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1.
BMJ Open ; 9(1): e026756, 2019 01 15.
Artigo em Inglês | MEDLINE | ID: mdl-30647050

RESUMO

INTRODUCTION: The creatine kinase circuit is central to the regulation of high-energy phosphate metabolism and the maintenance of cellular energy turnover. This circuit is fuelled by creatine, an amino acid derivative that can be obtained from a diet containing animal products, and by synthesis in the body de novo. A recent retrospective study conducted in a cohort of 287 pregnant women determined that maternal excreted levels of creatine may be associated with fetal growth. This prospective study aims to overcome some of the limitations associated with the previous study and thoroughly characterise creatine homeostasis throughout gestation in a low-risk pregnant population. METHODS AND ANALYSIS: This study is recruiting women with a singleton low-risk pregnancy who are attending Monash Health, in Melbourne, Australia. Maternal blood and urine samples, along with dietary surveys, are collected at five time points during pregnancy and then at delivery. Cord blood and placenta (including membranes and cord) are collected at birth. A biobank of tissue samples for future research is being established. Primary outcome measures will include creatine, creatine kinase and associated metabolites in antenatal bloods and urine, cord bloods and placenta, along with molecular analysis of the creatine transporter (SLC6A8) and synthesising enzymes L - arginine:glycine amidinotransferase (AGAT) and guanidinoacetate methyltransferase (GAMT) in placental tissues. Secondary outcome measures include dietary protein intake over pregnancy and any associations with maternal creatine, pregnancy events and birth outcomes. ETHICS AND DISSEMINATION: Ethical approval was granted in August 2015 from Monash Health (Ref: 14140B) and Monash University (Ref: 7785). Study outcomes will be disseminated at international conferences and published in peer-reviewed scientific journals. TRIAL REGISTRATION NUMBER: ACTRN12618001558213; Pre-results.


Assuntos
Creatina/metabolismo , Desenvolvimento Fetal , Placenta/metabolismo , Amidinotransferases/metabolismo , Austrália , Metabolismo Energético , Feminino , Guanidinoacetato N-Metiltransferase/metabolismo , Homeostase , Humanos , Proteínas do Tecido Nervoso/metabolismo , Estudos Observacionais como Assunto , Proteínas da Membrana Plasmática de Transporte de Neurotransmissores/metabolismo , Gravidez , Estudos Prospectivos , Projetos de Pesquisa
2.
Curr Protein Pept Sci ; 20(2): 184-193, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30370846

RESUMO

L-Homoarginine (hArg) ((2S)-amino-6-Carbamimidamidohexanoic acid) is a non-essential cationic amino acid that may be synthesised from the lysine catabolism or the transamination of its precursor (Arginine: Arg). These processes involve the use of the ornithine transcarbamoylase (OTC), an enzyme from the urea cycle or the arginine: glycine amidinotransferase (AGAT), an enzyme from the creatine biosynthesis pathway. These enzymes are tissue-specific, hence they synthesised L-hArg in animals and human organs such as the liver, kidneys, brains, and the small intestines. L-hArg plays some important roles in the pathophysiological conditions, endothelial functions, and the energy metabolic processes in different organs. These functions depend on the concentrations of the available LhArg in the body. These different concentrations of the L-hArg in the body are related to the different disease conditions such as the T2D mellitus, the cardiovascular and the cerebrovascular diseases, the chronic kidney diseases, the intrauterine growth restriction (IUGR) and the preeclampsia (PE) in pregnancy disorders, and even mortality. However, the applications of the L-hArg in both human and animal studies is in its juvenile stage, and the mechanism of action in this vital amino acid is not fully substantiated and requires more research attention. Hence, we review the evidence with the perspective of the LhArg usage in the monogastric and human nutrition and its related health implications.


Assuntos
Homoarginina , Amidinotransferases/metabolismo , Animais , Vias Biossintéticas/fisiologia , Doenças Cardiovasculares/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Metabolismo Energético , Feminino , Retardo do Crescimento Fetal/metabolismo , Homoarginina/biossíntese , Homoarginina/metabolismo , Homoarginina/farmacologia , Humanos , Gravidez , Insuficiência Renal Crônica/metabolismo
3.
Nutr Neurosci ; 22(5): 302-305, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-28971744

RESUMO

Arginine-glycine amidinotransferase (AGAT) deficiency is a rare inherited metabolic disorder that severely affects brain bioenergetics. Characterized by mental retardation, language impairment, and behavioral disorders, AGAT deficiency is a treatable condition, where long-term creatine supplementation usually restores brain creatine levels and improves its clinical features. In some cases of AGAT deficiency, creatine treatment might be somewhat limited due to possible shortcomings in performance and transport of creatine to the brain. Guanidinoacetic acid (GAA), a direct metabolic precursor of creatine, has recently been suggested as a possible alternative to creatine to tackle brain creatine levels in experimental medicine. AGAT patients might benefit from oral GAA due to upgraded bioavailability and convenient utilization of the compound, while possible drawbacks (e.g. brain methylation issues, neurotoxicity, and hyperhomocysteinemia) should be accounted as well.


Assuntos
Amidinotransferases/deficiência , Erros Inatos do Metabolismo dos Aminoácidos/dietoterapia , Creatina/metabolismo , Glicina/análogos & derivados , Deficiência Intelectual/dietoterapia , Distúrbios da Fala/dietoterapia , Amidinotransferases/metabolismo , Erros Inatos do Metabolismo dos Aminoácidos/metabolismo , Ensaios Clínicos como Assunto , Deficiências do Desenvolvimento/dietoterapia , Deficiências do Desenvolvimento/metabolismo , Glicina/uso terapêutico , Humanos , Deficiência Intelectual/metabolismo , Distúrbios da Fala/metabolismo , Resultado do Tratamento
4.
Acta Pharmacol Sin ; 40(4): 492-499, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-29950617

RESUMO

Myotoxicity is a significant factor contributing to the poor adherence and reduced effectiveness in the treatment of statins. Genetic variations and high drug plasma exposure are considered as critique causes for statin-induced myopathy (SIM). This study aims to explore the sequential influences of rosuvastatin (RST) pharmacokinetic and myopathy-related single-nucleotide polymorphisms (SNPs) on the plasma exposure to RST and its metabolites: rosuvastatin lactone (RSTL) and N-desmethyl rosuvastatin (DM-RST), and further on RST-induced myopathy. A total of 758 Chinese patients with coronary artery disease were enrolled and followed up SIM incidents for 2 years. The plasma concentrations of RST and its metabolites were determined through a validated ultra-performance liquid chromatography mass spectrometry method. Nine SNPs in six genes were genotyped by using the Sequenom MassArray iPlex platform. Results revealed that ABCG2 rs2231142 variations were highly associated with the plasma concentrations of RST, RSTL, and DM-RST (Padj < 0.01, FDR < 0.05). CYP2C9 rs1057910 significantly affected the DM-RST concentration (Padj < 0.01, FDR < 0.05). SLCO1B1 rs4149056 variant allele was significantly associated with high SIM risk (OR: 1.741, 95% CI: 1.180-2.568, P = 0.0052, FDR = 0.0468). Glycine amidinotransferase (GATM) rs9806699 was marginally associated with SIM incidents (OR: 0.617, 95% CI: 0.406-0.939, P = 0.0240, FDR = 0.0960). The plasma concentrations of RST and its metabolites were not significantly different between the SIM (n = 51) and control groups (n = 707) (all P > 0.05). In conclusion, SLCO1B1 and GATM genetic variants are potential biomarkers for predicting RST-induced myopathy, and their effects on SIM are unrelated to the high plasma exposure of RST and its metabolites.


Assuntos
Amidinotransferases/genética , Doença da Artéria Coronariana/tratamento farmacológico , Transportador 1 de Ânion Orgânico Específico do Fígado/genética , Doenças Musculares/induzido quimicamente , Rosuvastatina Cálcica/sangue , Amidinotransferases/sangue , Amidinotransferases/metabolismo , China , Doença da Artéria Coronariana/sangue , Doença da Artéria Coronariana/metabolismo , Variação Genética , Humanos , Transportador 1 de Ânion Orgânico Específico do Fígado/sangue , Transportador 1 de Ânion Orgânico Específico do Fígado/metabolismo , Doenças Musculares/genética , Polimorfismo de Nucleotídeo Único/genética , Rosuvastatina Cálcica/metabolismo , Rosuvastatina Cálcica/farmacocinética
5.
Amino Acids ; 50(10): 1391-1406, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-30003335

RESUMO

The L-arginine/nitric oxide synthase (NOS) pathway is considered to be altered in muscular dystrophy such as Becker muscular dystrophy (BMD). We investigated two pharmacological options aimed to increase nitric oxide (NO) synthesis in 20 male BMD patients (age range 21-44 years): (1) supplementation with L-citrulline (3 × 5 g/d), the precursor of L-arginine which is the substrate of neuronal NO synthase (nNOS); and (2) treatment with the antidiabetic drug metformin (3 × 500 mg/d) which activates nNOS in human skeletal muscle. We also investigated the combined use of L-citrulline (3 × 5 g/d) and metformin (3 × 500 mg/d). Before and after treatment, we measured in serum and urine samples the concentration of amino acids and metabolites of L-arginine-related pathways and the oxidative stress biomarker malondialdehyde (MDA). Compared to healthy subjects, BMD patients have altered NOS, arginine:glycine amidinotransferase (AGAT) and guanidinoacetate methyltransferase (GAMT) pathways. Metformin treatment resulted in concentration decrease of arginine and MDA in serum, and of homoarginine (hArg) and guanidinoacetate (GAA) in serum and urine. L-Citrulline supplementation resulted in considerable increase of the concentrations of amino acids and creatinine in the serum, and in their urinary excretion rates. Combined use of metformin and L-citrulline attenuated the effects obtained from their single administrations. Metformin, L-citrulline or their combination did not alter serum nitrite and nitrate concentrations and their urinary excretion rates. In conclusion, metformin or L-citrulline supplementation to BMD patients results in remarkable antidromic changes of the AGAT and GAMT pathways. In combination, metformin and L-citrulline at the doses used in the present study seem to abolish the biochemical effects of the single drugs in slight favor of L-citrulline.


Assuntos
Arginina/metabolismo , Citrulina/administração & dosagem , Metformina/administração & dosagem , Distrofia Muscular de Duchenne/tratamento farmacológico , Adulto , Amidinotransferases/metabolismo , Creatinina/sangue , Suplementos Nutricionais/análise , Feminino , Glicina/análogos & derivados , Glicina/sangue , Guanidinoacetato N-Metiltransferase/metabolismo , Homoarginina/sangue , Humanos , Masculino , Pessoa de Meia-Idade , Músculo Esquelético/metabolismo , Distrofia Muscular de Duchenne/enzimologia , Distrofia Muscular de Duchenne/metabolismo , Nitratos/sangue , Óxido Nítrico Sintase Tipo I/metabolismo , Resultado do Tratamento , Adulto Jovem
8.
J Am Soc Nephrol ; 29(7): 1849-1858, 2018 07.
Artigo em Inglês | MEDLINE | ID: mdl-29654216

RESUMO

Background For many patients with kidney failure, the cause and underlying defect remain unknown. Here, we describe a novel mechanism of a genetic order characterized by renal Fanconi syndrome and kidney failure.Methods We clinically and genetically characterized members of five families with autosomal dominant renal Fanconi syndrome and kidney failure. We performed genome-wide linkage analysis, sequencing, and expression studies in kidney biopsy specimens and renal cells along with knockout mouse studies and evaluations of mitochondrial morphology and function. Structural studies examined the effects of recognized mutations.Results The renal disease in these patients resulted from monoallelic mutations in the gene encoding glycine amidinotransferase (GATM), a renal proximal tubular enzyme in the creatine biosynthetic pathway that is otherwise associated with a recessive disorder of creatine deficiency. In silico analysis showed that the particular GATM mutations, identified in 28 members of the five families, create an additional interaction interface within the GATM protein and likely cause the linear aggregation of GATM observed in patient biopsy specimens and cultured proximal tubule cells. GATM aggregates-containing mitochondria were elongated and associated with increased ROS production, activation of the NLRP3 inflammasome, enhanced expression of the profibrotic cytokine IL-18, and increased cell death.Conclusions In this novel genetic disorder, fully penetrant heterozygous missense mutations in GATM trigger intramitochondrial fibrillary deposition of GATM and lead to elongated and abnormal mitochondria. We speculate that this renal proximal tubular mitochondrial pathology initiates a response from the inflammasome, with subsequent development of kidney fibrosis.


Assuntos
Amidinotransferases/genética , Síndrome de Fanconi/genética , Falência Renal Crônica/genética , Mitocôndrias/metabolismo , Mitocôndrias/patologia , Idoso , Amidinotransferases/metabolismo , Animais , Simulação por Computador , Síndrome de Fanconi/complicações , Síndrome de Fanconi/metabolismo , Síndrome de Fanconi/patologia , Feminino , Heterozigoto , Humanos , Lactente , Inflamassomos/metabolismo , Falência Renal Crônica/etiologia , Falência Renal Crônica/metabolismo , Falência Renal Crônica/patologia , Masculino , Camundongos , Camundongos Knockout , Conformação Molecular , Mutação , Mutação de Sentido Incorreto , Linhagem , Espécies Reativas de Oxigênio/metabolismo , Análise de Sequência de DNA , Adulto Jovem
9.
Toxicol Lett ; 290: 73-82, 2018 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-29574132

RESUMO

Cisplatin (CP) is a widely used drug in treatment of solid tumors. However, the use of CP was hampered by its serious side effects especially nephrotoxicity. This study aims to investigate the effect of resveratrol (RES) on CP-induced nephrotoxicity, particularly, the effect of RES on CP pharmacokinetics (PKs). Male white albino rats were divided to four group's six rats each. The first group received (1%) tween 80 in normal saline and served as control. The second group received RES (30 mg kg-1) per day for 14 consecutive day's i.p. The third and fourth groups were given a single i.p. injection of CP (6 mg kg-1) with or without pre-treatment of RES (30 mg kg-1per day for 14 consecutive days), respectively. Following administration of CP, plasma, urine and kidney platinum concentration were monitored to study PKs of CP. Five days after the CP injection, rats were killed; blood samples were collected; kidneys were dissected; and biochemical, immunohistochemical, and histological examinations were performed. Our results revealed that CP treatment significantly deteriorated kidney functions with subsequent alteration in redox balance of the kidney. On the other hand, RES successfully ameliorated CP-induced kidney injury and recovered normal kidney tissue redox status. Importantly, while RES pre-treatment did not significantly alter the plasma CP level, it dramatically decreased the urine concentration of CP and lowered its accumulation into the kidneys. Moreover, it increased CP plasma half-life (t1/2) with subsequent decrease in its elimination rate constant, indicating an important role of PKs modulation in RES protection against CP-induced renal damage. Taken together, RES may protect the kidney tissue from the deleterious effects of CP through constringe of CP renal accumulation and enhancement of CP-induced oxidative stress.


Assuntos
Antineoplásicos/toxicidade , Cisplatino/toxicidade , Rim/efeitos dos fármacos , Platina/farmacocinética , Estilbenos/farmacologia , Amidinotransferases/genética , Animais , Cisplatino/farmacocinética , Rim/metabolismo , Rim/patologia , Masculino , Óxido Nítrico Sintase Tipo III/genética , Ratos , Ratos Wistar , Resveratrol
10.
Cardiovasc Res ; 114(3): 417-430, 2018 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-29236952

RESUMO

Aims: Creatine buffers cellular adenosine triphosphate (ATP) via the creatine kinase reaction. Creatine levels are reduced in heart failure, but their contribution to pathophysiology is unclear. Arginine:glycine amidinotransferase (AGAT) in the kidney catalyses both the first step in creatine biosynthesis as well as homoarginine (HA) synthesis. AGAT-/- mice fed a creatine-free diet have a whole body creatine-deficiency. We hypothesized that AGAT-/- mice would develop cardiac dysfunction and rescue by dietary creatine would imply causality. Methods and results: Withdrawal of dietary creatine in AGAT-/- mice provided an estimate of myocardial creatine efflux of ∼2.7%/day; however, in vivo cardiac function was maintained despite low levels of myocardial creatine. Using AGAT-/- mice naïve to dietary creatine we confirmed absence of phosphocreatine in the heart, but crucially, ATP levels were unchanged. Potential compensatory adaptations were absent, AMPK was not activated and respiration in isolated mitochondria was normal. AGAT-/- mice had rescuable changes in body water and organ weights suggesting a role for creatine as a compatible osmolyte. Creatine-naïve AGAT-/- mice had haemodynamic impairment with low LV systolic pressure and reduced inotropy, lusitropy, and contractile reserve. Creatine supplementation only corrected systolic pressure despite normalization of myocardial creatine. AGAT-/- mice had low plasma HA and supplementation completely rescued all other haemodynamic parameters. Contractile dysfunction in AGAT-/- was confirmed in Langendorff perfused hearts and in creatine-replete isolated cardiomyocytes, indicating that HA is necessary for normal cardiac function. Conclusions: Our findings argue against low myocardial creatine per se as a major contributor to cardiac dysfunction. Conversely, we show that HA deficiency can impair cardiac function, which may explain why low HA is an independent risk factor for multiple cardiovascular diseases.


Assuntos
Amidinotransferases/metabolismo , Creatina/administração & dosagem , Homoarginina/administração & dosagem , Contração Miocárdica/efeitos dos fármacos , Miocárdio/enzimologia , Disfunção Ventricular Esquerda/tratamento farmacológico , Função Ventricular Esquerda/efeitos dos fármacos , Amidinotransferases/deficiência , Amidinotransferases/genética , Animais , Composição Corporal/efeitos dos fármacos , Metabolismo Energético/efeitos dos fármacos , Genótipo , Preparação de Coração Isolado , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mitocôndrias Cardíacas/efeitos dos fármacos , Mitocôndrias Cardíacas/enzimologia , Mitocôndrias Cardíacas/patologia , Fenótipo , Disfunção Ventricular Esquerda/enzimologia , Disfunção Ventricular Esquerda/genética , Disfunção Ventricular Esquerda/fisiopatologia
11.
J Biol Chem ; 292(52): 21366-21380, 2017 12 29.
Artigo em Inglês | MEDLINE | ID: mdl-29123034

RESUMO

Studies of virulence determinants in the bacterial phytopathogen Erwinia amylovora, the cause of devastating fire blight disease in apple and pear, have shown that HsvA, a putative amidinotransferase enzyme located in the Hrp pathogenicity island, is required for systemic infection in apple. However, the mechanism by which HsvA contributes to virulence is unclear. To investigate the role of HsvA in virulence, we carried out a series of biochemical and structural studies to characterize the amidinotransferase activity of HsvA. We found that HsvA displays a preference for linear aliphatic polyamines as the amidino acceptor substrate, especially for spermidine and putrescine (Km values of 33 µm and 3.9 mm, respectively). The three-dimensional structure, determined at 2.30 Å resolution using X-ray crystallography, revealed that the overall architecture of HsvA is similar to that of the human arginine-glycine amidinotransferase in the creatine biosynthesis pathway. The active site is located in the core of the protein at the base of a long, narrow substrate access channel. Specific amino acids near the entrance of the channel may serve as major determinants of the substrate specificity, including a glutamate residue at the rim of the channel entrance that appears to be positioned to interact with the distal primary amine in the putrescine substrate as well as the internal and distal amines in the spermidine substrate. These results suggest potential in vivo functions for HsvA as a virulence factor in fire blight and may also provide a basis for strategies to control fire blight by inhibiting HsvA activity.


Assuntos
Amidinotransferases/metabolismo , Erwinia amylovora/metabolismo , Amidinotransferases/fisiologia , Cristalografia por Raios X/métodos , Erwinia amylovora/patogenicidade , Ilhas Genômicas/genética , Ilhas Genômicas/fisiologia , Malus/microbiologia , Doenças das Plantas/microbiologia , Poliaminas/metabolismo , Pyrus/microbiologia , Virulência , Fatores de Virulência/metabolismo
12.
Int J Mol Sci ; 18(8)2017 Jul 31.
Artigo em Inglês | MEDLINE | ID: mdl-28758966

RESUMO

Creatine deficiency syndrome (CDS) comprises three separate enzyme deficiencies with overlapping clinical presentations: arginine:glycine amidinotransferase (GATM gene, glycine amidinotransferase), guanidinoacetate methyltransferase (GAMT gene), and creatine transporter deficiency (SLC6A8 gene, solute carrier family 6 member 8). CDS presents with developmental delays/regression, intellectual disability, speech and language impairment, autistic behaviour, epileptic seizures, treatment-refractory epilepsy, and extrapyramidal movement disorders; symptoms that are also evident in children with autism. The objective of the study was to test the hypothesis that genetic variability in creatine metabolism genes is associated with autism. We sequenced GATM, GAMT and SLC6A8 genes in 166 patients with autism (coding sequence, introns and adjacent untranslated regions). A total of 29, 16 and 25 variants were identified in each gene, respectively. Four variants were novel in GATM, and 5 in SLC6A8 (not present in the 1000 Genomes, Exome Sequencing Project (ESP) or Exome Aggregation Consortium (ExAC) databases). A single variant in each gene was identified as non-synonymous, and computationally predicted to be potentially damaging. Nine variants in GATM were shown to have a lower minor allele frequency (MAF) in the autism population than in the 1000 Genomes database, specifically in the East Asian population (Fisher's exact test). Two variants also had lower MAFs in the European population. In summary, there were no apparent associations of variants in GAMT and SLC6A8 genes with autism. The data implying there could be a lower association of some specific GATM gene variants with autism is an observation that would need to be corroborated in a larger group of autism patients, and with sub-populations of Asian ethnicities. Overall, our findings suggest that the genetic variability of creatine synthesis/transport is unlikely to play a part in the pathogenesis of autism spectrum disorder (ASD) in children.


Assuntos
Amidinotransferases/genética , Transtorno do Espectro Autista/genética , Creatinina/metabolismo , Variação Genética , Guanidinoacetato N-Metiltransferase/genética , Proteínas do Tecido Nervoso/genética , Proteínas da Membrana Plasmática de Transporte de Neurotransmissores/genética , Amidinotransferases/metabolismo , Transtorno do Espectro Autista/metabolismo , Criança , Pré-Escolar , Feminino , Guanidinoacetato N-Metiltransferase/metabolismo , Humanos , Masculino , Proteínas do Tecido Nervoso/metabolismo , Proteínas da Membrana Plasmática de Transporte de Neurotransmissores/metabolismo , Estudos Prospectivos
13.
Metab Brain Dis ; 32(6): 1951-1961, 2017 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-28808834

RESUMO

The creatine/phosphocreatine system is essential for cellular phosphate coupled energy storage and production. We investigated the utility of creatine monohydrate supplementation in two different creatine deficient knockout mouse models. Following weaning, female Arginine: Glycine Amidinotransferase (AGAT) and Guanidinoacetate: methyltransferase (GAMT) knockouts and wild type mice were studied based on their genotypes and dietary supplementation (creatine free or 2% creatine monohydrate supplemented diet) for 10 weeks, using a series of behavioral tests and biochemical analyzes. An improved Rota rod performance was observed in both AGAT (p = 0.02) and GAMT knockout mice (p < 0.001) supplemented with 2% creatine. During Morris water maze probe trial, creatine supplemented AGAT knockout mice took less time to reach virtual platform (p = 0.03) and more frequently crossed this area (p = 0.001) than mice on creatine free diet. Similar observations were recorded for GAMT knockout mice. Urinary creatinine concentrations for AGAT (p = 0.001) and GAMT (p = 0.05) knockout mice were increased following creatine supplementation. Creatine supplementation has a potential to improve neuro-muscular coordination, spatial learning in both AGAT and GAMT knockout mice. Long term Creatine supplementation results in increased urine creatinine concentrations indicating improved creatine metabolism in knockout mice.


Assuntos
Amidinotransferases/genética , Creatina/administração & dosagem , Guanidinoacetato N-Metiltransferase/genética , Aprendizagem em Labirinto/fisiologia , Atividade Motora/genética , Destreza Motora/fisiologia , Animais , Dieta , Aprendizagem em Labirinto/efeitos dos fármacos , Camundongos , Camundongos Knockout , Atividade Motora/efeitos dos fármacos , Destreza Motora/efeitos dos fármacos , Fenótipo
14.
Cell Metab ; 26(4): 660-671.e3, 2017 Oct 03.
Artigo em Inglês | MEDLINE | ID: mdl-28844881

RESUMO

Diet-induced thermogenesis is an important homeostatic mechanism that limits weight gain in response to caloric excess and contributes to the relative stability of body weight in most individuals. We previously demonstrated that creatine enhances energy expenditure through stimulation of mitochondrial ATP turnover, but the physiological role and importance of creatine energetics in adipose tissue have not been explored. Here, we have inactivated the first and rate-limiting enzyme of creatine biosynthesis, glycine amidinotransferase (GATM), selectively in fat (Adipo-Gatm KO). Adipo-Gatm KO mice are prone to diet-induced obesity due to the suppression of elevated energy expenditure that occurs in response to high-calorie feeding. This is paralleled by a blunted capacity for ß3-adrenergic activation of metabolic rate, which is rescued by dietary creatine supplementation. These results provide strong in vivo genetic support for a role of GATM and creatine metabolism in energy expenditure, diet-induced thermogenesis, and defense against diet-induced obesity.


Assuntos
Adipócitos/metabolismo , Amidinotransferases/metabolismo , Creatina/metabolismo , Dieta Hiperlipídica/efeitos adversos , Obesidade/etiologia , Obesidade/metabolismo , Termogênese , Adipócitos/patologia , Tecido Adiposo Marrom/metabolismo , Tecido Adiposo Marrom/fisiopatologia , Amidinotransferases/genética , Animais , Metabolismo Basal , Creatina/genética , Metabolismo Energético , Camundongos , Camundongos Knockout , Obesidade/genética , Obesidade/fisiopatologia
15.
Amino Acids ; 49(12): 2033-2044, 2017 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-28710553

RESUMO

L-Homoarginine (hArg) and guanidinoacetate (GAA) are produced from L-arginine (Arg) by the catalytic action of arginine:glycine amidinotransferase. Guanidinoacetate methyltransferase methylates GAA on its non-guanidine N atom to produce creatine. Arg and hArg are converted by nitric oxide synthase (NOS) to nitric oxide (NO). NO is oxidized to nitrite and nitrate which circulate in the blood and are excreted in the urine. Asymmetric dimethylarginine (ADMA), an NOS inhibitor, is widely accepted to be exclusively produced after asymmetric N G-methylation of Arg residues in proteins and their regular proteolysis. Low circulating and urinary hArg concentrations and high circulating concentrations of ADMA emerged as risk markers in the human renal and cardiovascular systems. While ADMA's distribution and metabolism are thoroughly investigated, such studies on hArg are sparse. The aim of the present pilot study was to investigate the distribution of exogenous hArg in plasma, liver, kidney, lung, and heart in a rat model of takotsubo cardiomyopathy (TTC). hArg hydrochloride solutions in physiological saline were injected intra-peritoneally at potentially pharmacological, non-toxic doses of 20, 220, or 440 mg/kg body weight. Vehicle (saline) served as control. As hArg has been reported to be a pro-oxidant, plasma and tissue malondialdehyde (MDA) was measured as a biomarker of lipid peroxidation. hArg administration resulted in dose-dependent maximum plasma hArg concentrations and distribution in all investigated organs. hArg disappeared from plasma with an elimination half-life ranging between 20 and 40 min. hArg administration resulted in relatively small changes in the plasma and tissue content of Arg, GAA, ADMA, creatinine, and of the NO metabolites nitrite and nitrate. Remarkable changes were observed for tissue GAA, notably in the kidney. Plasma and tissue MDA concentration did not change upon hArg administration, suggesting that even high-dosed hArg is not an oxidant. The lowest hArg dose of 20 mg/kg bodyweight increased 25-fold the mean hArg maximum plasma concentration. This hArg dose seems to be useful as the upper limit in forthcoming studies on the putative cardioprotective effects of hArg in our rat model of TTC.


Assuntos
Amidinotransferases/análise , Glicina/análogos & derivados , Homoarginina/farmacocinética , Rim/metabolismo , Cardiomiopatia de Takotsubo/metabolismo , Animais , Arginina/análogos & derivados , Arginina/sangue , Arginina/metabolismo , Glicina/sangue , Glicina/metabolismo , Guanidinoacetato N-Metiltransferase , Meia-Vida , Homoarginina/administração & dosagem , Homoarginina/sangue , Homoarginina/metabolismo , Humanos , Rim/efeitos dos fármacos , Peroxidação de Lipídeos/efeitos dos fármacos , Malondialdeído/sangue , Modelos Animais , Óxido Nítrico Sintase , Projetos Piloto , Ratos , Ratos Sprague-Dawley , Distribuição Tecidual
16.
Chemistry ; 23(45): 10714-10724, 2017 Aug 10.
Artigo em Inglês | MEDLINE | ID: mdl-28488371

RESUMO

Peptides are biologically occurring oligomers of amino acids linked by amide bonds and are indispensable for all living organisms. Many bioactive peptides are used as antibiotics, antivirus agents, insecticides, pheromones, and food preservatives. Nature employs several different strategies to form amide bonds. ATP-grasp enzymes that catalyze amide bond formation (ATP-dependent carboxylate-amine ligases) utilize a strategy of activating carboxylic acid as an acylphosphate intermediate to form amide bonds and are involved in many different biological processes in both primary and secondary metabolisms. The recent discovery of several new ATP-dependent carboxylate-amine ligases has expanded the diversity of this group of enzymes and showed their usefulness for generating oligopeptides. In this review, an overview of findings on amide bond formation catalyzed by ATP-grasp enzymes in the past decade is presented.


Assuntos
Carboxipeptidases/metabolismo , Oligopeptídeos/biossíntese , Trifosfato de Adenosina/metabolismo , Amidinotransferases/metabolismo , Cobalto/química , Ciclização , Combinação de Medicamentos , Metionina/biossíntese , Metionina/química , Oligopeptídeos/química , Peptídeo Sintases/metabolismo
17.
Placenta ; 52: 86-93, 2017 04.
Artigo em Inglês | MEDLINE | ID: mdl-28454702

RESUMO

INTRODUCTION: Creatine is an amino acid derivative that is involved in preserving ATP homeostasis. Previous studies suggest an important role for the creatine kinase circuit for placental ATP turnover. Creatine is obtained from both the diet and endogenous synthesis, usually along the renal-hepatic axis. However, some tissues with a high-energy demand have an inherent capacity to synthesise creatine. In this study, we determined if the term human placenta has the enzymatic machinary to synthesise creatine. METHODS: Eleven placentae were collected following elective term caesarean section. Samples from the 4 quadrants of each placenta were either fixed in formalin or frozen. qPCR was used to determine the mRNA expression of the creatine synthesising enzymes arginine:glycine amidinotransferase (AGAT) and guanidinoacetate methyltransferase (GAMT), and the creatine transporter (SLC6A8). Protein expression of AGAT and GAMT was quantified by Western blot, and observations of cell localisation of AGAT, GAMT and SLC6A8 made with immunohistochemistry. Synthesis of guanidinoacetate (GAA; creatine precursor) and creatine in placental homogenates was determined via GC-MS and HPLC, respectively. RESULTS: AGAT, GAMT and SLC6A8 mRNA and protein were detected in the human placenta. AGAT staining was identified in stromal and endothelial cells of the fetal capillaries. GAMT and SLC6A8 staining was localised to the syncytiotrophoblast of the fetal villi. Ex vivo, tissue homogenates produce both GAA (4.6 nmol mg protein-1h-1) and creatine (52.8 nmol mg protein-1h-1). DISCUSSION: The term human placenta has the capacity to synthesise creatine. These data present a new understanding of placental energy metabolism.


Assuntos
Amidinotransferases/metabolismo , Creatina/metabolismo , Guanidinoacetato N-Metiltransferase/metabolismo , Proteínas de Membrana Transportadoras/metabolismo , Placenta/metabolismo , Transporte Biológico , Creatina/biossíntese , Células Endoteliais/metabolismo , Metabolismo Energético/fisiologia , Feminino , Humanos , Gravidez , Células Estromais/metabolismo
18.
Proc Natl Acad Sci U S A ; 114(7): E1273-E1281, 2017 02 14.
Artigo em Inglês | MEDLINE | ID: mdl-28137860

RESUMO

Creatine, a nitrogenous organic acid, replenishes cytoplasmic ATP at the expense of mitochondrial ATP via the phosphocreatine shuttle. Creatine levels are maintained by diet and endogenous synthesis from arginine and glycine. Glycine amidinotransferase (GATM) catalyzes the rate-limiting step of creatine biosynthesis: the transfer of an amidino group from arginine to glycine to form ornithine and guanidinoacetate. We screened 36,530 third-generation germline mutant mice derived from N-ethyl-N-nitrosourea-mutagenized grandsires for intestinal homeostasis abnormalities after oral administration of dextran sodium sulfate (DSS). Among 27 colitis susceptibility phenotypes identified and mapped, one was strongly correlated with a missense mutation in Gatm in a recessive model of inheritance, and causation was confirmed by CRISPR/Cas9 gene targeting. Supplementation of homozygous Gatm mutants with exogenous creatine ameliorated the colitis phenotype. CRISPR/Cas9-targeted (Gatmc/c ) mice displayed a normal peripheral immune response and immune cell homeostasis. However, the intestinal epithelium of the Gatmc/c mice displayed increased cell death and decreased proliferation during DSS treatment. In addition, Gatmc/c colonocytes showed increased metabolic stress in response to DSS with higher levels of phospho-AMPK and lower levels of phosphorylation of mammalian target of rapamycin (phospho-mTOR). These findings establish an in vivo requirement for rapid replenishment of cytoplasmic ATP within colonic epithelial cells in the maintenance of the mucosal barrier after injury.


Assuntos
Colite/prevenção & controle , Creatina/farmacologia , Homeostase/efeitos dos fármacos , Intestinos/efeitos dos fármacos , Amidinotransferases/genética , Amidinotransferases/metabolismo , Animais , Sistemas CRISPR-Cas , Colite/genética , Colite/metabolismo , Colo/efeitos dos fármacos , Colo/metabolismo , Creatina/metabolismo , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Feminino , Mucosa Intestinal/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Mutação de Sentido Incorreto , Substâncias Protetoras/metabolismo , Substâncias Protetoras/farmacologia
19.
Orphanet J Rare Dis ; 12(1): 21, 2017 02 02.
Artigo em Inglês | MEDLINE | ID: mdl-28148286

RESUMO

BACKGROUND: Arginine:glycine amidinotransferase deficiency (AGAT-d) is a very rare inborn error of creatine synthesis mainly characterized by absence of brain Creatine (Cr) peak, intellectual disability, severe language impairment and behavioural disorder and susceptible to supplementary Cr treatment per os. Serial examinations by magnetic resonance spectroscopy are required to evaluate Cr recovery in brain during treatment of high doses of Cr per os, which have been proved beneficial and effective in treating main clinical symptoms. A long term study with detailed reports on clinical, neurochemical and neuropsychological outcomes of the first Italian patients affected by AGAT-d here reported can represent a landmark in management of this disorder thus enhancing medical knowledge and clinical practice. RESULTS: We have evaluated the long term effects of Cr supplementation management in four Italian patients affected by AGAT-d, correlating specific treatments with serial clinical, biochemical and magnetic resonance spectroscopy examinations as well as the neuropsychological outcome by standardized developmental scales. Consecutive MRS examinations have confirmed that Cr depletion in AGAT-d patients is reversible under Cr supplementation. Cr treatment is considered safe and well tolerated but side effects, including weight gain and kidney stones, have been reported. CONCLUSIONS: Early treatment prevents adverse developmental outcome, while patients diagnosed and treated at an older age showed partial but significant cognitive recovery with clear improvements in adaptive functioning.


Assuntos
Amidinotransferases/deficiência , Erros Inatos do Metabolismo dos Aminoácidos/tratamento farmacológico , Creatina/uso terapêutico , Família , Deficiência Intelectual/tratamento farmacológico , Distúrbios da Fala/tratamento farmacológico , Adolescente , Criança , Pré-Escolar , Creatina/administração & dosagem , Deficiências do Desenvolvimento/tratamento farmacológico , Suplementos Nutricionais , Esquema de Medicação , Feminino , Humanos , Lactente , Recém-Nascido , Itália , Masculino , Adulto Jovem
20.
Genet Med ; 19(2): 256-263, 2017 02.
Artigo em Inglês | MEDLINE | ID: mdl-28055022

RESUMO

Disclaimer: These ACMG Standards and Guidelines are intended as an educational resource for clinical laboratory geneticists to help them provide quality clinical laboratory genetic services. Adherence to these standards and guidelines is voluntary and does not necessarily assure a successful medical outcome. These Standards and Guidelines should not be considered inclusive of all proper procedures and tests or exclusive of others that are reasonably directed to obtaining the same results. In determining the propriety of any specific procedure or test, clinical laboratory geneticists should apply their professional judgment to the specific circumstances presented by the patient or specimen. Clinical laboratory geneticists are encouraged to document in the patient's record the rationale for the use of a particular procedure or test, whether or not it is in conformance with these Standards and Guidelines. They also are advised to take notice of the date any particular guideline was adopted, and to consider other relevant medical and scientific information that becomes available after that date. It also would be prudent to consider whether intellectual property interests may restrict the performance of certain tests and other procedures.Cerebral creatine deficiency syndromes are neurometabolic conditions characterized by intellectual disability, seizures, speech delay, and behavioral abnormalities. Several laboratory methods are available for preliminary and confirmatory diagnosis of these conditions, including measurement of creatine and related metabolites in biofluids using liquid chromatography-tandem mass spectrometry or gas chromatography-mass spectrometry, enzyme activity assays in cultured cells, and DNA sequence analysis. These guidelines are intended to standardize these procedures to help optimize the diagnosis of creatine deficiency syndromes. While biochemical methods are emphasized, considerations for confirmatory molecular testing are also discussed, along with variables that influence test results and interpretation.Genet Med 19 2, 256-263.


Assuntos
Amidinotransferases/deficiência , Erros Inatos do Metabolismo dos Aminoácidos/genética , Encefalopatias Metabólicas Congênitas/genética , Creatina/deficiência , Creatina/metabolismo , Guanidinoacetato N-Metiltransferase/deficiência , Deficiência Intelectual/genética , Transtornos do Desenvolvimento da Linguagem/genética , Retardo Mental Ligado ao Cromossomo X/genética , Transtornos dos Movimentos/congênito , Proteínas da Membrana Plasmática de Transporte de Neurotransmissores/deficiência , Proteínas Repressoras/genética , Distúrbios da Fala/genética , Amidinotransferases/sangue , Amidinotransferases/líquido cefalorraquidiano , Amidinotransferases/genética , Amidinotransferases/urina , Erros Inatos do Metabolismo dos Aminoácidos/sangue , Erros Inatos do Metabolismo dos Aminoácidos/líquido cefalorraquidiano , Erros Inatos do Metabolismo dos Aminoácidos/urina , Encefalopatias Metabólicas Congênitas/sangue , Encefalopatias Metabólicas Congênitas/líquido cefalorraquidiano , Encefalopatias Metabólicas Congênitas/urina , Técnicas de Laboratório Clínico/métodos , Creatina/sangue , Creatina/líquido cefalorraquidiano , Creatina/genética , Creatina/urina , Deficiências do Desenvolvimento/sangue , Deficiências do Desenvolvimento/líquido cefalorraquidiano , Deficiências do Desenvolvimento/genética , Deficiências do Desenvolvimento/urina , Testes Genéticos/normas , Genética Médica/normas , Genômica , Guanidinoacetato N-Metiltransferase/sangue , Guanidinoacetato N-Metiltransferase/líquido cefalorraquidiano , Guanidinoacetato N-Metiltransferase/genética , Guanidinoacetato N-Metiltransferase/urina , Guias como Assunto , Humanos , Deficiência Intelectual/sangue , Deficiência Intelectual/líquido cefalorraquidiano , Deficiência Intelectual/urina , Transtornos do Desenvolvimento da Linguagem/sangue , Transtornos do Desenvolvimento da Linguagem/líquido cefalorraquidiano , Transtornos do Desenvolvimento da Linguagem/urina , Retardo Mental Ligado ao Cromossomo X/sangue , Retardo Mental Ligado ao Cromossomo X/líquido cefalorraquidiano , Retardo Mental Ligado ao Cromossomo X/urina , Transtornos dos Movimentos/sangue , Transtornos dos Movimentos/líquido cefalorraquidiano , Transtornos dos Movimentos/genética , Transtornos dos Movimentos/urina , Proteínas da Membrana Plasmática de Transporte de Neurotransmissores/sangue , Proteínas da Membrana Plasmática de Transporte de Neurotransmissores/líquido cefalorraquidiano , Proteínas da Membrana Plasmática de Transporte de Neurotransmissores/genética , Proteínas da Membrana Plasmática de Transporte de Neurotransmissores/urina , Proteínas Repressoras/sangue , Proteínas Repressoras/líquido cefalorraquidiano , Proteínas Repressoras/urina , Distúrbios da Fala/sangue , Distúrbios da Fala/líquido cefalorraquidiano
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