmol/L), a precursor of nitric oxide (NO), also attenuated the impairment of vasorelaxation induced by homocysteine. However, in the combined presence of SOD and L-arginine, the inhibitory effect of homocysteine was reversed, which was very similar to the effect of 30 micromol/L captopril. These results suggest that captopril can prevent the inhibition of endothelium-dependent relaxation induced by homocysteine in isolated rat aorta, which may be related to scavenging oxygen free radicals and enhancing NO production.
Assuntos
Aorta Torácica/efeitos dos fármacos , Captopril/farmacologia , Endotélio Vascular/efeitos dos fármacos , Homocisteína/efeitos adversos , Homocisteína/farmacocinética , Relaxamento Muscular/efeitos dos fármacos , Acetilcolina/administração & dosagem , Acetilcolina/antagonistas & inibidores , Acetilcolina/farmacocinética , Animais , Aorta Torácica/fisiopatologia , Arginina/farmacocinética , Captopril/administração & dosagem , Relação Dose-Resposta a Droga , Endotélio Vascular/fisiopatologia , Homocisteína/antagonistas & inibidores , Masculino , Relaxamento Muscular/fisiologia , Músculo Liso Vascular/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Superóxido Dismutase/farmacocinética , Vasodilatação/efeitos dos fármacosRESUMO
Hyperhomocysteinemia occurs in homocystinuria, an inherited metabolic disease clinically characterized by thromboembolic episodes and a variable degree of neurological dysfunction whose pathophysiology is poorly known. In this study, we induced elevated levels of homocysteine (Hcy) in blood (500 microM), comparable to those of human homocystinuria, and in brain (60 nmol/g wet tissue) of young rats by injecting subcutaneously homocysteine (0.3-0.6 micromol/g of body weight) twice a day at 8-hr intervals from the 6th to the 28th postpartum day. Controls received saline in the same volumes. Na(+),K(+)-ATPase and Mg(2+)-ATPase activities were determined in the hippocampus of treated Hcy- and saline-treated rats. Chronic administration of Hcy significantly decreased (40%) Na(+),K(+)-ATPase activity but did not alter Mg(2+)-ATPase activity. Considering that Na(+),K(+)-ATPase plays a crucial role in the central nervous system, our results suggest that the brain dysfunction found in homocystinuria may be related to the reduction of brain Na(+),K(+)-ATPase activity.
Assuntos
Hipocampo/enzimologia , Homocisteína/sangue , ATPase Trocadora de Sódio-Potássio/metabolismo , Animais , Homocisteína/administração & dosagem , Homocisteína/farmacocinética , Ratos , Ratos WistarRESUMO
Folate and vitamin B(6) act in generating methyl groups for homocysteine remethylation, but the kinetic effects of folate or vitamin B(6) deficiency are not known. We used an intravenous primed, constant infusion of stable isotope-labeled serine, methionine, and leucine to investigate one-carbon metabolism in healthy control (n = 5), folate-deficient (n = 4), and vitamin B(6)-deficient (n = 5) human subjects. The plasma homocysteine concentration in folate-deficient subjects [15.9 +/- 2.1 (SD) micromol/l] was approximately two times that of control (7.4 +/- 1.7 micromol/l) and vitamin B(6)-deficient (7.7 +/- 2.1 micromol/l) subjects. The rate of methionine synthesis by homocysteine remethylation was depressed (P = 0.027) in folate deficiency but not in vitamin B(6) deficiency. For all subjects, the homocysteine remethylation rate was not significantly associated with plasma homocysteine concentration (r = -0.44, P = 0.12). The fractional synthesis rate of homocysteine from methionine was positively correlated with plasma homocysteine concentration (r = 0.60, P = 0.031), and a model incorporating both homocysteine remethylation and synthesis rates closely predicted plasma homocysteine levels (r = 0.85, P = 0.0015). Rates of homocysteine remethylation and serine synthesis were inversely correlated (r = -0.89, P < 0.001). These studies demonstrate distinctly different metabolic consequences of vitamin B(6) and folate deficiencies.