RESUMO
Succinic semialdehyde dehydrogenase deficiency (SSADHD) is an inherited metabolic disorder of γ-aminobutyrate (GABA) catabolism. Cerebral waste clearance along glymphatic perivascular spaces depends on aquaporin 4 (AQP4) water channels, the function of which was shown to be influenced by GABA. Sleep disturbances are associated independently with SSADHD and glymphatic dysfunction. This study aimed to determine whether indices of the hyperGABAergic state characteristic of SSADHD coincide with glymphatic dysfunction and sleep disturbances and to explicate the modulatory effect that GABA may have on the glymphatic system. The study included 42 individuals (21 with SSADHD; 21 healthy controls) who underwent brain MRIs and magnetic resonance spectroscopy (MRS) for assessment of glymphatic dysfunction and cortical GABA, plasma GABA measurements, and circadian clock gene expression. The SSADHD subjects responded to an additional Children's Sleep Habits Questionnaire (CSHQ). Compared with the control group, SSADHD subjects did not differ in sex and age but had a higher severity of enlarged perivascular spaces in the centrum semiovale (p < 0.001), basal ganglia (p = 0.01), and midbrain (p = 0.001), as well as a higher MRS-derived GABA/NAA peak (p < 0.001). Within the SSADHD group, the severity of glymphatic dysfunction was specific for a lower MRS-derived GABA/NAA (p = 0.04) and lower plasma GABA (p = 0.004). Additionally, the degree of their glymphatic dysfunction correlated with the CSHQ-estimated sleep disturbances scores (R = 5.18, p = 0.03). In the control group, EPVS burden did not correlate with age or cerebral and plasma GABA values. The modulatory effect that GABA may exert on the glymphatic system has therapeutic implications for sleep-related disorders and neurodegenerative conditions associated with glymphatic dysfunction.
Assuntos
Erros Inatos do Metabolismo dos Aminoácidos , Sistema Glinfático , Imageamento por Ressonância Magnética , Transtornos do Sono-Vigília , Succinato-Semialdeído Desidrogenase , Ácido gama-Aminobutírico , Humanos , Masculino , Feminino , Ácido gama-Aminobutírico/metabolismo , Erros Inatos do Metabolismo dos Aminoácidos/fisiopatologia , Erros Inatos do Metabolismo dos Aminoácidos/complicações , Transtornos do Sono-Vigília/fisiopatologia , Sistema Glinfático/fisiopatologia , Criança , Succinato-Semialdeído Desidrogenase/deficiência , Espectroscopia de Ressonância Magnética , Adolescente , Encéfalo/diagnóstico por imagem , Encéfalo/fisiopatologia , Encéfalo/metabolismo , Aquaporina 4 , Laringoestenose/fisiopatologia , Pré-Escolar , Deficiências do DesenvolvimentoRESUMO
To investigate the genotype-to-protein-to-phenotype correlations of succinic semialdehyde dehydrogenase deficiency (SSADHD), an inherited metabolic disorder of γ-aminobutyric acid catabolism. Bioinformatics and in silico mutagenesis analyses of ALDH5A1 variants were performed to evaluate their impact on protein stability, active site and co-factor binding domains, splicing, and homotetramer formation. Protein abnormalities were then correlated with a validated disease-specific clinical severity score and neurological, neuropsychological, biochemical, neuroimaging, and neurophysiological metrics. A total of 58 individuals (1:1 male/female ratio) were affected by 32 ALDH5A1 pathogenic variants, eight of which were novel. Compared to individuals with single homotetrameric or multiple homo and heterotetrameric proteins, those predicted not to synthesize any functional enzyme protein had significantly lower expression of ALDH5A1 (p = 0.001), worse overall clinical outcomes (p = 0.008) and specifically more severe cognitive deficits (p = 0.01), epilepsy (p = 0.04) and psychiatric morbidity (p = 0.04). Compared to individuals with predictions of having no protein or a protein impaired in catalytic functions, subjects whose proteins were predicted to be impaired in stability, folding, or oligomerization had a better overall clinical outcome (p = 0.02) and adaptive skills (p = 0.04). The quantity and type of enzyme proteins (no protein, single homotetramers, or multiple homo and heterotetramers), as well as their structural and functional impairments (catalytic or stability, folding, or oligomerization), contribute to phenotype severity in SSADHD. These findings are valuable for assessment of disease prognosis and management, including patient selection for gene replacement therapy. Furthermore, they provide a roadmap to determine genotype-to-protein-to-phenotype relationships in other autosomal recessive disorders.
Assuntos
Erros Inatos do Metabolismo dos Aminoácidos , Criança , Humanos , Masculino , Feminino , Erros Inatos do Metabolismo dos Aminoácidos/genética , Erros Inatos do Metabolismo dos Aminoácidos/metabolismo , Erros Inatos do Metabolismo dos Aminoácidos/patologia , Deficiências do Desenvolvimento/genética , Fenótipo , Succinato-Semialdeído Desidrogenase/genética , Succinato-Semialdeído Desidrogenase/metabolismoRESUMO
OBJECTIVE: Succinic semialdehyde dehydrogenase deficiency (SSADHD) is a rare inherited metabolic disorder caused by a defect of γ-aminobutyrate (GABA) catabolism. Despite the resultant hyper-GABAergic environment facilitated by the metabolic defect, individuals with this disorder have a paradoxically high prevalence of epilepsy. We aimed to study the characteristics of epilepsy in SSADHD and its concordance with GABA-related metabolites and neurophysiologic markers of cortical excitation. METHODS: Subjects in an international natural history study of SSADHD underwent clinical assessments, electroencephalography, transcranial magnetic stimulation (TMS), magnetic resonance spectroscopy for GABA/N-acetyl aspartate quantification, and plasma GABA-related metabolite measurements. RESULTS: A total of 61 subjects with SSADHD and 42 healthy controls were included in the study. Epilepsy was present in 49% of the SSADHD cohort. Over time, there was an increase in severity in 33% of the subjects with seizures. The presence of seizures was associated with increasing age (p = .001) and lower levels of GABA (p = .002), γ-hydroxybutyrate (GHB; p = .004), and γ-guanidinobutyrate (GBA; p = .003). Seizure severity was associated with increasing age and lower levels of GABA-related metabolites as well as lower TMS-derived resting motor thresholds (p = .04). The cutoff values with the highest discriminative ability to predict seizures were age > 9.2 years (p = .001), GABA < 2.57 µmol·L-1 (p = .002), GHB < 143.6 µmol·L-1 (p = .004), and GBA < .075 µmol·L-1 (p = .007). A prediction model for seizures in SSADHD was comprised of the additive effect of older age and lower plasma GABA, GHB, and GBA (area under the receiver operating characteristic curve of .798, p = .008). SIGNIFICANCE: Epilepsy is highly prevalent in SSADHD, and its onset and severity correlate with an age-related decline in GABA and GABA-related metabolite levels as well as TMS markers of reduced cortical inhibition. The reduction of GABAergic activity in this otherwise hyper-GABAergic disorder demonstrates a concordance between epileptogenesis and compensatory responses. These findings may furthermore inform the timing of molecular interventions for SSADHD.
Assuntos
Erros Inatos do Metabolismo dos Aminoácidos , Epilepsia , Oxibato de Sódio , Humanos , Criança , Erros Inatos do Metabolismo dos Aminoácidos/complicações , Erros Inatos do Metabolismo dos Aminoácidos/metabolismo , Deficiências do Desenvolvimento , Epilepsia/metabolismo , Ácido gama-Aminobutírico/metabolismo , Aminobutiratos , ConvulsõesRESUMO
Pathophysiology associated with Huntington's disease (HD) has been studied extensively in various cell and animal models since the 1993 discovery of the mutant huntingtin (mHtt) with abnormally expanded polyglutamine (polyQ) tracts as the causative factor. However, the sequence of early pathophysiological events leading to HD still remains elusive. To gain new insights into the early polyQ-induced pathogenic events, we expressed Htt exon1 (Httex1) with a normal (21), or an extended (42 or 63) number of polyQ in tobacco plants. Here, we show that transgenic plants accumulated Httex1 proteins with corresponding polyQ tracts, and mHttex1 induced protein aggregation and affected plant growth, especially root and root hair development, in a polyQ length-dependent manner. Quantitative proteomic analysis of young roots from severely affected Httex1Q63 and unaffected Httex1Q21 plants showed that the most reduced protein by polyQ63 is a GTP cyclohydrolase I (GTPCH) along with many of its related one-carbon (C1) metabolic pathway enzymes. GTPCH is a key enzyme involved in folate biosynthesis in plants and tetrahydrobiopterin (BH4) biosynthesis in mammals. Validating studies in 4-week-old R6/2 HD mice expressing a mHttex1 showed reduced levels of GTPCH and dihydrofolate reductase (DHFR, a key folate utilization/alternate BH4 biosynthesis enzyme), and impaired C1 and BH4 metabolism. Our findings from mHttex1 plants and mice reveal impaired expressions of GTPCH and DHFR and may contribute to a better understanding of mHtt-altered C1 and BH4 metabolism, and their roles in the pathogenesis of HD.
Assuntos
GTP Cicloidrolase , Doença de Huntington , Plantas Geneticamente Modificadas , Animais , Camundongos , Carbono , Ácido Fólico , GTP Cicloidrolase/metabolismo , Proteína Huntingtina/genética , Doença de Huntington/metabolismo , Agregados Proteicos , Proteômica , Tetra-Hidrofolato Desidrogenase/metabolismoRESUMO
BACKGROUND: Knowledge of metabolic processes affected by major hepatectomy (MHx), and the metabolic pathways involved in liver regeneration and recovery of function, is limited and mainly derived from animal models. Assessment of restoration of hepatic function is essential in human living liver donors (LD). METHODS: We used a targeted metabolomic approach to longitudinally quantify changes in plasma and urine biomarkers from healthy LD. The biomarkers were analyzed before MHx and at scheduled intervals up to 12 months thereafter. RESULTS: Marked changes were found in the concentration of 15 primary and secondary plasma bile acids. Most significant changes occurred 2 days after MHx and persisted for up to 3 months. In addition, there were significant changes in acylcarnitine, phospholipid, and amino acid metabolism. The sum of aromatic amino acids and the Fischer ratio, both metabolic markers of liver damage, and the symmetrically demethylated arginine to arginine ratio, a marker of kidney function, were affected. CONCLUSIONS: This is the first comprehensive longitudinal study investigating metabolic processes during recovery of liver function after MHx in LD. It provides further evidence of full restoration of metabolic processes 3 months after MHx and supports future investigation to understand how metabolic changes affect donors' hepatic function.
Assuntos
Regeneração Hepática , Fígado , Animais , Hepatectomia , Humanos , Doadores Vivos , Estudos LongitudinaisRESUMO
d-2-hydroxyglutarate (D2HG) is produced in the tricarboxylic acid cycle and is quickly converted to α-ketoglutarate by d-2-hydroxyglutarate dehydrogenase (D2HGDH). In a mouse model of colitis-associated colon cancer (CAC), urine level of D2HG during colitis correlates positively with subsequent polyp counts and severity of dysplasia. The i.p. injection of D2HG results in delayed recovery from colitis and severe tumorigenesis. The colonic expression of D2HGDH is decreased in ulcerative colitis (UC) patients at baseline who progress to cancer. Hypoxia-inducible factor (Hif)-1α is a key regulator of D2HGDH transcription. Our study identifies urine D2HG and tissue D2HGDH expression as biomarkers to identify patients at risk for progressing from colitis to cancer. The D2HG/D2HGDH pathway provides potential therapeutic targets for the treatment of CAC.
Assuntos
Colite/metabolismo , Colite/patologia , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Glutaratos/metabolismo , Animais , Apoptose , Azoximetano/química , Biomarcadores Tumorais/metabolismo , Biópsia , Células CACO-2 , Movimento Celular , Proliferação de Células , Sobrevivência Celular , Colite Ulcerativa/metabolismo , Colite Ulcerativa/patologia , Sulfato de Dextrana/química , Progressão da Doença , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Inflamação , Mucosa Intestinal/patologia , Camundongos , RiscoRESUMO
BACKGROUND: Using blood specimens from untreated early Parkinson's disease (PD) patients from the DATATOP trial, we found that subjects in the low serum vitamin B12 tertile experienced greater annualized change in ambulatory capacity score, whereas those with moderately elevated (>15 µmol/L) total homocysteine had greater annualized declines in the Mini-Mental State Exam. METHODS: In this this study we sought to determine whether levels of cerebrospinal fluid (CSF) B12 markers were also associated with progression of PD. RESULTS: The annualized change in the UPDRS "walking" item, a component of the ambulatory capacity score, was worse in the low B12 tertile. No association with change in the Mini-Mental State Exam was seen for those 7% with the highest baseline CSF total homocysteine. CONCLUSIONS: In these untreated early-PD subjects, low CSF B12 predicted greater worsening of the UPDRS "walking" item, whereas CSF total homocysteine was not associated with progression of cognitive impairment. These findings extend and partially support our findings in serum. © 2020 The Authors. Movement Disorders published by Wiley Periodicals, Inc. on behalf of International Parkinson and Movement Disorder Society.
Assuntos
Doença de Parkinson , Biomarcadores , Progressão da Doença , Humanos , Testes de Estado Mental e Demência , Vitamina B 12RESUMO
In humans, vitamin B12 deficiency causes peripheral and CNS manifestations. Loss of myelin in the peripheral nerves and the spinal cord (SC) contributes to peripheral neuropathy and motor deficits. The metabolic basis for the demyelination and brain disorder is unknown. The transcobalamin receptor-knockout mouse ( Cd320-/-) develops cobalamin (Cbl) deficiency in the nervous system, with mild anemia. A decreased S-adenosylmethionine: S-adenosylhomocysteine ratio and increased methionine were seen in the brain with no significant changes in neurotransmitter metabolites. The structural pathology in the SC presented as loss of myelin in the axonal tracts with inflammation. The sciatic nerve (SN) showed increased nonuniform, internodal segments suggesting demyelination, and remyelination in progress. Consistent with these changes, the Cd320-/- mouse showed an increased latency to thermal nociception. Further, lower amplitude of compound action potential in the SN suggested that the functional capacity of the heavily myelinated axons were preferentially compromised, leading to loss of peripheral sensation. Although the metabolic basis for the demyelination and the structural and functional alterations of the nervous system in Cbl deficiency remain unresolved, the Cd320-/- mouse provides a unique model to investigate the pathologic consequences of vitamin B12 deficiency. -Arora, K., Sequeira, J. M., Alarcon, J. M., Wasek, B., Arning, E., Bottiglieri, T., Quadros, E. V. Neuropathology of vitamin B12 deficiency in the Cd320-/- mouse.
Assuntos
Encéfalo/patologia , Doenças do Sistema Nervoso/patologia , Nociceptividade , Receptores de Superfície Celular/fisiologia , Deficiência de Vitamina B 12/complicações , Animais , Encéfalo/metabolismo , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Doenças do Sistema Nervoso/etiologia , Doenças do Sistema Nervoso/metabolismo , Neurotransmissores/metabolismo , Deficiência de Vitamina B 12/fisiopatologiaRESUMO
Methyl-donor deficiency is a risk factor for neurodegenerative diseases. Dietary deficiency of the methyl-donors methionine and choline [methionine-choline-deficient (MCD) diet] is a well-established model of nonalcoholic steatohepatitis (NASH), yet brain metabolism has not been studied in this model. We hypothesized that supplemental betaine would protect both the liver and brain in this model and that any benefit to the brain would be due to improved liver metabolism because betaine is a methyl-donor in liver methylation but is not metabolically active in the brain. We fed male Sprague-Dawley rats a control diet, MCD diet, or betaine-supplemented MCD (MCD+B) diet for 8 wk and collected blood and tissue. As expected, betaine prevented MCD diet-induced NASH. However, contrary to our prediction, it did not appear to do so by stimulating methylation; the MCD+B diet worsened hyperhomocysteinemia and depressed liver methylation potential 8-fold compared with the MCD diet. Instead, it significantly increased the expression of genes involved in ß-oxidation: fibroblast growth factor 21 and peroxisome proliferator-activated receptor α. In contrast to that of the liver, brain methylation potential was unaffected by diet. Nevertheless, several phospholipid (PL) subclasses involved in stabilizing brain membranes were decreased by the MCD diet, and these improved modestly with betaine. The protective effect of betaine is likely due to the stimulation of ß-oxidation in liver and the effects on PL metabolism in brain.-Abu Ahmad, N., Raizman, M., Weizmann, N., Wasek, B., Arning, E., Bottiglieri, T., Tirosh, O., Troen, A. M. Betaine attenuates pathology by stimulating lipid oxidation in liver and regulating phospholipid metabolism in brain of methionine-choline-deficient rats.
Assuntos
Betaína/uso terapêutico , Deficiência de Colina/tratamento farmacológico , Deficiência de Colina/metabolismo , Metabolismo dos Lipídeos/efeitos dos fármacos , Fígado/efeitos dos fármacos , Fígado/metabolismo , Metionina/deficiência , Metionina/metabolismo , Fosfolipídeos/metabolismo , Animais , Western Blotting , Masculino , Aprendizagem em Labirinto , Ratos , Ratos Sprague-DawleyRESUMO
Upon publication, it was noted that five of the on-line supplementary figures had incorrect figure: figure legend associations. These were supplementary Figs. 6, 7, 14, 15, and 23.
RESUMO
Metabolomic characterization of post-mortem tissues (frontal and parietal cortices, pons, cerebellum, hippocampus, cerebral cortex, liver and kidney) derived from a 37 y.o. male patient with succinic semialdehyde dehydrogenase deficiency (SSADHD) was performed in conjunction with four parallel series of control tissues. Amino acids, acylcarnitines, guanidino- species (guanidinoacetic acid, creatine, creatinine) and GABA-related intermediates were quantified using UPLC and mass spectrometric methods that included isotopically labeled internal standards. Amino acid analyses revealed significant elevation of aspartic acid and depletion of glutamine in patient tissues. Evidence for disruption of short-chain fatty acid metabolism, manifest as altered C4OH, C5, C5:1, C5DC (dicarboxylic) and C12OH carnitines, was observed. Creatine and guanidinoacetic acids were decreased and elevated, respectively. GABA-associated metabolites (total GABA, γ-hydroxybutyric acid, succinic semialdehyde, 4-guanidinobutyrate, 4,5-dihydroxyhexanoic acid and homocarnosine) were significantly increased in patient tissues, including liver and kidney. The data support disruption of fat, creatine and amino acid metabolism as a component of the pathophysiology of SSADHD, and underscore the observation that metabolites measured in patient physiological fluids provide an unreliable reflection of brain metabolism.
Assuntos
Erros Inatos do Metabolismo dos Aminoácidos/metabolismo , Aminoácidos/metabolismo , Encéfalo/metabolismo , Carnitina/análogos & derivados , Creatina/metabolismo , Creatinina/metabolismo , Deficiências do Desenvolvimento/metabolismo , Glicina/análogos & derivados , Succinato-Semialdeído Desidrogenase/deficiência , Ácido gama-Aminobutírico/análogos & derivados , Adulto , Erros Inatos do Metabolismo dos Aminoácidos/patologia , Encéfalo/patologia , Carnitina/metabolismo , Deficiências do Desenvolvimento/patologia , Glicina/metabolismo , Humanos , Masculino , Metabolômica , Succinato-Semialdeído Desidrogenase/metabolismo , Ácido gama-Aminobutírico/metabolismoRESUMO
The prevalence of stroke increases with age and the ability to absorb all nutrients from our diets decreases with age. Nutrition is a modifiable risk factor for stroke, which is a leading cause of death and disability in world-wide. Deficiencies in onecarbon metabolism, including in methyltetrahydrofolate reductase (MTHFR), have been linked to increased risk of stroke. The Mthfr+/- mice mouse model mimic the phenotype of the MTHFR677CâT polymorphism, such as elevated levels of homocystine. Using this mouse model, the aim of this study was to investigate the impact of dietary supplementation with 5-methylTHF, vitamin B12, and choline after ischemic stroke. Male Mthfr+/- and wildtype littermate control mice were aged (~1.5-year-old) and were placed on control diet (CD) 4-weeks prior to sensorimotor cortex damage using photothrombosis (PT), a model for ischemic stroke. Post-operatively, one group of Mthfr+/- and wildtype littermate mice were placed on 5-methylTHF, vitamin B12, and choline supplemented diet (SD). Four weeks after PT and SD motor function was assessed using the accelerating rotarod, forepaw asymmetry, and ladder beam walking tasks. Total homocysteine and cysteine levels were measured in blood. Brain tissue was processed to assess lesion volume and investigate biochemical and molecular changes. After PT and SD, Mthfr+/- mice were able to stay on the accelerating rotarod longer and used their impaired forepaw to explore more when compared to CD animals. Furthermore, total homocysteine levels in plasma and lesion volume were reduced in Mthfr+/+ and Mthfr+/- SD mice. Within the damage site, there were reduced levels of apoptotic cell death and increased neuroprotective cellular response in the brains of SD treated Mthfr+/- mice. This study reveals a critical role for onecarbon supplementation, with 5-methylTHF, vitamin B12, and choline, in supporting improvement after ischemic stroke damage.
Assuntos
Colina/farmacologia , Suplementos Nutricionais , Metilenotetra-Hidrofolato Redutase (NADPH2)/deficiência , Acidente Vascular Cerebral/fisiopatologia , Tetra-Hidrofolatos/farmacologia , Vitamina B 12/farmacologia , Envelhecimento , Animais , Encéfalo/efeitos dos fármacos , Encéfalo/patologia , Encéfalo/fisiopatologia , Masculino , Metilenotetra-Hidrofolato Redutase (NADPH2)/genética , Camundongos , Camundongos Endogâmicos C57BL , Recuperação de Função Fisiológica/efeitos dos fármacosRESUMO
Fabry disease is caused by deficient activity of α-galactosidase A and subsequent accumulation of glycosphingolipids (mainly globotriaosylceramide, Gb3), leading to multisystem organ dysfunction. Oxidative stress and nitric oxide synthase (NOS) uncoupling are thought to contribute to Fabry cardiovascular diseases. We hypothesized that decreased tetrahydrobiopterin (BH4) plays a role in the pathogenesis of Fabry disease. We found that BH4 was decreased in the heart and kidney but not in the liver and aorta of Fabry mice. BH4 was also decreased in the plasma of female Fabry patients, which was not corrected by enzyme replacement therapy (ERT). Gb3 levels were inversely correlated with BH4 levels in animal tissues and cultured patient cells. To investigate the role of BH4 deficiency in disease phenotypes, 12-month-old Fabry mice were treated with gene transfer-mediated ERT or substrate reduction therapy (SRT) for 6 months. In the Fabry mice receiving SRT but not ERT, BH4 deficiency was restored, concomitant with ameliorated cardiac and renal hypertrophy. Additionally, glutathione levels were decreased in Fabry mouse tissues in a sex-dependent manner. Renal BH4 levels were closely correlated with glutathione levels and inversely correlated with cardiac and kidney weight. In conclusion, this study showed that BH4 deficiency occurs in Fabry disease and may contribute to the pathogenesis of the disease through oxidative stress associated with a reduced antioxidant capacity of cells and NOS uncoupling. This study also suggested dissimilar efficacy of ERT and SRT in correcting pre-existing pathologies in Fabry disease.
Assuntos
Biopterinas/análogos & derivados , Terapia de Reposição de Enzimas , Doença de Fabry/genética , alfa-Galactosidase/genética , Animais , Biopterinas/deficiência , Biopterinas/genética , Biopterinas/metabolismo , Modelos Animais de Doenças , Doença de Fabry/mortalidade , Doença de Fabry/fisiopatologia , Feminino , Glutationa/metabolismo , Glicoesfingolipídeos/metabolismo , Humanos , Rim/metabolismo , Rim/patologia , Camundongos , Miocárdio/metabolismo , Miocárdio/patologia , Óxido Nítrico Sintase/genética , Óxido Nítrico Sintase/metabolismo , Estresse Oxidativo/genética , alfa-Galactosidase/biossíntese , alfa-Galactosidase/metabolismoRESUMO
Increased gamma-hydroxybutyric acid in urine and blood are metabolic hallmarks of succinic semialdehyde dehydrogenase deficiency, a defect of 4-aminobutyric acid metabolism. Here, we examined the hypothesis that succinic semialdehyde dehydrogenase deficiency could be identified via measurement of gamma-hydroxybutyric acid in newborn and post-newborn dried bloodspots. Quantitation of gamma-hydroxybutyric acid using liquid chromatography-tandem mass spectrometry in twelve archival newborn patient dried bloodspots was 360⯱â¯57⯵M (mean, standard error; range 111-767), all values exceeding the previously established cutoff for newborn detection of 78 µΜ established from 2831 dried bloodspots derived from newborns, neonates and children. Gamma-hydroxybutyric acid in post-newborn dried bloodspots (nâ¯=â¯19; ages 0.8-38â¯years) was 191⯱â¯65⯵M (mean, standard error; range 20-1218), exceeding the aforementioned GHB cutoff for patients approximately 10â¯years of age or younger. Further, gamma-hydroxybutyric acid in post-newborn dried bloodspots displayed a significant (pâ¯<â¯.0001) inverse correlation with age. This preliminary study suggests that succinic semialdehyde dehydrogenase deficiency may be identified in newborn and post-newborn dried bloodspots via quantitation of gamma-hydroxybutyric acid, while forming the platform for more extensive studies in affected and unaffected dried bloodspots.
Assuntos
Erros Inatos do Metabolismo dos Aminoácidos/diagnóstico , Deficiências do Desenvolvimento/diagnóstico , Teste em Amostras de Sangue Seco , Triagem Neonatal/métodos , Oxibato de Sódio/sangue , Succinato-Semialdeído Desidrogenase/deficiência , Adolescente , Adulto , Erros Inatos do Metabolismo dos Aminoácidos/sangue , Criança , Pré-Escolar , Deficiências do Desenvolvimento/sangue , Feminino , Humanos , Lactente , Recém-Nascido , Masculino , Succinato-Semialdeído Desidrogenase/sangue , Adulto JovemRESUMO
Murine succinic semialdehyde dehydrogenase deficiency (SSADHD) manifests with high concentrations of γ-aminobutyric acid (GABA) and γ-hydroxybutyrate (GHB) and low glutamine in the brain. To understand the pathogenic contribution of central glutamine deficiency, we exposed aldh5a1-/- (SSADHD) mice and their genetic controls (aldh5a1+/+ ) to either a 4% (w/w) glutamine-containing diet or a glutamine-free diet from conception until postnatal day 30. Endpoints included brain, liver and blood amino acids, brain GHB, ataxia scores, and open field testing. Glutamine supplementation did not improve aldh5a1-/- brain glutamine deficiency nor brain GABA and GHB. It decreased brain glutamate but did not change the ratio of excitatory (glutamate) to inhibitory (GABA) neurotransmitters. In contrast, glutamine supplementation significantly increased brain arginine (30% for aldh5a1+/+ and 18% for aldh5a1-/- mice), and leucine (12% and 18%). Glutamine deficiency was confirmed in the liver. The test diet increased hepatic glutamate in both genotypes, decreased glutamine in aldh5a1+/+ but not in aldh5a1-/- , but had no effect on GABA. Dried bloodspot analyses showed significantly elevated GABA in mutants (approximately 800% above controls) and decreased glutamate (approximately 25%), but no glutamine difference with controls. Glutamine supplementation did not impact blood GABA but significantly increased glutamine and glutamate in both genotypes indicating systemic exposure to dietary glutamine. Ataxia and pronounced hyperactivity were observed in aldh5a1-/- mice but remained unchanged by the diet intervention. The study suggests that glutamine supplementation improves peripheral but not central glutamine deficiency in experimental SSADHD. Future studies are needed to fully understand the pathogenic role of brain glutamine deficiency in SSADHD.
Assuntos
Erros Inatos do Metabolismo dos Aminoácidos/genética , Erros Inatos do Metabolismo dos Aminoácidos/metabolismo , Biomarcadores/sangue , Deficiências do Desenvolvimento/genética , Deficiências do Desenvolvimento/metabolismo , Glutamina/administração & dosagem , Succinato-Semialdeído Desidrogenase/deficiência , Erros Inatos do Metabolismo dos Aminoácidos/sangue , Aminoácidos/metabolismo , Animais , Encéfalo/patologia , Deficiências do Desenvolvimento/sangue , Suplementos Nutricionais , Modelos Animais de Doenças , Feminino , Humanos , Masculino , Fenômenos Fisiológicos da Nutrição Materna , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Succinato-Semialdeído Desidrogenase/sangue , Succinato-Semialdeído Desidrogenase/genética , Succinato-Semialdeído Desidrogenase/metabolismo , Ácido gama-Aminobutírico/metabolismoRESUMO
RATIONALE: Acquired resistance is an important driver of multidrug-resistant tuberculosis (TB), even with good treatment adherence. However, exactly what initiates the resistance and how it arises remain poorly understood. OBJECTIVES: To identify the relationship between drug concentrations and drug susceptibility readouts (minimum inhibitory concentrations [MICs]) in the TB cavity. METHODS: We recruited patients with medically incurable TB who were undergoing therapeutic lung resection while on treatment with a cocktail of second-line anti-TB drugs. On the day of surgery, antibiotic concentrations were measured in the blood and at seven prespecified biopsy sites within each cavity. Mycobacterium tuberculosis was grown from each biopsy site, MICs of each drug identified, and whole-genome sequencing performed. Spearman correlation coefficients between drug concentration and MIC were calculated. MEASUREMENTS AND MAIN RESULTS: Fourteen patients treated for a median of 13 months (range, 5-31 mo) were recruited. MICs and drug resistance-associated single-nucleotide variants differed between the different geospatial locations within each cavity, and with pretreatment and serial sputum isolates, consistent with ongoing acquisition of resistance. However, pretreatment sputum MIC had an accuracy of only 49.48% in predicting cavitary MICs. There were large concentration-distance gradients for each antibiotic. The location-specific concentrations inversely correlated with MICs (P < 0.05) and therefore acquired resistance. Moreover, pharmacokinetic/pharmacodynamic exposures known to amplify drug-resistant subpopulations were encountered in all positions. CONCLUSIONS: These data inform interventional strategies relevant to drug delivery, dosing, and diagnostics to prevent the development of acquired resistance. The role of high intracavitary penetration as a biomarker of antibiotic efficacy, when assessing new regimens, requires clarification.
Assuntos
Antituberculosos/farmacologia , Pulmão/efeitos dos fármacos , Pulmão/patologia , Tuberculose Resistente a Múltiplos Medicamentos/tratamento farmacológico , Tuberculose Resistente a Múltiplos Medicamentos/patologia , Adolescente , Adulto , Antituberculosos/uso terapêutico , Biópsia , Relação Dose-Resposta a Droga , Quimioterapia Combinada , Feminino , Humanos , Masculino , Testes de Sensibilidade Microbiana , Pessoa de Meia-Idade , Estudos Prospectivos , Adulto JovemRESUMO
Elevated levels of the ß-amyloid peptide (Aß) are thought to contribute to cognitive and behavioral impairments observed in Alzheimer's disease (AD). Protein phosphatase 2A (PP2A) participates in multiple molecular pathways implicated in AD, and its expression and activity are reduced in postmortem brains of AD patients. PP2A is regulated by protein methylation, and impaired PP2A methylation is thought to contribute to increased AD risk in hyperhomocysteinemic individuals. To examine further the link between PP2A and AD, we generated transgenic mice that overexpress the PP2A methylesterase, protein phosphatase methylesterase-1 (PME-1), or the PP2A methyltransferase, leucine carboxyl methyltransferase-1 (LCMT-1), and examined the sensitivity of these animals to behavioral and electrophysiological impairments caused by exogenous Aß exposure. We found that PME-1 overexpression enhanced these impairments, whereas LCMT-1 overexpression protected against Aß-induced impairments. Neither transgene affected Aß production or the electrophysiological response to low concentrations of Aß, suggesting that these manipulations selectively affect the pathological response to elevated Aß levels. Together these data identify a molecular mechanism linking PP2A to the development of AD-related cognitive impairments that might be therapeutically exploited to target selectively the pathological effects caused by elevated Aß levels in AD patients.
Assuntos
Peptídeos beta-Amiloides/fisiologia , Transtornos Cognitivos/fisiopatologia , Proteína Fosfatase 2/metabolismo , Animais , Comportamento Animal , Metilação , Camundongos , Camundongos TransgênicosRESUMO
Background: Suboptimal folate intake, a risk factor for birth defects, is common even in areas with folate fortification. A polymorphism in methylenetetrahydrofolate dehydrogenase 1 (MTHFD1), R653Q (MTHFD1 c.1958 G > A), has also been associated with increased birth defect risk, likely through reduced purine synthesis. Objective: We aimed to determine if the interaction of MTHFD1 synthetase deficiency and low folate intake increases developmental abnormalities in a mouse model for MTHFD1 R653Q. Methods: Female Mthfd1S+/+ and Mthfd1S+/- mice were fed control or low-folate diets (2 and 0.3 mg folic acid/kg diet, respectively) before mating and during pregnancy. Embryos and placentas were examined for anomalies at embryonic day 10.5. Maternal 1-carbon metabolites were measured in plasma and liver. Results: Delays and defects doubled in litters of Mthfd1S+/- females fed low-folate diets compared to wild-type females fed either diet, or Mthfd1S+/- females fed control diets [P values (defects): diet 0.003, maternal genotype 0.012, diet × maternal genotype 0.014]. These adverse outcomes were associated with placental dysmorphology. Intrauterine growth restriction was increased by embryonic Mthfd1S+/- genotype, folate deficiency, and interaction of maternal Mthfd1S+/- genotype with folate deficiency (P values: embryonic genotype 0.045, diet 0.0081, diet × maternal genotype 0.0019). Despite a 50% increase in methylenetetrahydrofolate reductase expression in low-folate maternal liver (P diet = 0.0007), methyltetrahydrofolate concentration decreased 70% (P diet <0.0001) and homocysteine concentration doubled in plasma (P diet = 0.0001); S-adenosylmethionine decreased 40% and S-adenosylhomocysteine increased 20% in low-folate maternal liver (P diet = 0.002 and 0.0002, respectively). Conclusions: MTHFD1 synthetase-deficient mice are more sensitive to low folate intake than wild-type mice during pregnancy. Reduced purine synthesis due to synthetase deficiency and altered methylation potential due to low folate may increase pregnancy complications. Further studies and individualized intake recommendations may be required for women homozygous for the MTHFD1 R653Q variant.
Assuntos
Anormalidades Congênitas/etiologia , Deficiência de Ácido Fólico/complicações , Ácido Fólico/administração & dosagem , Formiato-Tetra-Hidrofolato Ligase/deficiência , Genótipo , Meteniltetra-Hidrofolato Cicloidrolase/deficiência , Metilenotetra-Hidrofolato Desidrogenase (NADP)/deficiência , Enzimas Multifuncionais/deficiência , Polimorfismo Genético , Complicações na Gravidez/etiologia , Animais , Metilação de DNA , Dieta , Modelos Animais de Doenças , Feminino , Desenvolvimento Fetal , Retardo do Crescimento Fetal/etiologia , Ácido Fólico/sangue , Deficiência de Ácido Fólico/sangue , Deficiência de Ácido Fólico/genética , Deficiência de Ácido Fólico/metabolismo , Formiato-Tetra-Hidrofolato Ligase/genética , Formiato-Tetra-Hidrofolato Ligase/metabolismo , Ligases , Fígado/metabolismo , Meteniltetra-Hidrofolato Cicloidrolase/genética , Meteniltetra-Hidrofolato Cicloidrolase/metabolismo , Metilenotetra-Hidrofolato Desidrogenase (NADP)/genética , Metilenotetra-Hidrofolato Desidrogenase (NADP)/metabolismo , Metilenotetra-Hidrofolato Redutase (NADPH2)/metabolismo , Camundongos , Enzimas Multifuncionais/genética , Enzimas Multifuncionais/metabolismo , Placenta , Gravidez , Complicações na Gravidez/sangue , Complicações na Gravidez/genética , Complicações na Gravidez/metabolismo , Prenhez , S-Adenosil-Homocisteína/metabolismo , S-Adenosilmetionina/metabolismo , Tetra-Hidrofolatos/sangueRESUMO
Fabry disease is a glycosphingolipidosis caused by deficient activity of α-galactosidase A; it is one of a few diseases that are associated with priapism, an abnormal prolonged erection of the penis. The goal of this study was to investigate the pathogenesis of Fabry disease-associated priapism in a mouse model of the disease. We found that Fabry mice develop late-onset priapism. Neuronal nitric oxide synthase (nNOS), which was predominantly present as the 120-kDa N-terminus-truncated form, was significantly upregulated in the penis of 18-month-old Fabry mice compared to wild type controls (~fivefold). Endothelial NOS (eNOS) was also upregulated (~twofold). NO level in penile tissues of Fabry mice was significantly higher than wild type controls at 18 months. Gene transfer-mediated enzyme replacement therapy reversed abnormal nNOS expression in the Fabry mouse penis. The penile nNOS level was restored by antiandrogen treatment, suggesting that hyperactive androgen receptor signaling in Fabry mice may contribute to nNOS upregulation. However, the phosphodiesterase-5A expression level and the adenosine content in the penis, which are known to play roles in the development of priapism in other etiologies, were unchanged in Fabry mice. In conclusion, these data suggested that increased nNOS (and probably eNOS) content and the consequential elevated NO production and high arterial blood flow in the penis may be the underlying mechanism of priapism in Fabry mice. Furthermore, in combination with previous findings, this study suggested that regulation of NOS expression is susceptible to α-galactosidase A deficiency, and this may represent a general pathogenic mechanism of Fabry vasculopathy.
Assuntos
Doença de Fabry/complicações , Óxido Nítrico Sintase Tipo III/metabolismo , Óxido Nítrico Sintase Tipo I/metabolismo , Ereção Peniana , Pênis/enzimologia , Priapismo/etiologia , Animais , Modelos Animais de Doenças , Terapia de Reposição de Enzimas/métodos , Doença de Fabry/enzimologia , Doença de Fabry/fisiopatologia , Doença de Fabry/terapia , Terapia Genética/métodos , Masculino , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Óxido Nítrico/metabolismo , Pênis/fisiopatologia , Priapismo/enzimologia , Priapismo/fisiopatologia , Priapismo/terapia , Fluxo Sanguíneo Regional , Transdução de Sinais , Regulação para Cima , alfa-Galactosidase/biossíntese , alfa-Galactosidase/genéticaRESUMO
OBJECTIVE: Sepsis is characterized by microvascular dysfunction and thrombophilia. Several methionine metabolites may be relevant to this sepsis pathophysiology. S-adenosylmethionine (SAM) serves as the methyl donor for trans-methylation reactions. S-adenosylhomocysteine (SAH) is the by-product of these reactions and serves as the precursor to homocysteine. Relationships between plasma total homocysteine concentrations (tHcy) and vascular disease and thrombosis are firmly established. We hypothesized that SAM, SAH, and tHcy levels are elevated in patients with sepsis and associated with mortality. METHODS: This was a combined case-control and prospective cohort study consisting of 109 patients with sepsis and 50 control participants without acute illness. The study was conducted in the medical and surgical intensive care units of the University of Rochester Medical Center. Methionine, SAM, SAH, and tHcy concentrations were compared in patients with sepsis versus control participants and in sepsis survivors versus nonsurvivors. RESULTS: Patients with sepsis had significantly higher plasma SAM and SAH concentrations than control participants (SAM: 164 [107-227] vs73 [59-87 nM], P < .001; SAH: 99 [60-165] vs 35 [28-45] nM, P < .001). In contrast, plasma tHcy concentrations were lower in sepsis patients compared to healthy control participants (4 [2-6]) vs 7 [5-9] µM; P = .04). In multivariable analysis, quartiles of SAM, SAH, and tHcy were independently associated with sepsis ( P = .006, P = .05, and P < .001, respectively). Sepsis nonsurvivors had significantly higher plasma SAM and SAH concentrations than survivors (SAM: 223 [125-260] vs 136 [96-187] nM; P = .01; SAH: 139 [81-197] vs 86 [55-130] nM, P = .006). Plasma tHcy levels were similar in survivors vs nonsurvivors. The associations between SAM or SAH and hospital mortality were no longer significant after adjusting for renal dysfunction. CONCLUSIONS: Methionine metabolite concentrations are abnormal in sepsis and linked with clinical outcomes. Further study is required to determine whether these abnormalities have pathophysiologic significance.