RESUMEN
The placenta requires high levels of adenosine triphosphate to maintain a metabolically active state throughout gestation. The creatine-creatine kinase-phosphocreatine system is known to buffer adenosine triphosphate levels; however, the role(s) creatine-creatine kinase-phosphocreatine system plays in uterine and placental metabolism throughout gestation is poorly understood. In this study, Suffolk ewes were ovariohysterectomized on Days 30, 50, 70, 90, 110 and 125 of gestation (n = 3-5 ewes/per day, except n = 2 on Day 50) and uterine and placental tissues subjected to analyses to measure metabolites, mRNAs, and proteins related to the creatine-creatine kinase-phosphocreatine system. Day of gestation affected concentrations and total amounts of guanidinoacetate and creatine in maternal plasma, amniotic fluid and allantoic fluid (P < 0.05). Expression of mRNAs for arginine:glycine amidinotransferase, guanidinoacetate methyltransferase, creatine kinase B, and solute carrier 16A12 in endometria and for arginine:glycine amidinotransferase and creatine kinase B in placentomes changed significantly across days of gestation (P < 0.05). The arginine:glycine amidinotransferase protein was more abundant in uterine luminal epithelium on Days 90 and 125 compared to Days 30 and 50 (P < 0.01). The chorionic epithelium of placentomes expressed guanidinoacetate methyltransferase and solute carrier 6A13 throughout gestation. Creatine transporter (solute carrier 6A8) was expressed by the uterine luminal epithelium and trophectoderm of placentomes throughout gestation. Creatine kinase (creatine kinase B and CKMT1) proteins were localized primarily to the uterine luminal epithelium and to the placental chorionic epithelium of placentomes throughout gestation. Collectively, these results demonstrate cell-specific and temporal regulation of components of the creatine-creatine kinase-phosphocreatine system that likely influence energy homeostasis for fetal-placental development.
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Creatina , Placenta , Embarazo , Femenino , Animales , Ovinos , Placenta/metabolismo , Creatina/metabolismo , Guanidinoacetato N-Metiltransferasa/metabolismo , Fosfocreatina/metabolismo , Creatina Quinasa/metabolismo , Adenosina Trifosfato/metabolismo , ArgininaRESUMEN
Guanidinoacetate methyltransferase deficiency (GAMTD) is a treatable neurodevelopmental disorder with normal or nonspecific imaging findings. Here, we reported a 14-month-old girl with GAMTD and novel findings on brain magnetic resonance imaging (MRI).A 14-||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||month-old female patient was referred to Myelin Disorders Clinic due to onset of seizures and developmental regression following routine vaccination at 4 months of age. Brain MRI, prior to initiation of treatment, showed high signal intensity in T2-weighted imaging in bilateral thalami, globus pallidus, subthalamic nuclei, substantia nigra, dentate nuclei, central tegmental tracts in the brainstem, and posterior periventricular white matter which was masquerading for mitochondrial leukodystrophy. Basic metabolic tests were normal except for low urine creatinine; however, exome sequencing identified a homozygous frameshift deletion variant [NM_000156: c.491del; (p.Gly164AlafsTer14)] in the GAMT. Biallelic pathogenic or likely pathogenic variants cause GAMTD. We confirmed the homozygous state for this variant in the proband, as well as the heterozygote state in the parents by Sanger sequencing.MRI features in GAMTD can mimic mitochondrial leukodystrophy. Pediatric neurologists should be aware of variable MRI findings in GAMTD since they would be misleading to other diagnoses.
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Trastornos del Desarrollo del Lenguaje , Trastornos del Movimiento , Niño , Humanos , Femenino , Lactante , Irán , Trastornos del Desarrollo del Lenguaje/genética , Trastornos del Desarrollo del Lenguaje/diagnóstico , Trastornos del Desarrollo del Lenguaje/metabolismo , Guanidinoacetato N-Metiltransferasa/metabolismo , NeuroimagenRESUMEN
BACKGROUND: Guanidinoacetate (GAA) induces epileptogenesis and neurotoxicity in the brain. As epileptic animal models have been reported to show elevated cerebral GAA levels, the processing mechanism of GAA in the brain is important for maintaining brain homeostasis. We have revealed that GAA in the cerebrospinal fluid (CSF) is removed by incorporation into the choroid plexus epithelial cells (CPxEpic), which form the blood-CSF barrier (BCSFB). However, the processing mechanism of GAA incorporated into CPxEpic remains unknown. We have reported that monocarboxylate transporter 12 (MCT12) functions as an efflux transporter of GAA and creatine, a metabolite of GAA, in the kidneys and liver. Therefore, we aimed to clarify the role of MCT12 in GAA dynamics in CPxEpic. METHODS: Protein expression and localization in CPxEpic were evaluated using immunohistochemistry. Metabolic analysis was performed using high-performance liquid chromatography (HPLC) 24 h after the addition of [14C]GAA to TR-CSFB3 cells, which are conditionally immortalized rat CPxEpic. The efflux transport of [14C]creatine was evaluated in TR-CSFB3 cells after transfection with MCT12 small interfering RNA (siRNA). The CSF-to-brain parenchyma transfer of creatine was measured after intracerebroventricular injection in rats. RESULTS: Immunohistochemical staining revealed that MCT12-derived signals merged with those of the marker protein at the apical membrane of CPxEpic, suggesting that MCT12 is localized on the apical membrane of CPxEpic. The expression levels of guanidinoacetate N-methyltransferase (GAMT), which catalyzes the conversion of GAA to creatine, in TR-CSFB3 cells was also indicated, and GAA was considered to be metabolized to creatine after influx transport into CPxEpic, after which creatine was released into the CSF. Creatine release from TR-CSFB3 cells decreased following MCT12 knockdown. The contribution ratio of MCT12 to the release of creatine was more than 50%. The clearance of CSF-to-brain parenchyma transfer of creatine was 4.65 µL/(min·g brain), suggesting that biosynthesized creatine in CPxEpic is released into the CSF and supplied to the brain parenchyma. CONCLUSIONS: In CPxEpic, GAA is metabolized to creatine via GAMT. Biosynthesized creatine is then released into the CSF via MCT12 and supplied to the brain parenchyma.
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Creatina , Guanidinoacetato N-Metiltransferasa , Animales , Plexo Coroideo/metabolismo , Creatina/química , Creatina/metabolismo , Células Epiteliales/metabolismo , Glicina/análogos & derivados , Guanidinoacetato N-Metiltransferasa/metabolismo , RatasRESUMEN
Early-stage gastric cancer (GC) is asymptomatic. How to diagnose the early-stage GC is challenging. The sensitivity and specificity of diagnosing signatures for early-stage patients are still poor. Elastic-net-based analysis was used to identify potential diagnostic signatures of early-stage GC. The expression level of candidate gene was evaluated by immunohistochemistry staining. The potential function of candidate gene was verified by overexpressing in vitro. Consensus genes (including GAMT) were identified using the different strengths of the penalty. Surprisingly, GAMT was still identified even if some multicollinear variables were deleted directly. IHC staining showed that there are no GAMT-positive signals in the cell nuclei of all tumor tissues, while GAMT does express in nuclei of adjacent normal tissue. There are 16.33% positive cell nuclei in paracancerous tissues. In addition, the number of larger-area colonies of overexpression-GAMT group, empty-vector group, and AGS group is 70±29.21, 151.33±15.95, and 111.67±22.03, respectively. Number of larger colonies in group with overexpression of GAMT is significantly less than control groups. Elastic-net-penalty-based workflow is a effective tool to identify diagnostic biomarker for early-stage solid tumor. GAMT has strong potential to be the diagnostic biomarker for the early-stage GC.
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Biomarcadores de Tumor/metabolismo , Elasticidad , Guanidinoacetato N-Metiltransferasa/metabolismo , Neoplasias Gástricas/diagnóstico , Línea Celular Tumoral , Proliferación Celular , Análisis por Conglomerados , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Humanos , Estadificación de Neoplasias , Modelos de Riesgos Proporcionales , Reproducibilidad de los Resultados , Neoplasias Gástricas/genética , Neoplasias Gástricas/patologíaRESUMEN
The enzymes glycine amidinotransferase, mitochondrial (GATM also known as AGAT) and guanidinoacetate N-methyltransferase (GAMT) function together to synthesize creatine from arginine, glycine, and S-Adenosyl methionine. Deficiency in either enzyme or the creatine transporter, CT1, results in a devastating neurological disorder, Cerebral Creatine Deficiency Syndrome (CCDS). To better understand the pathophysiology of CCDS, we mapped the distribution of GATM and GAMT at single cell resolution, leveraging RNA sequencing analysis combined with in vivo immunofluorescence (IF). Using the mouse as a model system, we find that GATM and GAMT are coexpressed in several tissues with distinct and overlapping cellular sources, implicating local synthesis as an important mechanism of creatine metabolism in numerous organs. Extending previous findings at the RNA level, our analysis demonstrates that oligodendrocytes express the highest level of Gatm and Gamt of any cell type in the body. We confirm this finding in the mouse brain by IF, where GATM localizes to the mitochondria of oligodendrocytes, whereas both oligodendrocytes and cerebral cortical neurons express GAMT. Interestingly, the latter is devoid of GATM. Single nucleus assay for transposase-accessible chromatin sequencing (snATAC-seq) analysis of 4 brain regions highlights a similar primacy of oligodendrocytes in the expression of GATM and GAMT in the human central nervous system. Importantly, an active putative regulatory element within intron 2 of human GATM is detected in oligodendrocytes but not neurons.
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Amidinotransferasas/metabolismo , Encéfalo/metabolismo , Creatina/metabolismo , Guanidinoacetato N-Metiltransferasa/metabolismo , Oligodendroglía/metabolismo , Animales , Ratones , Mitocondrias/metabolismo , Neuronas/metabolismoAsunto(s)
Amidinotransferasas/deficiencia , Errores Innatos del Metabolismo de los Aminoácidos/metabolismo , Creatina/deficiencia , Glicina/análogos & derivados , Guanidinoacetato N-Metiltransferasa/deficiencia , Discapacidad Intelectual/metabolismo , Trastornos del Desarrollo del Lenguaje/metabolismo , Trastornos del Movimiento/congénito , Enfermedades Musculares/tratamiento farmacológico , Trastornos del Habla/metabolismo , Amidinotransferasas/metabolismo , Animales , Creatina/metabolismo , Creatina/uso terapéutico , Discapacidades del Desarrollo/metabolismo , Glicina/deficiencia , Glicina/metabolismo , Glicina/uso terapéutico , Guanidinoacetato N-Metiltransferasa/metabolismo , Humanos , Inhibidores de Hidroximetilglutaril-CoA Reductasas/toxicidad , Ratones , Trastornos del Movimiento/metabolismo , Enfermedades Musculares/inducido químicamente , Enfermedades Musculares/metabolismo , FosforilaciónRESUMEN
Statins prevent cardiovascular diseases, yet their use is limited by the muscle disturbances they cause. Rarely, statin-induced myopathy is autoimmune, but more commonly it is due to direct muscle toxicity. Available evidence suggests that statin-induced creatine deficiency might be a major cause of this toxicity, and that creatine supplementation prevents it. Statins inhibit guanidinoacetate methyl transferase (GAMT), the last enzyme in the synthesis of creatine; thus, they decrease its intracellular content. Such decreased content could cause mitochondrial impairment, since creatine is the final acceptor of the phosphate group of adenosine triphosphate (ATP) at the end of mitochondrial oxidative phosphorylation. Decreased cellular synthesis of ATP would follow. Accordingly, ATP synthesis is decreased in statin-treated cells. In vitro, creatine supplementation prevents the opening of the mitochondrial permeability transition pore that is caused by statins. Clinically, creatine administration prevents statin myopathy in statin-intolerant patients. Additional research is warranted to hopefully confirm these findings. However, creatine is widely used by athletes with no adverse events, and has demonstrated to be safe even in double-blind, placebo-controlled trials of elderly individuals. Thus, it should be trialed, under medical supervision, in patients who cannot assume statin due to the occurrence of muscular symptoms.
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Creatina/administración & dosificación , Inhibidores de Hidroximetilglutaril-CoA Reductasas/efectos adversos , Enfermedades Musculares/metabolismo , Atletas , Ensayos Clínicos como Asunto , Creatina/farmacología , Regulación Enzimológica de la Expresión Génica/efectos de los fármacos , Guanidinoacetato N-Metiltransferasa/metabolismo , Humanos , Enfermedades Musculares/inducido químicamente , Enfermedades Musculares/tratamiento farmacológico , Fosforilación Oxidativa/efectos de los fármacosRESUMEN
Creatine is a metabolite involved in cellular energy homeostasis. In this study, we examined placental creatine content, and expression of the enzymes required for creatine synthesis, transport and the creatine kinase reaction, in pregnancies complicated by low birthweight. We studied first trimester chorionic villus biopsies (CVBs) of small for gestational age (SGA) and appropriately grown infants (AGA), along with third trimester placental samples from fetal growth restricted (FGR) and healthy gestation-matched controls. Placental creatine and creatine precursor (guanidinoacetate-GAA) levels were measured. Maternal and cord serum from control and FGR pregnancies were also analyzed for creatine concentration. mRNA expression of the creatine transporter (SLC6A8); synthesizing enzymes arginine:glycine aminotransferase (GATM) and guanidinoacetate methyltransferase (GAMT); mitochondrial (mtCK) and cytosolic (BBCK) creatine kinases; and amino acid transporters (SLC7A1 & SLC7A2) was assessed in both CVBs and placental samples. Protein levels of AGAT (arginine:glycine aminotransferase), GAMT, mtCK and BBCK were also measured in placental samples. Key findings; total creatine content of the third trimester FGR placentae was 43% higher than controls. The increased creatine content of placental tissue was not reflected in maternal or fetal serum from FGR pregnancies. Tissue concentrations of GAA were lower in the third trimester FGR placentae compared to controls, with lower GATM and GAMT mRNA expression also observed. No differences in the mRNA expression of GATM, GAMT or SLC6A8 were observed between CVBs from SGA and AGA pregnancies. These results suggest placental creatine metabolism in FGR pregnancies is altered in late gestation. The relevance of these changes on placental bioenergetics should be the focus of future investigations.
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Creatina/metabolismo , Guanidinoacetato N-Metiltransferasa/metabolismo , Placenta/metabolismo , Placenta/fisiopatología , Adulto , Femenino , Desarrollo Fetal/genética , Desarrollo Fetal/fisiología , Guanidinoacetato N-Metiltransferasa/genética , Humanos , Embarazo , Primer Trimestre del Embarazo/metabolismo , Tercer Trimestre del Embarazo/metabolismo , ARN Mensajero/metabolismoRESUMEN
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.
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Creatina/metabolismo , Desarrollo Fetal , Placenta/metabolismo , Amidinotransferasas/metabolismo , Australia , Metabolismo Energético , Femenino , Guanidinoacetato N-Metiltransferasa/metabolismo , Homeostasis , Humanos , Proteínas del Tejido Nervioso/metabolismo , Estudios Observacionales como Asunto , Proteínas de Transporte de Neurotransmisores en la Membrana Plasmática/metabolismo , Embarazo , Estudios Prospectivos , Proyectos de InvestigaciónRESUMEN
PURPOSE: To obtain high-resolution Cr and PCr maps of mouse skeletal muscle using a polynomial and Lorentzian line-shape fitting (PLOF) CEST method. METHODS: Wild-type mice and guanidinoacetate N-methyltransferase-deficient (GAMT-/-) mice that have low Cr and PCr concentrations in muscle were used to assign the Cr and PCr peaks in the Z-spectrum at 11.7 T. A PLOF method was proposed to simultaneously extract and quantify the Cr and PCr by assuming a polynomial function for the background and 2 Lorentzian functions for the CEST peaks at 1.95 ppm and 2.5 ppm. RESULTS: The Z-spectra of phantoms revealed that PCr has 2 CEST peaks (2 ppm and 2.5 ppm), whereas Cr only showed 1 peak at 2 ppm. Comparison of the Z-spectra of wild-type and GAMT-/- mice indicated that, contrary to brain, there was no visible protein guanidinium peak in the skeletal-muscle Z-spectrum, which allowed us to extract clean PCr and Cr CEST signals. High-resolution PCr and Cr concentration maps of mouse skeletal muscle were obtained by the PLOF CEST method after calibration with in vivo MRS. CONCLUSIONS: The PLOF method provides an efficient way to map Cr and PCr concentrations simultaneously in the skeletal muscle at high MRI field.
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Creatina/análisis , Espectroscopía de Resonancia Magnética/métodos , Músculo Esquelético/metabolismo , Fosfocreatina/análisis , Algoritmos , Animales , Medios de Contraste , Femenino , Guanidinoacetato N-Metiltransferasa/genética , Guanidinoacetato N-Metiltransferasa/metabolismo , Ratones , Ratones Endogámicos BALB C , Modelos Teóricos , Fantasmas de Imagen , Fosfocreatina/análogos & derivados , Fosfocreatina/sangreRESUMEN
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.
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Arginina/metabolismo , Citrulina/administración & dosificación , Metformina/administración & dosificación , Distrofia Muscular de Duchenne/tratamiento farmacológico , Adulto , Amidinotransferasas/metabolismo , Creatinina/sangre , Suplementos Dietéticos/análisis , Femenino , Glicina/análogos & derivados , Glicina/sangre , Guanidinoacetato N-Metiltransferasa/metabolismo , Homoarginina/sangre , Humanos , Masculino , Persona de Mediana Edad , Músculo Esquelético/metabolismo , Distrofia Muscular de Duchenne/enzimología , Distrofia Muscular de Duchenne/metabolismo , Nitratos/sangre , Óxido Nítrico Sintasa de Tipo I/metabolismo , Resultado del Tratamiento , Adulto JovenRESUMEN
Chronic intermittent hypoxia (ChIH) is a dominant feature of obstructive sleep apnoea (OSA) and is associated to metabolic alterations and oxidative stress (OS). Although management of OSA is well established, the research of new biomarkers that are independent of confounding factors remains necessary to improve the early detection of comorbidity and therapeutic follow-up. In this study, the urinary metabonomic profile associated to intermittent hypoxia was evaluated in a mouse model. When exposed to intermittent hypoxia, animals showed a significant alteration in energy metabolism towards anaerobic pathways and signs of OS imbalance. A compensatory response was observed over time. Our data also indicates an excess production of vitamin B3, liver function modulations and a stimulation of creatine synthesis which could be used to evaluate the ChIH repercussions. As well, TMAO and allantoin could constitute interesting biomarker candidates, respectively in the context of cardiovascular risk and OS associated to OSA.
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Hipoxia/orina , Metabolómica/métodos , Estrés Oxidativo/fisiología , Análisis de Varianza , Animales , Creatina/metabolismo , Modelos Animales de Enfermedad , Prueba de Tolerancia a la Glucosa , Guanidinoacetato N-Metiltransferasa/metabolismo , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Espectroscopía de Resonancia Magnética/métodos , Ratones , Factores de Tiempo , Tritio/metabolismoRESUMEN
Creatine is synthesized by S-adenosylmethionine:guanidinoacetate N-methyltransferase (GAMT), and the creatine/phosphocreatine shuttle system mediated by creatine kinase (CK) is essential for storage and regeneration of high-energy phosphates in cells. Although the importance of this system in brain development is evidenced by the hereditary nature of creatine deficiency syndrome, the spatiotemporal cellular expression patterns of GAMT in developing brain remain unknown. Here we show that two waves of high GAMT expression occur in developing mouse brain. The first involves high expression in mitotic cells in the ventricular zone of the brain wall and the external granular layer of the cerebellum at the embryonic and neonatal stages. The second was initiated by striking up-regulation of GAMT in oligodendrocytes during the second and third postnatal weeks (i.e., the active myelination stage), which continued to adulthood. Distinct temporal patterns were also evident in other cell types. GAMT was highly expressed in perivascular pericytes and smooth muscle cells after birth, but not in adults. In neurons, GAMT levels were low to moderate in neuroblasts residing in the ventricular zone, increased during the second postnatal week when active dendritogenesis and synaptogenesis occur, and decreased to very low levels thereafter. Moderate levels were observed in astrocytes throughout development. The highly regulated, cell type-dependent expression of GAMT suggests that local creatine biosynthesis plays critical roles in certain phases of neural development. In accordance with this idea, we observed increased CK expression in differentiating neurons; this would increase creatine/phosphocreatine shuttle system activity, which might reflect increased energy demand.
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Creatina/metabolismo , Guanidinoacetato N-Metiltransferasa/metabolismo , Neuronas/metabolismo , S-Adenosilmetionina/metabolismo , Animales , Encéfalo/crecimiento & desarrollo , Glicina/metabolismo , Metiltransferasas/metabolismo , Ratones Endogámicos C57BL , Fosfocreatina/metabolismoRESUMEN
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.
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Amidinotransferasas/genética , Trastorno del Espectro Autista/genética , Creatinina/metabolismo , Variación Genética , Guanidinoacetato N-Metiltransferasa/genética , Proteínas del Tejido Nervioso/genética , Proteínas de Transporte de Neurotransmisores en la Membrana Plasmática/genética , Amidinotransferasas/metabolismo , Trastorno del Espectro Autista/metabolismo , Niño , Preescolar , Femenino , Guanidinoacetato N-Metiltransferasa/metabolismo , Humanos , Masculino , Proteínas del Tejido Nervioso/metabolismo , Proteínas de Transporte de Neurotransmisores en la Membrana Plasmática/metabolismo , Estudios ProspectivosRESUMEN
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.
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Amidinotransferasas/metabolismo , Creatina/metabolismo , Guanidinoacetato N-Metiltransferasa/metabolismo , Proteínas de Transporte de Membrana/metabolismo , Placenta/metabolismo , Transporte Biológico , Creatina/biosíntesis , Células Endoteliales/metabolismo , Metabolismo Energético/fisiología , Femenino , Humanos , Embarazo , Células del Estroma/metabolismoRESUMEN
Guanidinoacetate methyltransferase (GAMT) deficiency is a rare inherited disorder characterized by creatine (Cr) depletion and guanidinoacetate (GAA) accumulation in body fluids. We report the first identified Chinese case, diagnosed in a 4-year-old girl with onset of global developmental. Low Cr and high GAA levels were detected in her serum and urine, and low Cr level in her brain. Compound heterozygous variants in GAMT gene were found, including a previously reported variant at c.491dupG which was inherited from her mother and a novel variant at c.564G>T, which was inherited from her father. The Cr and GAA levels returned back to normal after 3 months of treatment. After one year of treatment, the patient stopped taking antiepileptic drugs and her electroencephalogram (EEG) was also back to normal. The girl was followed up for five years and exhibited good results beyond our expectation. The results have shown that protein restriction with high-dose ornithine and creatine supplements have strong therapeutic potential for our patient.
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Guanidinoacetato N-Metiltransferasa/deficiencia , Trastornos del Desarrollo del Lenguaje/tratamiento farmacológico , Trastornos del Desarrollo del Lenguaje/genética , Trastornos del Movimiento/congénito , Preescolar , Creatina/farmacología , Creatina/uso terapéutico , Relación Dosis-Respuesta a Droga , Femenino , Estudios de Seguimiento , Guanidinoacetato N-Metiltransferasa/genética , Guanidinoacetato N-Metiltransferasa/metabolismo , Humanos , Trastornos del Desarrollo del Lenguaje/enzimología , Trastornos del Movimiento/tratamiento farmacológico , Trastornos del Movimiento/enzimología , Trastornos del Movimiento/genética , Ornitina/farmacología , Ornitina/uso terapéutico , Resultado del TratamientoRESUMEN
Epilepsy commonly presents in childhood as part of a syndrome, and some such children may reach adult services without an underlying syndromic diagnosis. For adult neurologists taking over their care, it is often unclear how hard to search for an underlying diagnosis. The diagnostic yield may be small and such a diagnosis may not change management. Young adults with learning difficulties are also challenging to investigate, as they may not tolerate standard epilepsy tests.We present such a case in which simple tests identified a unifying diagnosis. With the new diagnosis came a new treatment that had a significant impact on seizures and quality of life.
Asunto(s)
Epilepsia/etiología , Epilepsia/terapia , Guanidinoacetato N-Metiltransferasa/deficiencia , Trastornos del Desarrollo del Lenguaje/complicaciones , Trastornos del Movimiento/congénito , Adulto , Creatina/sangre , Creatina/uso terapéutico , Creatina/orina , Guanidinoacetato N-Metiltransferasa/sangre , Guanidinoacetato N-Metiltransferasa/genética , Guanidinoacetato N-Metiltransferasa/metabolismo , Guanidinoacetato N-Metiltransferasa/orina , Humanos , Trastornos del Desarrollo del Lenguaje/genética , Trastornos del Desarrollo del Lenguaje/metabolismo , Masculino , Trastornos del Movimiento/complicaciones , Trastornos del Movimiento/genética , Trastornos del Movimiento/metabolismo , Mutación/genéticaRESUMEN
Long interspersed element-1 (LINE-1) is a mammalian transposable element, and its genomic insertion could cause neurological disorders in humans. Incidentally, LINE-1 is present in intron 3 of the microphthalmia-associated transcription factor (Mitf) gene of the black-eyed white mouse (Mitfmi-bw allele). Mice homozygous for the Mitfmi-bw allele show the white coat color with black eye and deafness. Here, we explored the functional consequences of the LINE-1 insertion in the Mitf gene using homozygous Mitfmi-bw mice on the C3H background (C3H-bw mice) or on the C57BL/6 background (bw mice). The open-field test showed that C3H-bw mice moved more irregularly in an unfamiliar environment during the 20-min period, compared to wild-type mice, suggesting the altered emotionality. Moreover, C3H-bw mice showed the lower serum creatinine levels, which may reflect the creatine deficiency. In fact, morphologically abnormal neurons and astrocytes were detected in the frontal cortex of bw mice. The immunohistochemical analysis of bw mouse tissues showed the lower intensity for expression of guanidinoacetate methyltransferase, a key enzyme in creatine synthesis, in neurons of the frontal cortex and in glomeruli and renal tubules. Thus, Mitf may ensure the elongation of axons and dendrites by maintaining creatine synthesis in the frontal cortex.