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J G Okun

Publications and source records attributed to J G Okun.

18 recordsLinked to original sources
PubMed2007

Guideline for the diagnosis and management of glutaryl-CoA dehydrogenase deficiency (glutaric aciduria type I).

Glutaryl-CoA dehydrogenase (GCDH) deficiency is an autosomal recessive disease with an estimated overall prevalence of 1 in 100 000 newborns. Biochemically, the disease is characterized by accumulation of glutaric acid, 3-hydroxyglutaric acid, glutaconic acid, and glutarylcarnitine, which can be detected by gas chromatography-mass spectrometry of organic acids or tandem mass spectrometry of acylcarnitines. Clinically, the disease course is usually determined by acute encephalopathic crises precipitated by infectious diseases, immunizations, and surgery during infancy or childhood. The characteristic neurological sequel is acute striatal injury and, subsequently, dystonia. During the last three decades attempts have been made to establish and optimize therapy for GCDH deficiency. Maintenance treatment consisting of a diet combined with oral supplementation of L: -carnitine, and an intensified emergency treatment during acute episodes of intercurrent illness have been applied to the majority of patients. This treatment strategy has significantly reduced the frequency of acute encephalopathic crises in early-diagnosed patients. Therefore, GCDH deficiency is now considered to be a treatable condition. However, significant differences exist in the diagnostic procedure and management of affected patients so that there is a wide variation of the outcome, in particular of pre-symptomatically diagnosed patients. At this time of rapid expansion of neonatal screening for GCDH deficiency, the major aim of this guideline is to re-assess the common practice and to formulate recommendations for diagnosis and management of GCDH deficiency based on the best available evidence.

Child↗
PubMed2005

Measurement of bile acid CoA esters by high-performance liquid chromatography-electrospray ionisation tandem mass spectrometry (HPLC-ESI-MS/MS).

The novel and rapid assay presented here combines high-performance liquid chromatography and electrospray ionisation tandem mass spectrometry (HPLC-ESI-MS/MS) to directly measure and quantify the CoA esters of 3alpha,7alpha,12alpha-trihydroxy- and 3alpha,7alpha-dihydroxy-5beta-cholestan-26-oic acid (THCA and DHCA). The latter are converted inside peroxisomes to the primary bile acids, cholic and chenodeoxycholic acids, respectively. Prior to MS/MS, esters were separated by reversed-phase HPLC on a C(18) column using an isocratic mobile phase (acetonitrile/water/2-propanol) and subsequently detected by multiple reaction monitoring. For quantification, the CoA ester of deuterium-labelled 3alpha,7alpha,12alpha-trihydroxy-5beta-cholan-24-oic acid (d(4)-CA) was used as internal standard. To complete an assay took less than 8 min. To verify the validity of the assay, the effect of peroxisomal proteins on the efficacy of extraction of the CoA esters was tested. To this end, variable amounts of the CoA esters were spiked with a fixed amount of either intact peroxisomes or peroxisomal matrix proteins and then extracted using a solid-phase extraction system. The CoA esters could be reproducibly recovered in the range of 0.1-4 micromol l(-1) (linear correlation coefficient R(2) > 0.99), with a detection limit of 0.1 micromol l(-1). In summary, electrospray ionization tandem mass spectrometry combined with HPLC as described here proved to be a rapid and versatile technique for the determination of bile acid CoA esters in a mixture with peroxisomal proteins. This suggests this technique to become a valuable tool in studies dealing with the multi-step biosynthesis of bile acids and its disturbances in disorders like the Zellweger syndrome.

Acyl Coenzyme A↗
PubMed2005

Methylmalonic acid--an endogenous toxin?

Methylmalonic acid was previously considered as major neurotoxin in methylmalonic acidurias. In contrast, recent studies support the notion that other metabolites deriving from propionyl-coenzyme A, inducing synergistic inhibition of mitochondrial energy metabolism, are more important than methylmalonic acid to understand the neuropathogenesis of this disease. However, it is not yet known whether methylmalonic acid is involved in the induction of other organ manifestations in this disease, such as chronic renal failure.

Energy Metabolism↗
PubMed2005

Massive insulin secretion in response to anaerobic exercise in exercise-induced hyperinsulinism.

Exercise-induced hyperinsulinism (EIHI) is a recently described entity characterised by recurrent episodes of hypoglycaemia induced by physical exercise. The index patient for this disorder and a matched control were subjected to aerobic and anaerobic exercise tests on a cycle ergometer. Aerobic exercise was performed at an intensity of 60% of the respective 4 mmol/l lactate threshold (40 min). Anaerobic exercise with an intensity corresponding to 130% VO2max lead to exertion within 2-3 min and elicited comparable maximal lactate levels in both subjects (10-11 mmol/l). The patient experienced a massive increase in insulin from 34 to 649 mU/l after the anaerobic test, and a lower increase in insulin from 27 to 79 mU/l during the aerobic test. Insulin concentration remained unchanged during both tests in the control. Epinephrine increased in the EIHI patient, which was probably a counterregulatory response to hypoglycaemia. The activity of lactate dehydrogenase of the index patient in isolated leukocytes as well as the response to inhibition of oxamate was normal. The hypothesis of abnormal transport or metabolism of lactate/pyruvate in the beta-cells of patients with EIHI was further supported by the parallel increase of lactate and insulin in this study elicited in particular by anaerobic exercise.

Adult↗
PubMed2004

Excitotoxicity and bioenergetics in glutaryl-CoA dehydrogenase deficiency.

Glutaryl-CoA dehydrogenase deficiency is an inherited organic acid disorder with predominantly neurological presentation. The biochemical hallmark of this disease is an accumulation and enhanced urinary excretion of two key organic acids, glutaric acid and 3-hydroxyglutaric acid. If untreated, acute striatal damage is often precipitated by febrile illnesses during a vulnerable period of brain development in infancy or early childhood, resulting in a dystonic dyskinetic movement disorder. 3-hydroxyglutaric and glutaric acids are structurally similar to glutamate, the main excitatory amino acid of the human brain, and are considered to play an important role in the pathophysiology of this disease. 3-hydroxyglutaric acid induces excitotoxic cell damage specifically via activation of N-methyl-D-aspartate receptors. It has also been suggested that secondary amplification loops potentiate the neurotoxic properties of these organic acids. Probable mechanisms for this effect include cytokine-stimulated NO production, a decrease in energy metabolism, and reduction of cellular creatine phosphate levels. Finally, maturation-dependent changes in the expression of neuronal glutamate receptors may affect the vulnerability of the immature brain to excitotoxic cell damage in this disease.

Amino Acid Metabolism, Inborn Errors↗
PubMed2004

Animal models for glutaryl-CoA dehydrogenase deficiency.

In vitro studies suggest that excitotoxic cell damage is an underlying mechanism for the acute striatal damage in glutaryl-CoA dehydrogenase (GCDH) deficiency. It is believed to result from an imbalance of glutamatergic and GABAergic neurotransmission induced by the accumulating organic acids 3-hydroxyglutaric acid (3-OH-GA) and to a lesser extent glutaric acid (GA). Stereotaxic administration of 3-OH-GA and GA into the rat striatum have confirmed these results, but may not truly represent the effect of chronic exposure to these compounds. In an attempt to better understand the pathophysiology of GCDH deficiency in vivo , two animal models have been utilized. A mouse that lacks GCDH activity in all tissues was generated by gene targeting in embryonic stem cells. These animals develop the characteristic biochemical phenotype of the human disease. Pathologically, these mice have a diffuse spongiform myelinopathy similar to that in human patients; however, there is no evidence for acute striatal damage or sensitivity to acute encephalopathy induced by catabolism or inflammatory cytokines. A naturally occurring animal model, the fruit-eating bat Rousettus aegypticus, lacks hepatic and renal GCDH activity, but retains cerebral enzyme activity. Like the mouse, these bats develop the characteristic biochemical phenotype of glutaryl-CoA dehydrogenase deficiency, but lack overt neurological symptoms such as dystonia. It is not known whether they also develop the spongiform myelinopathy seen in the Gcdh-deficient mice. Otherwise, these constellations would suggest that cerebral GCDH deficiency is responsible for the development of neuronal damage. The lack of striatal damage in these two rodent models may also be related to species differences. However, they also highlight our lack of a comprehensive understanding of additional factors that might modulate the susceptibiliy of neurons to accumulating 3-OH-GA and GA in GCDH deficiency. Unravelling these mechanisms may be the key to understanding the pathophysiology of this unique disease and to the development of neuroprotective strategies.

Amino Acid Metabolism, Inborn Errors↗
PubMed2004

Challenges for basic research in glutaryl-CoA dehydrogenase deficiency.

During the last decades, efforts have been made to elucidate the complex mechanisms underlying neuronal damage in glutaryl-CoA dehydrogenase deficiency. A combination of in vitro and in vivo investigations have facilitated the development of several hypotheses, including the probable pathogenic role of accumulating glutaric acid and 3-hydroxyglutaric acid. However, there are still many shortcomings that limit an evidence-based approach to treating this inborn error of metabolism. Major future goals should include generation of a suitable animal model for acute striatal necrosis, investigation of the formation, distribution and exact intra- and extracellular concentrations of accumulating metabolites, a deeper understanding of striatal vulnerability, and systematic investigation of effects on cerebral gene expression during development and of the modulatory role of inflammatory cytokines.

Amino Acid Metabolism, Inborn Errors↗
PubMed2004

Maintenance treatment of glutaryl-CoA dehydrogenase deficiency.

This paper summarizes the published experience as well as results of the 3rd International Workshop on Glutaryl-CoA Dehydrogenase Deficiency held in October 2003 in Heidelberg, Germany, on the topic treatment of patients with glutaryl-CoA dehydrogenase (GCDH) deficiency. So far no international recommendation for treatment of GCDH deficiency exists. Such an approach is hampered by several facts, namely the lack of an in-depth understanding of the pathophysiology of the disease, the lack of prospective studies, including the evaluation of drug monotherapy, and lack of objective documentation of clinical changes (e.g. video documentation) during pharmacotherapy.

Amino Acid Metabolism, Inborn Errors↗
PubMed2004

Looking forward--an evidence-based approach to glutaryl-CoA dehydrogenase deficiency.

Three decades after the first description of glutaryl-CoA dehydrogenase deficiency, major progress has been achieved in the prevention of acute striatal necrosis and neurological sequelae in affected children, if diagnosis is made early and treatment is started before manifestation of acute encephalopathic crises. However, all concepts for diagnostic work-up, monitoring, and treatment are solely experience-based, and 10-35% of early-diagnosed children do not or only incompletely benefit from the current management. They still develop neurological deterioration and sequelae despite early implementation of dietary treatment, carnitine supplementation and emergency treatment during acute intercurrent illnesses. International efforts should be made to move management of affected children from experience-based to evidence-based medicine. Major tools for this optimization are the establishment of an international patients' database, the implementation of an international prospective clinical study, and the development of international guidelines for diagnostic work-up, monitoring and therapy.

Amino Acid Metabolism, Inborn Errors↗
PubMed2004

S-Acetylglutathione normalizes intracellular glutathione content in cultured fibroblasts from patients with glutathione synthetase deficiency.

Glutathione synthetase deficiency is an autosomal recessive inherited metabolic defect in the gamma-glutamyl cycle. Decreased intracellular glutathione levels are one of the characteristic biochemical features. In this study we show that addition of S-acetylglutathione to the medium raised intracellular glutathione content in cultured fibroblasts from patients with glutathione synthetase deficiency. This has implications for the treatment of patients with this inborn error of metabolism.

Cells, Cultured↗
PubMed2003

Glutaryl-CoA dehydrogenase deficiency: region-specific analysis of organic acids and acylcarnitines in post mortem brain predicts vulnerability of the putamen.

The neurometabolic disorder glutaryl-CoA dehydrogenase (GCDH) deficiency is biochemically characterised by an accumulation of the marker metabolites 3-hydroxyglutaric acid, glutaric acid, and glutarylcarnitine. If untreated, the disease is complicated by acute encephalopathic crises, resulting in neurodegeneration of vulnerable brain regions, in particular the putamen. 3-hydroxyglutaric acid is considered the major neurotoxin in this disease. There are only preliminary data concerning glutaric acid concentrations in the brains of affected children and the distribution of 3-hydroxyglutaric acid and glutarylcarnitine has not been described. In the present study, we investigated post mortem the distribution of 3-hydroxyglutaric and glutaric acids as well as glutarylcarnitine in 14 different brain regions, internal organs, and body fluids (urine, plasma, cerebrospinal fluid) in a 14-year-old boy. 3-Hydroxyglutaric acid showed the highest concentration (62 nmol/g protein) in the putamen among all brain areas investigated. The glutarylcarnitine concentration was also highest in the putamen (7.1 nmol/g protein). We suggest that the regional-specific differences in the relative concentrations of 3-hydroxyglutaric acid contribute to the pattern of neuronal damage in this disease. These results provide an explanatory basis for the high vulnerability of the putamen in this disease, adding to the strong corticostriatal glutamatergic input into the putamen and the high excitotoxic susceptibility of neostriatal medium spiny neurons.

Acidosis↗
PubMed2001

3-Ureidopropionate contributes to the neuropathology of 3-ureidopropionase deficiency and severe propionic aciduria: a hypothesis.

3-Ureidopropionate (3-UPA) is a physiologic metabolite in pyrimidine degradation. Pathological accumulation of 3-UPA in body fluids is found in 3-ureidopropionase deficiency and severe forms of propionic aciduria. Both diseases clinically present with a severe neuropathology involving gray and white matter as well as with a dystonic dyskinetic movement disorder. To date nothing is known about the toxic nature of this metabolite. The aim of the present study was to elucidate whether 3-UPA may act as endogenous neurotoxin. Exposure of cultured chick neurons to 3-UPA induced a concentration- and time-dependent neurodegeneration. Neuronal damage was reduced by the antioxidant alpha-tocopherol and the N-methyl-D-aspartate (NMDA) receptor antagonist MK-801. In contrast, the non-NMDA receptor antagonist CNQX, the metabotropic glutamate receptor antagonist L-AP3, and succinate showed no protective effect. Furthermore, 3-UPA elicited an increased production of reactive oxygen species followed by a delayed increase in intracellular calcium concentrations. Activity measurement of single respiratory chain complexes I-V revealed an inhibition of complex V activity, but not of the electron-transferring complexes I-IV by 3-UPA. In contrast, 3-UPA did not affect the mitochondrial beta-oxidation of fatty acids. In conclusion, our results provide strong evidence that 3-UPA acts as endogenous neurotoxin via inhibition of mitochondrial energy metabolism, resulting in the initiation of secondary, energy-dependent excitotoxic mechanisms.

Animals↗
PubMed2001

Three molecules of ubiquinone bind specifically to mitochondrial cytochrome bc1 complex.

Bifurcated electron flow to high potential "Rieske" iron-sulfur cluster and low potential heme b(L) is crucial for respiratory energy conservation by the cytochrome bc(1) complex. The chemistry of ubiquinol oxidation has to ensure the thermodynamically unfavorable electron transfer to heme b(L). To resolve a central controversy about the number of ubiquinol molecules involved in this reaction, we used high resolution magic-angle-spinning nuclear magnetic resonance experiments to show that two out of three n-decyl-ubiquinones bind at the ubiquinol oxidation center of the complex. This substantiates a proposed mechanism in which a charge transfer between a ubiquinol/ubiquinone pair explains the bifurcation of electron flow.

Animals↗
PubMed2000

Binding of detergents and inhibitors to bovine complex I - a novel purification procedure for bovine complex I retaining full inhibitor sensitivity.

Mitochondrial complex I exhibits some peculiar and poorly understood features regarding the effects of detergents on activity and sensitivity to hydrophobic inhibitors that are not seen with other membrane complexes using ubiquinone as a substrate. Therefore, we investigated the interaction of complex I from bovine heart mitochondria with different types of detergents by monitoring activity, degree of inhibition and inhibitor binding in the presence of increasing concentrations of detergent. It is shown that apart from their nature as solubilizing and delipidating agents the polyoxyethylene-ether detergents Triton X-100, Brij-35 and Thesit act as specific inhibitors of complex I and compete with classical complex I inhibitors for a common binding domain. These findings were used to develop a novel large-scale chromatographic procedure for isolation of inhibitor-sensitive NADH:ubiquinone oxidoreductase (complex I) from bovine heart mitochondria. The enzyme was purified by selective solubilization in Triton X-100 and subsequent hydroxylapatite, ion-exchange and gel-exclusion chromatography. By switching detergents from Triton X-100 to dodecylmaltoside after hydroxylapatite chromatography the procedure yields highly pure, monodisperse and fully inhibitor-sensitive enzyme.

Amines↗
PubMed1999

Three classes of inhibitors share a common binding domain in mitochondrial complex I (NADH:ubiquinone oxidoreductase).

We have developed two independent methods to measure equilibrium binding of inhibitors to membrane-bound and partially purified NADH:ubiquinone oxidoreductase (complex I) to characterize the binding sites for the great variety of hydrophobic compounds acting on this large and complicated enzyme. Taking advantage of a partial quench of fluorescence upon binding of the fenazaquin-type inhibitor 2-decyl-4-quinazolinyl amine to complex I in bovine submitochondrial particles, we determined a Kd of 17 +/- 3 nM and one binding site per complex I. Equilibrium binding studies with [3H]dihydrorotenone and the aminopyrimidine [3H]AE F119209 (4(cis-4-[3H]isopropyl cyclohexylamino)-5-chloro-6-ethyl pyrimidine) using partially purified complex I from Musca domestica exhibited little unspecific binding and allowed reliable determination of dissociation constants. Competition experiments consistently demonstrated that all tested hydrophobic inhibitors of complex I share a common binding domain with partially overlapping sites. Although the rotenone site overlaps with both the piericidin A and the capsaicin site, the latter two sites do not overlap. This is in contrast to the interpretation of enzyme kinetics that have previously been used to define three classes of complex I inhibitors. The existence of only one large inhibitor binding pocket in the hydrophobic part of complex I is discussed in the light of possible mechanisms of proton translocation.

Animals↗
PubMed1999

A single external enzyme confers alternative NADH:ubiquinone oxidoreductase activity in Yarrowia lipolytica.

NADH:ubiquinone oxidoreductases catalyse the first step within the diverse pathways of mitochondrial NADH oxidation. In addition to the energy-conserving form commonly called complex I, fungi and plants contain much simpler alternative NADH:ubiquinone oxido-reductases that catalyze the same reaction but do not translocate protons across the inner mitochondrial membrane. Little is known about the distribution and function of these enzymes. We have identified YLNDH2 as the only gene encoding an alternative NADH:ubiquinone oxidoreductase (NDH2) in the obligate aerobic yeast Yarrowia lipolytica. Cells carrying a deletion of YLNDH2 were fully viable; full inhibition by piericidin A indicated that complex I activity was the sole NADH:ubiquinone oxidoreductase activity left in the deletion strains. Studies with intact mitochondria revealed that NDH2 in Y. lipolytica is oriented towards the external face of the mitochondrial inner membrane. This is in contrast to the situation seen in Saccharomyces cerevisiae, Neurospora crassa and in green plants, where internal alternative NADH:ubiquinone oxidoreductases have been reported. Phylogenetic analysis of known NADH:ubiquinone oxidoreductases suggests that during evolution conversion of an ancestral external alternative NADH:ubiquinone oxidoreductase to an internal enzyme may have paved the way for the loss of complex I in fermenting yeasts like S. cerevisiae.

Amino Acid Sequence↗
PubMed1997

Role of deprotonation events in ubihydroquinone:cytochrome c oxidoreductase from bovine heart and yeast mitochondria.

The pH dependence of bovine and yeast cytochrome bc1 complex catalyzing electron transfer from ubi- and plastohydroquinone to cytochrome c have been analyzed. The pH dependence of the steady-state rate was found to be governed by two protonable groups, one of which (pK approximately 6.6) has to be deprotonated while the other (pK approximately 9.2) has to be protonated to allow catalysis. Using ubideuteroquinone instead of ubihydroquinone as a substrate resulted in 1.4- and 1.7-fold lower steady-state rates for the bovine and yeast enzymes, respectively. The activation energy at pH 8.0 was 33 kJ/mol for the bovine and 44 kJ/mol for the yeast enzyme and exhibited a linear decrease between pH 5.4 and 9.2. For ubihydroquinone the slope was very close to a value of -5.7 kJ/mol expected if the activation energy depended on a single deprotonation event. When plastohydroquinone was used instead, the slope more than doubled, indicating that a second deprotonation contributed to the activation barrier with this nonphysiological substrate. In contrast to previous kinetic models for the cytochrome bc1 complex, which propose that the activation barrier is associated with the formation of ubisemiquinone at the ubihydroquinone oxidation center, our results strongly suggest that the best approximation of the transition state is the singly deprotonated form of ubihydroquinone. This supports the recently proposed proton-gated charge transfer mechanism, which has control of catalysis by the first deprotonation of ubihydroquinone as one of its key features [Brandt, U. (1996) FEBS Lett. 387, 1-6]. All results reported here can be rationalized in a straightforward way based on other aspects of the same hypothesis.

Animals↗