Durante a fase de crescimento e terminação, 1890 frangos de corte machos da linhagem Cobb®500 foram submetidos a altas temperaturas e alimentados com dietas experimentais contendo três fontes de metionina (DL-Met, DL-HMTBA e DL-Met/ DL-HMTBA), três níveis de vitamina E (15 UI, 45 UI e 100 UI) e duas relações Arg:Lis (0,9 e 1,5), cujo delineamento fatorial 3 x 3 x 2 totalizou 18 tratamentos, com sete repetições de 15 aves cada. Ao final do período experimental, os parâmetros de desempenho zootécnico como consumo de ração, peso corporal, conversão alimentar e rendimento de carcaça, peito e pernas foram avaliados e, após análise estatística, comprovou-se que não houve interação entre os fatores. Dessa maneira, os efeitos individuais dos elementos testados foram analisados e observou-se que a utilização de diferentes fontes de metionina e a inclusão de vitamina E nas quantidades testadas não apresentaram diferenças significativas. Porém, a maior inclusão de arginina na dieta proporcionou melhora nos resultados de ganho de peso, peso corporal e conversão alimentar, mas não resultou em diferenças significativas nos parâmetros de carcaça avaliados. / During the growing and the finishing, 1890 male broiler chickens of Cobb 500 ® were subjected to high temperatures and fed with experimental diets containing three sources of methionine (DL -Met ,DL - HMTBA and DL-Met/DL - HMTBA), three levels of vitamin E (15 IU, 45 IU and 100 IU) and two relations Arg:Lys (0.9 and 1.5), whose factorial design 3 x 3 x 2 totaled 18 treatments with seven replicates of 15 birds each. At the end of the experimental period, the growth performance parameters such as feed intake, body weight, feed conversion and carcass, breast and legs yieldwere evaluated and, after statistical analysis, it was shown that there was no interaction between factors. Thus, the effects of the tested individual elements were analyzed and it was found that the use of different sources of methionine and the inclusion of vitamin E in the amounts tested showed no significant differences. However, the increased inclusion of arginine in the diet improved results in weight gain, body weight and feed conversion, but resulted in no significant differences in carcass parameters.

A colina é um nutriente essencial nas dietas de frangos de corte. Os valores de exigências nutricionais são variáveis e foram estabelecidos há muitos anos. Assim, o objetivou-se com este trabalho reavaliar as respostas de frangos de corte de alto potencial genético de 1 a 7, 1 a 14 e 1 a 21 dias de idade como também modelar as respostas por diferentes funções matemáticas. Foram conduzidos dois experimentos sob dois níveis de metionina na dieta, tendo como base o método dose-resposta. Para os ensaios, foram utilizados 2.160 pintos de corte machos da linhagem Cobb 500 (1.080 em cada experimento), distribuídos em um delineamento inteiramente casualizado, com seis tratamentos, seis repetições e 30 aves por unidade experimental. Os tratamentos consistiram em rações com níveis de 390, 715, 1.040, 1.365, 1.690 e 2.015 mg/kg de colina. As rações experimentais foram formuladas utilizando-se a técnica da suplementação. Foi formulada uma ração basal à base de milho, farelo de soja, concentrado proteico de soja, açúcar e amido contendo 390 mg/kg de colina total e 0,596% de metionina digestível. O cloreto de colina (62,5%) comercial foi suplementado para obter os demais níveis. O segundo experimento consistiu nos mesmos tratamentos do primeiro, porém com atendimento de 0,440% da metionina digestível. As variáveis mensuradas em ambos os ensaios foram: consumo de ração, consumo de colina, conversão alimentar e ganho de peso. Para modelar as respostas considerou-se o consumo de colina como variável independente e ganho de peso como variável dependente. As respostas foram modeladas por cinco modelos matemáticos: broken line, broken line com ascendência quadrática, cinética de saturação, logístico e monomolecular e o melhor modelo foi selecionado pelo critério de Akaike. As análises estatísticas foram realizadas com auxílio do programa computacional SAS® utilizando o procedimento PROC GLM para análise das pressuposições e análise de variância e PROC NLIN para o ajuste dos modelos. Os modelos que melhor se ajustaram foram: Broken line com ascendência quadrática (1,26 mg/g de GP ensaio 1) e Logístico (1,26 mg/g de GP ensaio 2) para o período de 1 a 7 dias. No período de 1 a 14 dias os modelos selecionados para o experimento com metionina normal e reduzida foram Logístico (1,16 mg/g de GP) e Broken line com ascendência quadrática (1,28 mg/g de GP), respectivamente. Já para o período de 1 a 21 foram o cinética de saturação, estimando uma exigência de 1,35 mg/g de GP para o experimento com metionina normal, e Broken line com ascendência quadrática estimando uma exigência de 1,21 mg de colina/g de GP para o experimento com metionina reduzida. / Choline is an essential nutrient in the diets of chicks. Supplementation levels in feeds is variable and requirements have been established many years ago. Thus, the objective of this study was to reassess the responses of high genetic potential chicks from 1-7, 1-14 and 1 to 21 days as well as modeling the responses of different mathematical functions. Two trials were conducted using two dietary levels of methionine, based on the dose-response method. For the trials were used 2,160 Cobb 500 male broiler chicks (1,080 birds in each experiment), distributed in a completely randomized design with six treatments and six replicates and 30 birds per experimental unit. The treatments consisted of diets with levels of 390, 715, 1,040, 1,365, 1,690 and 2,015 mg/kg of choline. The diets were formulated using the supplementation method. The basal diet was formulated based on corn, soybean meal, soy protein concentrate, sugar and starch, and contained 390 mg/kg of choline and 0.596% of digestible methionine. Commercial choline chloride (62.5%) was supplemented to obtain the other levels. The second trial consisted of the same treatments of the first, but with 0.440% of digestible methionine. The variables measured in both trials were: feed intake (FI), intake of choline (IC), feed conversion (FC) and weight gain (WG). To model the responses, the variables used were the intake of choline as an independent variable and weight gain as the dependent variable. The responses were modeled by five mathematical models: broken line (ascending linear and quadratic segments), saturation kinetic, 4-parameter logistics and monomolecular models and the best model was selected by Akaike information criterion. Statistical analyzes were performed using the computer program SAS® using PROC GLM for analysis of the assumptions and analysis of variance and PROC NLIN to fit the models. The models that best fit were: broken line quadratic segments (1.26 mg/g of WG - trial 1) and Logistics (1.26 mg/g of WG - trial 2) for the period 1-7 days. In the period 1-14 days, the models selected for the experiment with normal and deficient methionine were Logistic (1.16 mg/g WG) and broken line quadratic segments (1.28 mg/g WG), respectively. For the period 1-21days, the models were the saturation kinetics, estimating a requirement of 1.35 mg/g of WG for the experiment with normal methionine, and broken line quadratic segments estimating a requirement of 1.21 mg choline/g of WG for the experiment with deficient methionine.

Certaines cellules cancéreuses sont méthionine dépendantes cependant les mécanismes de cette méthionine dépendance sont inconnus. Les cellules initiatrices de tumeur, qui représentent un faible pourcentage des cellules d’une tumeur, sont impliquées dans la récidive du cancer, phénocopient les cellules souches cancéreuses et forment des sphéroïdes 3D ou «tumor spheres (TS)» dans des conditions de culture non adhérentes. Nous montrons que, contrairement aux cellules monocouches adhérentes U251, les TS dérivées de cellules de glioblastome U251 ont besoin de méthionine exogène pour se développer. Cette méthionine-dépendance est caractérisée par une courbe en forme de cloche dans laquelle la croissance des TS est ralentie par des concentrations élevées de méthionine (> 0,01mM). Pendant la restriction en méthionine, le 5-méthyle-tétrahydrofolate restaure la formation des TS. Si les TS sont privées de méthionine pendant 24h, puis supplémentées en acide folique, elles présentent des concentrations d'isoformes des folates significativement inférieures à celles retrouvées dans les cellules adhérentes maintenues dans les mêmes conditions. Ceci suggère que le cycle des folates est réprimé dans les TS comparativement aux cellules adhérentes. L'annotation fonctionnelle des données ARN-Seq montre des changements nets dans plusieurs fonctions moléculaires et dévoile dans les TS un cycle cellulaire réduit, une augmentation du caractère « stemness » et une diminution du métabolisme des folates affectant particulièrement DHFR, SHMT et MTFHD. L'analyse du méthylome révèle des changements de méthylation dans le cycle cellulaire, la signature « stemness » et le cycle des folates, malgré des profils globaux de méthylation de l'ADN qui restent stables. Cependant, contrairement à la méthylation importante des promoteurs observée pour le cycle cellulaire et les gènes « stemness » (+ 25%), seuls 10 gènes du cycle des folates sur 139 gènes impliqués dans le métabolisme des mono-carbones sont significativement modifiés. En conclusion, un cycle des folates avec activité réduite fait partie de la reprogrammation métabolique qui déclenche la dédifférenciation en cellules souches cancéreuses et cette répression ne s'explique qu’en partie par la modification de méthylation des promoteurs / Some cancer cells are methionine dependent however little is known about the mechanisms of this dependency. Tumor initiating cells are a rare population of cancer cells, implicated in disease recurrence, that phenocopy cancer stem cells and form 3D spheroids or ‘tumor spheres (TS)» under non adherent conditions. We show that, unlike U251 adherent monolayer cells, TS derived from U251 glioblastoma cells need exogenous methionine to grow. This methionine dependency is characterized by a bell shape curve in which high methionine concentrations (>0.01mM) slow down TS growth. During methionine restriction, 5- methyltetrahydrofolate restores TS formation. When TS are deprived from methionine for 24h, then supplemented with folic acid, they exhibit lower levels of folate isoforms than adherent cells maintained in the same conditions, suggesting that folate cycle is repressed in TS relative to adherent cells. Functional annotation of the RNA-seq data shows clear changes in several molecular functions and reveals in TS a reduced cell cycle, an increased stemness and a diminished folate metabolism affecting particularly DHFR, SHMT and MTFHD. Methylome analysis shows methylation changes in cell cycle, stemness and folate cycle, despite global DNA methylation patterns remaining stable. However, unlike the important promoter methylation observed for cell cycle and stemness genes (+25%), only 10 folate cycle genes out of 139 genes involved in one-carbon metabolism are significantly altered. In conclusion, reduced folate cycle is part of the metabolic reprogramming triggering dedifferenciation into cancer stem cells and this repression is only partly explained by the alteration of promoter methylation

A metionina é doadora de grupos metil para metilação do DNA, processo responsável por modificações da expressão gênica. Diante da importância da metionina para o crescimento e desenvolvimento normais, variações desse aminoácido na dieta podem alterar a estabilidade do DNA. O objetivo desse trabalho foi avaliar o efeito de dietas deficiente e suplementada em metionina sobre a instabilidade genômica e o estresse oxidativo em camundongos e suas mães tratadas durante gestação e lactação e destas também foi realizada a expressão de RNAm de Mat1a, Bhmt e Cbsdas vias de transmetilação, remetilação e transsulfuração da metionina, respectivamente, em fígado. As fêmeas foram divididas nos grupos de dietas de metionina (controle, 0,3% DL-metionina; suplementado, 2,0%; e deficiente 0%) até o fim da lactação (10 semanas) e, para cada grupo de fêmeas, os filhotes foram subdivididos e também receberam essas dietas durante 18 semanas após desmame. Foram avaliados o consumo de ração, massa corpórea e massa relativa de fígado e rinse a taxa de sobrevivência dos filhotes. Também foram realizadas a avaliação da peroxidação lipídica (quantificação das substâncias reativas ao ácido tiobarbitúrico, TBARS), da quantificação de glutationa (GSH)e da atividade da catalase, da instabilidade genômica (ensaio do cometa) e a análise do RNAm deMat1a, BhmteCbs somente em fígado das fêmeas. A dieta deficiente resultou em menor consumo de ração e massa corpórea em ambas fases e reduziu a sobrevivência dos filhotes que receberam essa dieta.A suplementação reduziu a concentração de TBARS nas fêmeas em ambos tecidos e a deficiência não diferiu. Nos filhotes, a suplementação reduziu a concentração de TBARS em fígado, enquanto que a deficiência aumentou. A suplementação aumentou a concentração de GSH em fígado das fêmeas, assim como a deficiência em rins. Nos filhotes, houve uma inversão de respostas, ou seja, a suplementação reduziu a concentração de GSH em fígado, assim como a deficiência, também observada em rins. Não houve diferença na catalase das fêmeas, mas houve redução em ambos tecidos dos filhotes. A suplementação e a deficiência reduziram os danos ao DNA hepático das fêmeas, mas em rins a suplementação aumentou e a deficiência reduziu. Nos filhotes, a suplementação e a deficiência aumentaram os danos ao DNA em ambos tecidos. A suplementação de metionina em fêmeas não alterou a expressão dos RNAm avaliados e a deficiência reduziu em Cbs somente. Concluiu-se que na fase materna, a suplementação ou a deficiência de metionina não resultou em estresse oxidativo, mas a suplementação reduziu a instabilidade genômica no fígado e aumentou no rim. A deficiência resultou em menor instabilidade nos dois tecidos. Na fase descendente, a suplementação e a deficiência de metionina apresentaram variação de estresse oxidativo em ambos tecidos e também resultaram em maior instabilidade genômica. / Methionine is the main methyl donor for the DNA methylation, a process responsible for gene expression modifications. Since this essential amino acid is required for normal growth and development, variations of this compound in the diet may lead to alterations on DNA stability. Thus, this study aimed to evaluate the effect of deficient and supplemented methionine diets on oxidative stress and genomic instability in mice and their dams treated during pregnancy and lactation, and the expression of Mat1a, Bhmt and Cbs mRNA of transmethylation, remetthylation, and transulfuration pathways, respectively, in dams livers. The dams were divided into three methionine diets groups (control, 0,3% DL-methionine; supplemented, 2,0%; and deficient, 0%) until the end of lactation (10 weeks). For each dams groups, the offspring were subdivided and were also treated with the same diets during 18 weeks after weaning. The parameters evaluated were food intake, body weight, relative liver and kidney weights, and survival of the offspring. Also, it was carried out theevaluation of lipid peroxidation (thiobarbituric acid reactive substances, TBARS), quantification of glutathione (GSH) and catalase activity, and genomic instability (comet assay) in liver and kidneys; andMat1a, Bhmt, and CbsmRNA analysis only in dams liver. The deficient diet resulted in lower food intake and body weights in both phases and reduced the survival of the offspring that were treated with this diet. The supplemented diet reduced the TBARS concentration in both tissues of dams and the deficient diet did not differ. In the offspring, the supplementation reduced liver TBARS, whereas the deficiency raised. The supplementation increased the liver GSH concentration of dams, as well as the deficiency in kidneys. In the offspring, the responses were different; the supplementation reduced the liver GSH, as well as the deficiency, also observed in kidneys. There were no differences of catalase parameter of dams, but there was a reduction in both tissues of the offspring. Both supplementation and deficiency reduced the liver DNA damage of dams, however the supplementation increased and the deficiency reduced the DNA damage of kidneys. In the offspring, both diets increased the DNA damage in both tissues. The methionine supplementation did not differ the mRNA expression and the deficiency only reduced the Cbs, mRNA expression. It was concluded that the methionine supplementation or deficiency did not resulted in oxidative stress in dams, but the supplementation reduced the genomic instability of liver and raised the kidney one. The deficiency resulted in lower genomic instability in both tissues in dams. In the offspring, both methionine supplementation and deficiency presented variation of oxidative stress in both tissues and resulted more genomic instability.

Made available in DSpace on 2016-04-28T14:59:43Z (GMT). No. of bitstreams: 1 2005-Fabiana Goes de Almeida Nobre.pdf: 412358 bytes, checksum: fd0b391dfbb07e357a843eb24048d913 (MD5) Previous issue date: 2005-08-30 / This research was carried out in the Institute of Zootecnia / Universidade Federal Rural do Rio de Janeiro, Serop?dica, Brazil, with the objective of evaluating the levels of methionine+cystine in the slow-growing broiler chickens, raised in conventional pen and in free-range farming. The study consisted of 360 broilers at 28 days of age distributed in 15 boxes in the conventional pen and 15 free-range farming areas with Tifton 85 grass. The experimental line used was a completely randomized design in a factorial arrangement (5 X 2) with 5 levels of supplementation of DL-metionine (0.000; 0.1515; 0.3030; 0.4545; 0.6060%) and two kinds of raising pens. The basal diet of corn and soybean-meal had a 16.039% CP and 2.900Kcal/Kg ME with 0.56% of metionine+cystine total. Feed intake, weight gain, and feed efficiency were evaluated during the experimental periods 1, 2 and 3 (respectively 28 to 49, 28 to 70 and 28 to 85 days of age. At the end of each experimental period the birds were slaughtered simulating the ages of slaughter in the conventional, organic and free-range systems, respectively 49, 70, and 85 days of age. The following weights chilled carcass, cold carcass, breast, legs, heart, gizzard, abdominal fat, and carcass income. The results yielded the following ideal levels: 1.05% for feed efficiency and 0.93% based on weight gain in period 1; 0.85% for the highest weight of gizzards raised in conventional pen at 49 days of age; 0.89% for feed efficiency in period 2; 0.87% for the highest weight of abdominal fat in the broilers raised in free-range farming area slaughtered at 70 days of age; 0.89% was the lowest weight of broiler gizzards raised in free-range farming areas slaughtered at 85 days of age. The level of 0,71% of metionine+cystine was more economic in all periods. The permanence of broilers in good living conditions had an effect on the relation between feed ingestion and weight gain specially when slowgrowing broiler chickens slaughtered later. / Um experimento foi conduzido no Setor de Avicultura do Instituto de Zootecnia da Universidade Federal Rural do Rio de Janeiro, com o objetivo de avaliar os n?veis de metionina+cistina na alimenta??o de frangos de corte de linhagem comercial de crescimento lento, criados em galp?o convencional e em abrigos com livre acesso a piquetes. A partir dos 28 dias de idade, 360 aves foram distribu?das em 15 boxes de um galp?o convencional e 15 abrigos com acesso a piquetes formados com a forrageira Tifton 85. O delineamento experimental utilizado foi o inteiramente casualizado, em um esquema fatorial 5X2, com cinco n?veis de suplementa??o de DL-metionina (0,000; 0,1515; 0,3030; 0,4545; 0,6060%) e dois tipos de alojamento. A dieta basal, a base de milho e farelo de soja, continha 16,039%PB e 2.900 kcalEM/kg com 0,56% de metionina+cistina total. Foram avaliados: consumo de ra??o, ganho de peso e convers?o alimentar nos per?odos experimentais I, II e III ( respectivamente, 28 a 49, 28 a 70 e 28 a 85 dias de idade). No final de cada per?odo experimental foram feitas avalia??es, correspondendo ?s idades de abate em sistema convencional, org?nico e caipira, respectivamente, 49, 70 e 85 dias de idade, para peso da carca?a quente, peso da carca?a fria, rendimento de carca?a, de peito e pernas, al?m do peso do cora??o, moela e gordura abdominal. Por an?lises de regress?o, observou-se que no per?odo I, o n?vel de 1,05% seria o melhor, considerando-se a convers?o alimentar e 0,93% quando se avaliou o ganho de peso, nos dois tipos de alojamentos. O peso da moela, no entanto, sofreu influ?ncia dos n?veis e do alojamento, com os maiores resultados, respectivamente, para 0,85% e galp?o convencional. No per?odo II, a convers?o alimentar foi melhor para ponto correspondente a um n?vel de 0,89%, e o maior peso de gordura abdominal em 0,87%, considerando ambos os alojamentos. Entretanto o n?vel de 0,89%, quando se avaliou o abate no per?odo III, correspondeu ao menor peso da moela dos frangos com acesso a piquetes. Pela an?lise de custo, o n?vel de 0,71% de metionina+cistina seria considerado o mais econ?mico para todos os per?odos. A perman?ncia dos frangos em condi??es favor?veis de bem-estar, teve influ?ncia na rela??o entre o consumo de ra??o e o ganho de peso, principalmente quando abatidos tardiamente.

Chlamydomonas reinhardtii can utilize CO2 for autotrophic growth (HSM plus 5% CO2) or acetate for mixotrophic growth (TAP). This study was to elucidate the differential regulation of methionine sulfoxide reductase (MSR) gene expression between HSM plus 5% CO2 and TAP cultured cells, and also to determine the difference of gene expression in response to high light (1,000 £gE m-2 s-1). The role of photosynthetic electron transport (PET) in the regulation of MSR gene expression was also examined by the use of PET inhibitors. High light inhibited PSII activity (Fv/Fm and Fv'/Fm') of HSM plus 5% CO2 and TAP cultured cells., while the responses of CrMSR gene expression in mixotrophically grown cells were different from autotrophically grown cells, High light increased the expression of CrMSRA1, CrMSRA2, CrMSRA3, CrMSRA5, CrMSRB1.2, and CrMSRB2.1, but inhibited the expression of CrMSRA4 and CrMSRB2.2 in autotrophically grown cells. The expression of CrMSRA3, CrMSRA5, and CrMSRB2.1 in mixotrophically grown cells was increased by high light but that of CrMSRA1, CrMSRA4, and CrMSRB2.2 was inhbited. The number of MSR isoform that was up-regulated by high light was greater in autotrophically grown cell than that in mixotrophically grown cells. Using the PET inhibitors (3-(3,4-dichlorophenyl)-1,1- dimethylurea (DCMU) and 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone (DBMIB)), most of the CrMSRA expression was regulated by reduced QA for autotrophically grown cells while reduced PQ was the main site for mixotrophically grown cells by high light. The expression of CrMSRB in autotrophically grown cells was mainy modulated by QA (-) or Cytb6f (-), while that was not affected by PET, except a role of Cytb6f (-) on the high light-induced CrMSRB2.2 expression. We fouind that CrMSRB gene expression in autotrophically grown cells was highly affected by PET but not for micotrophically grtown cells. The present result that H2O2 did not accumulate in autotrophically and mixotrophically grown cells suggests that H2O2 may be not involved in the regulation of high light regulation of CrMSR gene expression. The present study shows that the mRNA expression of CrMSR isoforms in Chlamydomonas was diffrerentially regulated between autotrophically and mixttrophically grown cells. The relationship between the utilization of different C source and CrMSR gene expression will be discussed.

Activity of the Ryanodine Receptor (RyR2) (aka cardiac Ca2+ release channel) plays a pivotal role in contraction of the heart. S-adenosyl-l-methionine (SAM) is a biological methyl group donor that has close structural similarity to ATP, an important physiological regulator of RyR2. This work provides evidence that SAM can act as a RyR2 regulatory ligand in a manner independent from its recognized role as a biological methyl group donor. RyR2 activation appears to arise from the direct interaction of SAM, via its adenosyl moiety, with the RyR2 adenine nucleotide binding sites. Because uncertainty remains regarding the structural motifs involved in RyR2 modulation by ATP and its metabolites, this finding has important implications for clarifying the structural basis of ATP regulation of RyR2. During the course of this project, direct measurements of single RyR2 activity revealed that SAM has distinct effects on RyR2 conductance. From the cytosolic side of the channel, SAM produced a single clearly resolved subconductance state. The effects of SAM on channel conductance were dependent on SAM concentration and membrane holding potential. A second goal of this work was to distinguish between the two possible mechanisms by which SAM could reduce RyR2 conductance: i) SAM interfering directly with ion permeation via binding within the conduction pathway (pore block), or ii) SAM binding a regulatory (or allosteric) site thereby stabilizing or inducing a reduced conductance conformation of the channel. It was determined that SAM does not directly interact with the RyR2 conduction pathway. To account for these observations an allosteric model for the effect of SAM on RyR2 conductance is proposed. According to this model, SAM binding stabilizes an inherent RyR2 subconductance conformation. The voltage dependence of the SAM related subconductance state is accounted for by direct effects of voltage on channel conformation which indirectly alter the affinity of RyR2 for SAM. Patterns in the transitions between RyR2 conductance states in the presence of SAM may provide insight into the structure-activity relationship of RyR2 which can aid in the development of therapeutic strategies targeting this channel.

Eine erhöhte Konzentration von Homocystein im Plasma (Hyperhomocysteinämie) gilt als unabhängiger Risikoindikator für neuronale und kardiovaskuläre Erkrankungen. Der Präcursor des Homocysteins, Methionin, ist eine essentielle Aminosäure, die bei Fehlernährung übermäßig verzehrt werden kann. Es wurde untersucht, ob tatsächlich durch langfristigen erhöhten Methioninverzehr via Erhöhung des Homocysteinspiegels im Plasma ein reales Gesundheitsrisiko besteht. Als Modell wurde eine Füttterungsstudie an Ratten gewählt, deren Bedingungen bezüglich Fütterungsdauer und Methioningehalt der Diät (0,4 % Methionin für 4 Wochen) an eine mögliche Fehlernährung des Menschen angepasst waren. Bei diesen Ratten war die Plasmahomocystein-Konzentration ca. 2-fach höher, als bei Ratten, die im gleichen Zeitraum eine "normale" Diät mit einem 3-4 fach niedrigeren Methioningehalt bekamen. Neben der Auswirkung der Diät auf den Homocystein-Stoffwechsel wurde geprüft, welche der in der Literatur dargestellten potentiellen Pathomechanismen für Hyperhomocysteinämie-induzierte Schäden Anwendung in diesem Modell finden. Obwohl die Konzentration von Homocystein im Plasma verändert war, wurde keine Beeinträchtigung der Gefäßfunktion gefunden. Auch die Plasmakonzentration von asymmetrischem Dimethylarginin, einem weiteren Risikoindikator für Herz-Kreislauf-Erkrankungen blieb unverändert, obwohl dieser Parameter bei Hyperhomocysteinämie oftmals erhöht ist. Eine Konzentrationsverdoppelung von Homocystein durch erhöhte alimentäre Methioninzufuhr ohne gleichzeitige Erhöhung von ADMA scheint also keine Verschlechterung der Gefäßfunktion bei Ratten zu bewirken. In diesem Punkt kann man von in anderen Modellen der Hyperhomocysteinämie gefundenen Resultaten keine Rückschlüsse auf die durch alimentäres Methionin verursachte Hyperhomocysteinämie ziehen. Andere Modelle zur Induktion einer Hyperhomocysteinämie ist Homocyst(e)in-Fütterung. In weiteren Rattenstudien wurden Effekte homocystin- und methioninreiche Diät verglichen. In diesen Studien zeigte sich, dass bei ähnlichen applizierten Dosen (tägliche Aufnahme ca. 1,0 bzw. 1,4 g/kg Körpergewicht) methionin- und homocystinreiche Diät bei Ratten zu vergleichbaren Plasma-Homocysteinspiegeln führen (methioninreich: 27,32 ± 2,80 µmol/l; homocystinreich: 40,61 ± 2,22 µmol/l). Als Vergleichsparameter zur Beurteilung pathophysiologischer Veränderungen diente zum einen der Gewebsgehalt an Homocystein, zum anderen wurden die intrazellulären Konzentrationen von S-Adenosyl-Methionin (SAM) und S-Adenosyl-Homocystein (SAH) betrachtet. Es zeigten sich besonders in Leber und Niere signifikante Unterschiede zwischen einer methioninreichen und einer homocystinreichen Diät bei Ratten. Dies ist eine mögliche Erklärung dafür, dass die bei vierwöchiger methioninreicher Diät gefundenen Ergebnisse nicht mit Literaturdaten übereinstimmen. Abschließend wurde der Frage nachgegangen, ob eine homocytinreiche Ernährung, wie in der vorherigen Studie angewandt, überhaupt möglich ist. Da zwar die Gehalte von Methionin in fast allen Lebensmitteln bekannt sind, nicht aber die von Homocystein, wurde untersucht, in welchen Konzentrationen Homocystein in Lebensmitteln enthalten ist. In Schwarzbier (0,03 mg/l), Weißbrot (0,95 mg/kg), Roquefort-Käse (0,50 mg/kg), Thunfisch (0,25 mg/kg) und Schweineleber (0,31 mg/kg) konnte Homocystein bestimmt werden. Da die Homocysteinkonzentrationen in diesen Beispiellebensmitteln mindestens um den Faktor 105 geringer waren als die Methioninkonzentrationen ist ein Einfluss von alimentärem Homocystein auf den Plasmaspiegel sehr unwahrscheinlich. Es wurde weiterhin geprüft, ob durch Darmbakterien ein Teil des alimentär aufgenommenen Methionins bereits im Dünndarm in Homocyt(e)in umgewandelt werden könnte. Dabei wurde eine vermehrte Homocysteinproduktion nach Methionin-Zugabe zu Dünndarmisolaten gefunden. Quantitativ kommt dieser Homocysteinquelle eine untergeordnete Bedeutung zu.

Methionine dependence, the inability of cells to grow when the amino acid methionine is replaced in culture medium by its metabolic precursor homocysteine, is characteristic of many cancer cell lines. Most cells proliferate normally under these conditions. The methionine dependent tumorigenic human melanoma cell line MeWo-LC1 was derived from the methionine independent non-tumorigenic line MeWo. The MeWo-LC1 cell line has been shown to have a cellular phenotype similar to that of cells from patients with the cblC inborn error of cobalamin metabolism, with decreased synthesis of cobalamin coenzymes and decreased activity of the cobalamin dependent enzymes methionine synthase and methylmalonyl-CoA mutase. Inability of cblC cells to complement the defect in cobalamin metabolism in MeWo-LC1 suggested that the defect was caused by decreased activity of the MMACHC gene product. However, no potentially disease causing mutations could be detected in the coding sequence of MMACHC in MeWo-LC1. No MMACHC expression could be detected in MeWo-LC1, and there was virtually complete methylation of a CpG island at the 5' end of the MMACHC gene in MeWo-LC1, consistent with inactivation of the gene by methylation; the CpG island was partially methylated in MeWo and only lightly methylated in control fibroblasts. Transfection of MeWo-LC1 with wild type MMACHC with a constitutive promoter resulted in correction of the defect in cobalamin metabolism and restoration of the ability of cells to grow in medium containing homocysteine. We conclude that epigenetic inactivation of the MMACHC gene is responsible for methionine dependence in MeWo-LC1.

This study characterizes B vitamin supplement use prior to and during pregnancy, changes in dietary one-carbon nutrient intakes (folate, vitamin B12, vitamin B6, choline, betaine and methionine) and most significant dietary sources. In Canadian women (Toronto, Ontario) supplemental (n=364) and dietary intakes (using a food frequency questionnaire) (n=290) were assessed during early and late pregnancy. Majority reported using a B vitamin-containing supplement prior (60%) to and during early (93%) and late (89%) pregnancy. Median supplemental intakes of folic acid, B12 and B6 were 1000 µg/d, 2.6 µg/d and 1.9 mg/d, respectively. Dietary one-carbon nutrient intakes did not change appreciably between early and late pregnancy. Most significant sources of folate and B6 were fruits and vegetables, of folic acid were cereals and grains and of B12 were dairy and egg products. Overall, this study provides novel information about one-carbon nutrient intakes in pregnancy which are crucial in maternal and child health.