Your browser doesn't support javascript.
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 341
Filtrar
1.
J Inherit Metab Dis ; 2020 Jan 18.
Artigo em Inglês | MEDLINE | ID: mdl-31955429

RESUMO

BACKGROUND: A maladaptive shift from fat to carbohydrate (CHO) oxidation during exercise is thought to underlie myopathy and exercise-induced rhabdomyolysis in patients with fatty acid oxidation (FAO) disorders. We hypothesized that ingestion of a ketone ester (KE) drink prior to exercise could serve as an alternative oxidative substrate supply to boost muscular ATP homeostasis. To establish a rational basis for therapeutic use of KE supplementation in FAO, we tested this hypothesis in patients deficient in Very Long-Chain acyl-CoA Dehydrogenase (VLCAD). METHODS: Five patients (range 17-45 y; 4M/1F) patients were included in an investigator-initiated, randomized, blinded, placebo-controlled, 2-way cross-over study. Patients drank either a KE+CHO mix or an isocaloric CHO equivalent and performed 35 min upright cycling followed by 10 minutes supine cycling inside a Magnetic Resonance scanner at individual maximal FAO work rate (fatmax; ~40% VO2 max). The protocol was repeated after a one-week interval with the alternate drink. Primary outcome measures were quadriceps phosphocreatine (PCr), Pi and pH dynamics during exercise and recovery assayed by in vivo 31 P-MR spectroscopy. Secondary outcomes included plasma and muscle metabolites and respiratory gas exchange recordings. RESULTS: Ingestion of KE rapidly induced mild ketosis and increased muscle BHB content. During exercise at FATMAX, VLCADD-specific plasma acylcarnitine levels, quadriceps glycolytic intermediate levels and in vivo Pi/PCr ratio were all lower in KE+CHO than CHO. CONCLUSION: These results provide a rational basis for future clinical trials of synthetic ketone ester supplementation therapy in patients with FAO disorders. This article is protected by copyright. All rights reserved.

2.
J Appl Genet ; 2019 Oct 18.
Artigo em Inglês | MEDLINE | ID: mdl-31628608

RESUMO

Zellweger spectrum disorders (ZSD) constitute a group of rare autosomal recessive disorders characterized by a defect in peroxisome biogenesis due to mutations in one of 13 PEX genes. The broad clinical heterogeneity especially in late-onset presenting patients and a mild phenotype complicates and delays the diagnostic process. Here, we report a case of mild ZSD, due to novel PEX1 variants. The patient presented with an early hearing loss, bilateral cataracts, and leukodystrophy on magnetic resonance (MR) images. Normal results of serum very-long-chain fatty acids (VLCFA) and phytanic acid were found. Molecular diagnostics were performed to uncover the etiology of the clinical phenotype. Using whole exome sequencing, there have been found two variants in the PEX1 gene-c.3450T>A (p.Cys1150*) and c.1769T>C (p.Leu590Pro). VLCFA measurement in skin fibroblasts and C26:0-lysoPC in dried blood spot therefore was performed. Both results were in line with the diagnosis of ZSD. To conclude, normal results of routine serum VLCFA and branched-chain fatty acid measurement do not exclude mild forms of ZSD. The investigation of C26:0-lysoPC should be included in the diagnostic work-up in patients with cataract, hearing loss, and leukodystrophy on MR images suspected to suffer from ZSD.

3.
Brain ; 142(11): 3382-3397, 2019 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-31637422

RESUMO

CTP:phosphoethanolamine cytidylyltransferase (ET), encoded by PCYT2, is the rate-limiting enzyme for phosphatidylethanolamine synthesis via the CDP-ethanolamine pathway. Phosphatidylethanolamine is one of the most abundant membrane lipids and is particularly enriched in the brain. We identified five individuals with biallelic PCYT2 variants clinically characterized by global developmental delay with regression, spastic para- or tetraparesis, epilepsy and progressive cerebral and cerebellar atrophy. Using patient fibroblasts we demonstrated that these variants are hypomorphic, result in altered but residual ET protein levels and concomitant reduced enzyme activity without affecting mRNA levels. The significantly better survival of hypomorphic CRISPR-Cas9 generated pcyt2 zebrafish knockout compared to a complete knockout, in conjunction with previously described data on the Pcyt2 mouse model, indicates that complete loss of ET function may be incompatible with life in vertebrates. Lipidomic analysis revealed profound lipid abnormalities in patient fibroblasts impacting both neutral etherlipid and etherphospholipid metabolism. Plasma lipidomics studies also identified changes in etherlipids that have the potential to be used as biomarkers for ET deficiency. In conclusion, our data establish PCYT2 as a disease gene for a new complex hereditary spastic paraplegia and confirm that etherlipid homeostasis is important for the development and function of the brain.

4.
Am J Hum Genet ; 105(3): 534-548, 2019 Sep 05.
Artigo em Inglês | MEDLINE | ID: mdl-31422819

RESUMO

Early-infantile encephalopathies with epilepsy are devastating conditions mandating an accurate diagnosis to guide proper management. Whole-exome sequencing was used to investigate the disease etiology in four children from independent families with intellectual disability and epilepsy, revealing bi-allelic GOT2 mutations. In-depth metabolic studies in individual 1 showed low plasma serine, hypercitrullinemia, hyperlactatemia, and hyperammonemia. The epilepsy was serine and pyridoxine responsive. Functional consequences of observed mutations were tested by measuring enzyme activity and by cell and animal models. Zebrafish and mouse models were used to validate brain developmental and functional defects and to test therapeutic strategies. GOT2 encodes the mitochondrial glutamate oxaloacetate transaminase. GOT2 enzyme activity was deficient in fibroblasts with bi-allelic mutations. GOT2, a member of the malate-aspartate shuttle, plays an essential role in the intracellular NAD(H) redox balance. De novo serine biosynthesis was impaired in fibroblasts with GOT2 mutations and GOT2-knockout HEK293 cells. Correcting the highly oxidized cytosolic NAD-redox state by pyruvate supplementation restored serine biosynthesis in GOT2-deficient cells. Knockdown of got2a in zebrafish resulted in a brain developmental defect associated with seizure-like electroencephalography spikes, which could be rescued by supplying pyridoxine in embryo water. Both pyridoxine and serine synergistically rescued embryonic developmental defects in zebrafish got2a morphants. The two treated individuals reacted favorably to their treatment. Our data provide a mechanistic basis for the biochemical abnormalities in GOT2 deficiency that may also hold for other MAS defects.

5.
Sci Rep ; 9(1): 10502, 2019 Jul 19.
Artigo em Inglês | MEDLINE | ID: mdl-31324846

RESUMO

The peroxisomal ABC transporter, Comatose (CTS), a full length transporter from Arabidopsis has intrinsic acyl-CoA thioesterase (ACOT) activity, important for physiological function. We used molecular modelling, mutagenesis and biochemical analysis to identify amino acid residues important for ACOT activity. D863, Q864 and T867 lie within transmembrane helix 9. These residues are orientated such that they might plausibly contribute to a catalytic triad similar to type II Hotdog fold thioesterases. When expressed in Saccharomyces cerevisiae, mutation of these residues to alanine resulted in defective of ß-oxidation. All CTS mutants were expressed and targeted to peroxisomes and retained substrate-stimulated ATPase activity. When expressed in insect cell membranes, Q864A and S810N had similar ATPase activity to wild type but greatly reduced ACOT activity, whereas the Walker A mutant K487A had greatly reduced ATPase and no ATP-dependent ACOT activity. In wild type CTS, ATPase but not ACOT was stimulated by non-cleavable C14 ether-CoA. ACOT activity was stimulated by ATP but not by non-hydrolysable AMPPNP. Thus, ACOT activity depends on functional ATPase activity but not vice versa, and these two activities can be separated by mutagenesis. Whether D863, Q864 and T867 have a catalytic role or play a more indirect role in NBD-TMD communication is discussed.

6.
J Inherit Metab Dis ; 42(5): 878-889, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31268564

RESUMO

Multiple acyl-CoA dehydrogenase deficiency (MADD) is an ultra-rare inborn error of mitochondrial fatty acid oxidation (FAO) and amino acid metabolism. Individual phenotypes and treatment response can vary markedly. We aimed to identify markers that predict MADD phenotypes. We performed a retrospective nationwide cohort study; then developed an MADD-disease severity scoring system (MADD-DS3) based on signs and symptoms with weighed expert opinions; and finally correlated phenotypes and MADD-DS3 scores to FAO flux (oleate and myristate oxidation rates) and acylcarnitine profiles after palmitate loading in fibroblasts. Eighteen patients, diagnosed between 1989 and 2014, were identified. The MADD-DS3 entails enumeration of eight domain scores, which are calculated by averaging the relevant symptom scores. Lifetime MADD-DS3 scores of patients in our cohort ranged from 0 to 29. FAO flux and [U-13 C]C2-, C5-, and [U-13 C]C16-acylcarnitines were identified as key variables that discriminated neonatal from later onset patients (all P < .05) and strongly correlated to MADD-DS3 scores (oleate: r = -.86; myristate: r = -.91; [U-13 C]C2-acylcarnitine: r = -.96; C5-acylcarnitine: r = .97; [U-13 C]C16-acylcarnitine: r = .98, all P < .01). Functional studies in fibroblasts were found to differentiate between neonatal and later onset MADD-patients and were correlated to MADD-DS3 scores. Our data may improve early prediction of disease severity in order to start (preventive) and follow-up treatment appropriately. This is especially relevant in view of the inclusion of MADD in population newborn screening programs.

7.
Hum Mutat ; 40(10): 1899-1904, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31187905

RESUMO

Primary carnitine deficiency is caused by a defect in the active cellular uptake of carnitine by Na+ -dependent organic cation transporter novel 2 (OCTN2). Genetic diagnostic yield for this metabolic disorder has been relatively low, suggesting that disease-causing variants are missed. We Sanger sequenced the 5' untranslated region (UTR) of SLC22A5 in individuals with possible primary carnitine deficiency in whom no or only one mutant allele had been found. We identified a novel 5'-UTR c.-149G>A variant which we characterized by expression studies with reporter constructs in HeLa cells and by carnitine-transport measurements in fibroblasts using a newly developed sensitive assay based on tandem mass spectrometry. This variant, which we identified in 57 of 236 individuals of our cohort, introduces a functional upstream out-of-frame translation initiation codon. We show that the codon suppresses translation from the wild-type ATG of SLC22A5, resulting in reduced OCTN2 protein levels and concomitantly lower transport activity. With an allele frequency of 24.2% the c.-149G>A variant is the most frequent cause of primary carnitine deficiency in our cohort and may explain other reported cases with an incomplete genetic diagnosis. Individuals carrying this variant should be clinically re-evaluated and monitored to determine if this variant has clinical consequences.

8.
Biochim Biophys Acta Mol Basis Dis ; 1865(10): 2774-2787, 2019 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-31207289

RESUMO

Zellweger spectrum disorders (ZSDs) are autosomal recessive diseases caused by defective peroxisome assembly. They constitute a clinical continuum from severe early lethal to relatively milder presentations in adulthood. Liver disease is a prevalent symptom in ZSD patients. The underlying pathogenesis for the liver disease, however, is not fully understood. We report a hypomorphic ZSD mouse model, which is homozygous for Pex1-c.2531G>A (p.G844D), the equivalent of the most common pathogenic variant found in ZSD, and which predominantly presents with liver disease. After introducing the Pex1-G844D allele by knock-in, we characterized homozygous Pex1-G844D mice for survival, biochemical parameters, including peroxisomal and mitochondrial functions, organ histology, and developmental parameters. The first 20 post-natal days (P20) were critical for survival of homozygous Pex1-G844D mice (~20% survival rate). Lethality was likely due to a combination of cholestatic liver problems, liver dysfunction and caloric deficit, probably as a consequence of defective bile acid biosynthesis. Survival beyond P20 was nearly 100%, but surviving mice showed a marked delay in growth. Surviving mice showed similar hepatic problems as described for mild ZSD patients, including hepatomegaly, bile duct proliferation, liver fibrosis and mitochondrial alterations. Biochemical analyses of various tissues showed the absence of functional peroxisomes accompanied with aberrant levels of peroxisomal metabolites predominantly in the liver, while other tissues were relatively spared. ur findings show that homozygous Pex1-G844D mice have a predominant liver disease phenotype, mimicking the hepatic pathology of ZSD patients, and thus constitute a good model to study pathogenesis and treatment of liver disease in ZSD patients.

10.
N Engl J Med ; 380(15): 1433-1441, 2019 04 11.
Artigo em Inglês | MEDLINE | ID: mdl-30970188

RESUMO

We report an inborn error of metabolism caused by an expansion of a GCA-repeat tract in the 5' untranslated region of the gene encoding glutaminase (GLS) that was identified through detailed clinical and biochemical phenotyping, combined with whole-genome sequencing. The expansion was observed in three unrelated patients who presented with an early-onset delay in overall development, progressive ataxia, and elevated levels of glutamine. In addition to ataxia, one patient also showed cerebellar atrophy. The expansion was associated with a relative deficiency of GLS messenger RNA transcribed from the expanded allele, which probably resulted from repeat-mediated chromatin changes upstream of the GLS repeat. Our discovery underscores the importance of careful examination of regions of the genome that are typically excluded from or poorly captured by exome sequencing.


Assuntos
Erros Inatos do Metabolismo dos Aminoácidos/genética , Ataxia/genética , Deficiências do Desenvolvimento/genética , Glutaminase/deficiência , Glutaminase/genética , Glutamina/metabolismo , Repetições de Microssatélites , Mutação , Atrofia/genética , Cerebelo/patologia , Pré-Escolar , Feminino , Genótipo , Glutamina/análise , Humanos , Masculino , Fenótipo , Reação em Cadeia da Polimerase , Sequenciamento Completo do Genoma
11.
Metabolites ; 9(3)2019 Mar 05.
Artigo em Inglês | MEDLINE | ID: mdl-30841653

RESUMO

Oxidative stress plays a role in the onset and progression of a number of diseases, such as Alzheimer's disease, diabetes and cancer, as well as ageing. Oxidative stress is caused by an increased production of reactive oxygen species and reduced antioxidant activity, resulting in the oxidation of glutathione. The ratio of reduced to oxidised glutathione is often used as a marker of the redox state in the cell. Whereas a variety of methods have been developed to measure glutathione in blood samples, methods to measure glutathione in cultured cells are scarce. Here we present a protocol to measure glutathione levels in cultured human and yeast cells using ultra-performance liquid chromatography-tandem mass spectrometry (UPLC⁻MS/MS).

12.
J Inherit Metab Dis ; 42(3): 414-423, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30761551

RESUMO

Most infants with very-long-chain acyl-CoA dehydrogenase deficiency (VLCADD) identified by newborn screening (NBS) are asymptomatic at the time of diagnosis and remain asymptomatic. If this outcome is due to prompt diagnosis and initiation of therapy, or because of identification of individuals with biochemical abnormalities who will never develop symptoms, is unclear. Therefore, a 10-year longitudinal national cohort study of genetically confirmed VLCADD patients born before and after introduction of NBS was conducted. Main outcome measures were clinical outcome parameters, acyl-CoA dehydrogenase very long chain gene analysis, VLCAD activity, and overall capacity of long-chain fatty acid oxidation (LC-FAO flux) in lymphocytes and cultured skin fibroblasts. Median VLCAD activity in lymphocytes of 54 patients, 21 diagnosed pre-NBS and 33 by NBS was, respectively, 5.4% (95% confidence interval [CI]: 4.0-8.3) and 12.6% (95% CI: 10.7-17.7; P < 0.001) of the reference mean. The median LC-FAO flux was 33.2% (95% CI: 22.8-48.3) and 41% (95% CI: 40.8-68; P < 0.05) of the control mean, respectively. Clinical characteristics in 23 pre-NBS and 37 NBS patients revealed hypoglycemic events in 12 vs 2 patients, cardiomyopathy in 5 vs 4 patients and myopathy in 14 vs 3 patients. All patients with LC-FAO flux <10% developed symptoms. Of the patients with LC-FAO flux >10% 7 out of 12 diagnosed pre-NBS vs none by NBS experienced hypoglycemic events. NBS has a clear beneficial effect on the prevention of hypoglycemic events in patients with some residual enzyme activity, but does not prevent hypoglycemia nor cardiac complications in patients with very low residual enzyme activity. The effect of NBS on prevalence and prevention of myopathy-related complications remains unclear.

13.
Mitochondrion ; 47: 298-308, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-30802674

RESUMO

Hyperammonemia is an important contributing factor to hepatic encephalopathy in end-stage liver failure patients. Therefore reducing hyperammonemia is a requisite of bioartificial liver support (BAL). Ammonia elimination by human liver HepaRG cells occurs predominantly through reversible fixation into amino acids, whereas the irreversible conversion into urea is limited. Compared to human liver, the expression and activity of the three urea cycle (UC) enzymes carbamoyl-phosphate synthase1 (CPS1), ornithine transcarbamoylase (OTC) and arginase1, are low. To improve HepaRG cells as BAL biocomponent, its rate limiting factor of the UC was determined under two culture conditions: static and dynamic medium flow (DMF) achieved by shaking. HepaRG cells increasingly converted escalating arginine doses into urea, indicating that arginase activity is not limiting ureagenesis. Neither was OTC activity, as a stable HepaRG line overexpressing OTC exhibited a 90- and 15.7-fold upregulation of OTC transcript and activity levels, without improvement in ureagenesis. However, a stable HepaRG line overexpressing CPS1 showed increased mitochondrial stress and reduced hepatic differentiation without promotion of the CPS1 transcript level or ureagenesis under static-culturing conditions, yet, it exhibited a 4.3-fold increased ureagenesis under DMF. This was associated with increased CPS1 transcript and activity levels amounting to >2-fold, increased mitochondrial abundance and hepatic differentiation. Unexpectedly, the transcript levels of several other UC genes increased up to 6.8-fold. We conclude that ureagenesis can be improved in HepaRG cells by CPS1 overexpression, however, only in combination with DMF-culturing, suggesting that both the low CPS1 level and static-culturing, possibly due to insufficient mitochondria, are limiting UC.

14.
J Inherit Metab Dis ; 42(2): 197-208, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30723938

RESUMO

The laboratory diagnosis of inborn errors of metabolism has been revolutionized in recent years, thanks to the amazing developments in the field of DNA sequencing including whole exome and whole genome sequencing (WES and WGS). Interpretation of the results coming from WES and/or WGS analysis is definitely not trivial especially since the biological relevance of many of the variants identified by WES and/or WGS, have not been tested experimentally and prediction programs like POLYPHEN-2 and SIFT are far from perfect. Correct interpretation of WES and/or WGS results can only be achieved by performing functional studies at multiple levels (different metabolomics platforms, enzymology, in vitro and in vivo flux analysis), often requires studies in model organisms like zebra fish, Caenorhabditis elegans, Saccharomyces cerevisiae, mutant mice and others, and also requires the input of many different disciplines to make this Translational Metabolism approach effective.

15.
J Inherit Metab Dis ; 42(2): 303-312, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30793331

RESUMO

INTRODUCTION: Currently, no therapies are available for Zellweger spectrum disorders (ZSDs), a group of genetic metabolic disorders characterised by a deficiency of functional peroxisomes. In a previous study, we showed that oral cholic acid (CA) treatment can suppress bile acid synthesis in ZSD patients and, thereby, decrease plasma levels of toxic C27 -bile acid intermediates, one of the biochemical abnormalities in these patients. However, no effect on clinically relevant outcome measures could be observed after 9 months of CA treatment. It was noted that, in patients with advanced liver disease, caution is needed because of possible hepatotoxicity. METHODS: An extension study of the previously conducted pretest-posttest design study was conducted including 17 patients with a ZSD. All patients received oral CA for an additional period of 12 months, encompassing a total of 21 months of treatment. Multiple clinically relevant parameters and markers for bile acid synthesis were assessed after 15 and 21 months of treatment. RESULTS: Bile acid synthesis was still suppressed after 21 months of CA treatment, accompanied with reduced levels of C27 -bile acid intermediates in plasma. These levels significantly increased again after discontinuation of CA. No significant changes were found in liver tests, liver elasticity, coagulation parameters, fat-soluble vitamin levels or body weight. CONCLUSIONS: Although CA treatment did lead to reduced levels of toxic C27 -bile acid intermediates in ZSD patients without severe liver fibrosis or cirrhosis, no improvement of clinically relevant parameters was observed after 21 months of treatment. We discuss the implications for CA therapy in ZSD based on these results.

16.
Biochim Biophys Acta Mol Cell Biol Lipids ; 1864(5): 654-661, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30731133

RESUMO

Previous studies have shown that the cardiolipin (CL)-deficient yeast mutant, crd1Δ, has decreased levels of acetyl-CoA and decreased activities of the TCA cycle enzymes aconitase and succinate dehydrogenase. These biochemical phenotypes are expected to lead to defective TCA cycle function. In this study, we report that signaling and anaplerotic metabolic pathways that supplement defects in the TCA cycle are essential in crd1Δ mutant cells. The crd1Δ mutant is synthetically lethal with mutants in the TCA cycle, retrograde (RTG) pathway, glyoxylate cycle, and pyruvate carboxylase 1. Glutamate levels were decreased, and the mutant exhibited glutamate auxotrophy. Glyoxylate cycle genes were up-regulated, and the levels of glyoxylate metabolites succinate and citrate were increased in crd1Δ. Import of acetyl-CoA from the cytosol into mitochondria is essential in crd1Δ, as deletion of the carnitine-acetylcarnitine translocase led to lethality in the CL mutant. ß-oxidation was functional in the mutant, and oleate supplementation rescued growth defects. These findings suggest that TCA cycle deficiency caused by the absence of CL necessitates activation of anaplerotic pathways to replenish acetyl-CoA and TCA cycle intermediates. Implications for Barth syndrome, a genetic disorder of CL metabolism, are discussed.


Assuntos
Cardiolipinas/genética , Ciclo do Ácido Cítrico , Regulação Fúngica da Expressão Gênica , Proteínas de Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/genética , Acetilcoenzima A/genética , Acetilcoenzima A/metabolismo , Cardiolipinas/metabolismo , Deleção de Genes , Glioxilatos/metabolismo , Mitocôndrias/genética , Mitocôndrias/metabolismo , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo
17.
Brain ; 142(3): 542-559, 2019 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-30668673

RESUMO

Biallelic pathogenic variants in PLPBP (formerly called PROSC) have recently been shown to cause a novel form of vitamin B6-dependent epilepsy, the pathophysiological basis of which is poorly understood. When left untreated, the disease can progress to status epilepticus and death in infancy. Here we present 12 previously undescribed patients and six novel pathogenic variants in PLPBP. Suspected clinical diagnoses prior to identification of PLPBP variants included mitochondrial encephalopathy (two patients), folinic acid-responsive epilepsy (one patient) and a movement disorder compatible with AADC deficiency (one patient). The encoded protein, PLPHP is believed to be crucial for B6 homeostasis. We modelled the pathogenicity of the variants and developed a clinical severity scoring system. The most severe phenotypes were associated with variants leading to loss of function of PLPBP or significantly affecting protein stability/PLP-binding. To explore the pathophysiology of this disease further, we developed the first zebrafish model of PLPHP deficiency using CRISPR/Cas9. Our model recapitulates the disease, with plpbp-/- larvae showing behavioural, biochemical, and electrophysiological signs of seizure activity by 10 days post-fertilization and early death by 16 days post-fertilization. Treatment with pyridoxine significantly improved the epileptic phenotype and extended lifespan in plpbp-/- animals. Larvae had disruptions in amino acid metabolism as well as GABA and catecholamine biosynthesis, indicating impairment of PLP-dependent enzymatic activities. Using mass spectrometry, we observed significant B6 vitamer level changes in plpbp-/- zebrafish, patient fibroblasts and PLPHP-deficient HEK293 cells. Additional studies in human cells and yeast provide the first empirical evidence that PLPHP is localized in mitochondria and may play a role in mitochondrial metabolism. These models provide new insights into disease mechanisms and can serve as a platform for drug discovery.

18.
Mol Genet Metab ; 126(1): 43-52, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30470562

RESUMO

Succinate-CoA ligase (SUCL) is a heterodimer consisting of an alpha subunit encoded by SUCLG1, and a beta subunit encoded by either SUCLA2 or SUCLG2 catalyzing an ATP- or GTP-forming reaction, respectively, in the mitochondrial matrix. The deficiency of this enzyme represents an encephalomyopathic form of mtDNA depletion syndromes. We describe the fatal clinical course of a female patient with a pathogenic mutation in SUCLG1 (c.626C > A, p.Ala209Glu) heterozygous at the genomic DNA level, but homozygous at the transcriptional level. The patient exhibited early-onset neurometabolic abnormality culminating in severe brain atrophy and dystonia leading to death by the age of 3.5 years. Urine and plasma metabolite profiling was consistent with SUCL deficiency which was confirmed by enzyme analysis and lack of mitochondrial substrate-level phosphorylation (mSLP) in skin fibroblasts. Oxygen consumption- but not extracellular acidification rates were altered only when using glutamine as a substrate, and this was associated with mild mtDNA depletion and no changes in ETC activities. Immunoblot analysis revealed no detectable levels of SUCLG1, while SUCLA2 and SUCLG2 protein expressions were largely reduced. Confocal imaging of triple immunocytochemistry of skin fibroblasts showed that SUCLG2 co-localized only partially with the mitochondrial network which otherwise exhibited an increase in fragmentation compared to control cells. Our results outline the catastrophic consequences of the mutated SUCLG1 leading to strongly reduced SUCL activity, mSLP impairment, mislocalization of SUCLG2, morphological alterations in mitochondria and clinically to a severe neurometabolic disease, but in the absence of changes in mtDNA levels or respiratory complex activities.


Assuntos
Mitocôndrias/patologia , Doenças Mitocondriais/diagnóstico , Mutação , Succinato-CoA Ligases/genética , Pré-Escolar , DNA Mitocondrial/genética , Evolução Fatal , Feminino , Heterozigoto , Homozigoto , Humanos , Mitocôndrias/metabolismo , Fosforilação , Succinato-CoA Ligases/sangue , Succinato-CoA Ligases/urina
19.
Subcell Biochem ; 89: 345-365, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30378031

RESUMO

Peroxisomes catalyze a number of essential metabolic functions and impairments in any of these are usually associated with major clinical signs and symptoms. In contrast to mitochondria which are autonomous organelles that can catalyze the degradation of fatty acids, certain amino acids and other compounds all by themselves, peroxisomes are non-autonomous organelles which are highly dependent on the interaction with other organelles and compartments to fulfill their role in metabolism. This includes mitochondria, the endoplasmic reticulum, lysosomes, and the cytosol. In this paper we will discuss the central role of peroxisomes in different metabolic interaction networks in humans, including fatty acid oxidation, ether phospholipid biosynthesis, bile acid synthesis, fatty acid alpha-oxidation and glyoxylate metabolism.


Assuntos
Redes e Vias Metabólicas , Peroxissomos/metabolismo , Ácidos e Sais Biliares/biossíntese , Ácidos Graxos/metabolismo , Glioxilatos/metabolismo , Humanos , Oxirredução
20.
Eur J Hum Genet ; 26(12): 1752-1758, 2018 12.
Artigo em Inglês | MEDLINE | ID: mdl-30089828

RESUMO

Pontocerebellar hypoplasia (PCH) is a heterogeneous neurodegenerative disorder with a prenatal onset. Using whole-exome sequencing, we identified variants in the gene Coenzyme A (CoA) synthase (COASY) gene, an enzyme essential in CoA synthesis, in four individuals from two families with PCH, prenatal onset microcephaly, and arthrogryposis. In family 1, compound heterozygous variants were identified in COASY: c.[1549_1550delAG]; [1486-3 C>G]. In family 2, all three affected siblings were homozygous for the c.1486-3 C>G variant. In both families, the variants segregated with the phenotype. RNA analysis showed that the c.1486-3 C>G variant leads to skipping of exon 7 with partial retention of intron 7, disturbing the reading frame and resulting in a premature stop codon (p.(Ala496Ilefs*20)). No CoA synthase protein was detected in patient cells by immunoblot analysis and CoA synthase activity was virtually absent. Partial CoA synthase defects were previously described as a cause of COASY Protein-Associated Neurodegeneration (CoPAN), a type of Neurodegeneration and Brain Iron Accumulation (NBIA). Here we demonstrate that near complete loss of function variants in COASY are associated with lethal PCH and arthrogryposis.


Assuntos
Artrogripose/genética , Doenças Cerebelares/genética , Mutação com Perda de Função , Microcefalia/genética , Transferases/genética , Feto Abortado/anormalidades , Artrogripose/patologia , Células Cultivadas , Doenças Cerebelares/patologia , Humanos , Recém-Nascido , Masculino , Microcefalia/patologia , Síndrome , Transferases/metabolismo
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA