Showing papers on "Urea cycle published in 2006"

Valproate-induced hyperammonemic encephalopathy

Abstract: Valproate-induced hyperammonemic encephalopathy (VHE) is an unusual complication characterized by a decreasing level of consciousness, focal neurological deficits, cognitive slowing, vomiting, drowsiness, and lethargy. We have thoroughly reviewed the predisposing factors and their screening, the biochemical and physiopathological mechanisms involved, the different treatments described, and those that are being investigated. Etiopathogenesis is not completely understood, although hyperammonemia has been postulated as the main cause of the clinical syndrome. The increase in serum ammonium level is due to several mechanisms, the most important one appearing to be the inhibition of carbamoylphosphate synthetase-I, the enzyme that begins the urea cycle. Polytherapy with several drugs, such as phenobarbital and topiramate, seems to contribute to hyperammonemia. Hyperammonemia leads to an increase in the glutamine level in the brain, which produces astrocyte swelling and cerebral edema. There are several studies that suggest that treatment with supplements of carnitine can lead to an early favorable clinical response due to the probable carnitine deficiency induced by a valproate (VPA) treatment. Development of the progressive confusional syndrome, associated with an increase in seizure frequency after VPA treatment onset, obliges us to rule out VHE by screening for blood ammonium levels and the existence of urea cycle enzyme deficiency, such as ornithine carbamoyltransferase deficiency. Electroencephalography (EEG) is characterized by signs of severe encephalopathy with continuous generalized slowing, a predominance of theta and delta activity, occasional bursts of frontal intermittent rhythmic delta activity, and triphasic waves. These EEG findings, as well as clinical manifestations and hyperammonemia, tend to normalize after VPA withdrawal.

Molecular Mechanisms of Urea Transport in Plants

Abstract: Urea is a soil nitrogen form available to plant roots and a secondary nitrogen metabolite liberated in plant cells. Based on growth complementation of yeast mutants and “in-silico analysis”, two plant families have been identified and partially characterized that mediate membrane transport of urea in heterologous expression systems. AtDUR3 is a single Arabidopsis gene belonging to the sodium solute symporter family that cotransports urea with protons at high affinity, while members of the tonoplast intrinsic protein (TIP) subfamily of aquaporins transport urea in a channel-like manner. The following review summarizes current knowledge on the membrane localization, energetization and regulation of these two types of urea transporters and discusses their possible physiological roles in planta.

Clinical, biochemical, and molecular spectrum of hyperargininemia due to arginase I deficiency

Abstract: The urea cycle consists of six consecutive enzymatic reactions that convert waste nitrogen into urea. Urea cycle disorders are a group of inborn errors of hepatic metabolism that often result in life threatening hyperammonemia and hyperglutaminemia. Deficiencies of all of the enzymes of the cycle have been described and although each specific disorder results in the accumulation of different precursors, hyperammonemia and hyperglutaminemia are common biochemical hallmarks of these disorders. Arginase is the enzyme involved in the last step of the urea cycle. It catalyzes the conversion of arginine to urea and ornithine. The latter reenters the mitochondrion to continue the cycle. Hyperargininemia is an autosomal recessive disorder caused by a defect in the arginase I enzyme. Unlike other urea cycle disorders, this condition is not generally associated with a hyperammonemic encephalopathy in the neonatal period. It typically presents later in childhood between 2 and 4 years of age with predominantly neurological features. If untreated, it progresses with gradual developmental regression. A favorable outcome can be achieved if dietary treatment and alternative pathway therapy are instituted early in the disease course. With this approach, further neurological deterioration is prevented and partial recovery of skills ensues. Early diagnosis of this disorder through newborn screening programs may lead to a better outcome. This review article summarizes the clinical characterization of this disorder; as well as its biochemical, enzymatic, and molecular features. Treatment, prenatal diagnosis and diagnosis through newborn screening are also discussed.

Sodium fluoroacetate poisoning.

Abstract: Sodium fluoroacetate was introduced as a rodenticide in the US in 1946. However, its considerable efficacy against target species is offset by comparable toxicity to other mammals and, to a lesser extent, birds and its use as a general rodenticide was therefore severely curtailed by 1990. Currently, sodium fluoroacetate is licensed in the US for use against coyotes, which prey on sheep and goats, and in Australia and New Zealand to kill unwanted introduced species. The extreme toxicity of fluoroacetate to mammals and insects stems from its similarity to acetate, which has a pivotal role in cellular metabolism. Fluoroacetate combines with coenzyme A (CoA-SH) to form fluoroacetyl CoA, which can substitute for acetyl CoA in the tricarboxylic acid cycle and reacts with citrate synthase to produce fluorocitrate, a metabolite of which then binds very tightly to aconitase, thereby halting the cycle. Many of the features of fluoroacetate poisoning are, therefore, largely direct and indirect consequences of impaired oxidative metabolism. Energy production is reduced and intermediates of the tricarboxylic acid cycle subsequent to citrate are depleted. Among these is oxoglutarate, a precursor of glutamate, which is not only an excitatory neurotransmitter in the CNS but is also required for efficient removal of ammonia via the urea cycle. Increased ammonia concentrations may contribute to the incidence of seizures. Glutamate is also required for glutamine synthesis and glutamine depletion has been observed in the brain of fluoroacetate-poisoned rodents. Reduced cellular oxidative metabolism contributes to a lactic acidosis. Inability to oxidise fatty acids via the tricarboxylic acid cycle leads to ketone body accumulation and worsening acidosis. Adenosine triphosphate (ATP) depletion results in inhibition of high energy-consuming reactions such as gluconeogenesis. Fluoroacetate poisoning is associated with citrate accumulation in several tissues, including the brain. Fluoride liberated from fluoroacetate, citrate and fluorocitrate are calcium chelators and there are both animal and clinical data to support hypocalcaemia as a mechanism of fluoroacetate toxicity. However, the available evidence suggests the fluoride component does not contribute. Acute poisoning with sodium fluoroacetate is uncommon. Ingestion is the major route by which poisoning occurs. Nausea, vomiting and abdominal pain are common within 1 hour of ingestion. Sweating, apprehension, confusion and agitation follow. Both supraventricular and ventricular arrhythmias have been reported and nonspecific ST- and T-wave changes are common, the QTc may be prolonged and hypotension may develop. Seizures are the main neurological feature. Coma may persist for several days. Although several possible antidotes have been investigated, they are of unproven value in humans. The immediate, and probably only, management of fluoroacetate poisoning is therefore supportive, including the correction of hypocalcaemia.

The dogfish shark (Squalus acanthias) increases both hepatic and extrahepatic ornithine urea cycle enzyme activities for nitrogen conservation after feeding.

Abstract: Urea not only is utilized as a major osmolyte in marine elasmobranchs but also constitutes their main nitrogenous waste. This study investigated the effect of feeding, and thus elevated nitrogen intake, on nitrogen metabolism in the Pacific spiny dogfish Squalus acanthias. We determined the activities of ornithine urea cycle (O‐UC) and related enzymes in liver and nonhepatic tissues. Carbamoyl phosphate synthetase III (the rate‐limiting enzyme of the O‐UC) activity in muscle is high compared with liver, and the activities in both tissues increased after feeding. The contribution of muscle to urea synthesis in the dogfish body appears to be much larger than that of liver when body mass is considered. Furthermore, enhanced activities of the O‐UC and related enzymes (glutamine synthetase, ornithine transcarbamoylase, arginase) were seen after feeding in both liver and muscle and were accompanied by delayed increases in plasma urea, trimethylamine oxide, total free amino acids, alanine, and chloride ...

The Wnt/β‐catenin pathway: master regulator of liver zonation?

Abstract: The liver contains two systems for the removal of ammonia—the urea cycle and the enzyme glutamine synthetase. These systems are expressed in a complementary fashion in two distinct populations of hepatocytes, referred to as periportal and perivenous cells. One of the unresolved problems in hepatology has been to elucidate the molecular mechanisms responsible for induction and maintenance of the cellular heterogeneity for ammonia detoxification. There is now a potential molecular explanation for the zonation of the urea cycle and glutamine synthetase based on the Wnt/β-catenin pathway[1]. BioEssays 28: 1072–1077, 2006. © 2006 Wiley Periodicals, Inc.

Hepatic encephalopathy and nitrogen metabolism

Abstract: Preface.- Foundation of the 'International Society of Hepatic Encephalopathy and Nitrogen Metabolism' (ISHEN).- I. Physiology and pathophysiology of astrocytes.- Water transport in the brain: basic concepts and astrocyte swelling.- Osmotic and Oxidative stress in hepatic encephalopathy.- NADPH oxidase is a major source of swelling-induced generation of reactive oxygen species in rat astrocytes.- Detoxification of hydrogen peroxide by astrocytes.- Glutamate release from astrocytes, impact on neuronal function.- Hepatic stellate cells and astrocytes both star-shaped but not next of kin.- Localization and function of the ATP -binding cassette C (ABCC/MRP) efflux pumps in human brain.- Glial mechanism of axomal growth protection from ammonia.- Selenoprotein P: a link between liver and brain.- II Pathogenesis of HE.- The peripheral benzodiazepine receptor and neurosteroids in the pathogenesis of hepatic encephalopathy and ammonia neurotoxicity.- Glutamatergic synaptic regulation deficit in liver failture: A review of molecular mechanisms.- Synaptic plasticity in animal models relevant for hepatic encephalopathy.- NMDA receptors in hyperammonemia and hepatic encephalopathy.- Prehepatic portal hypertension and mitochondrial dysfunction in brain hipppo-campus.- Nuclear magnetic resonance studies in experimental animal models of hepatic encephalopathy.- The role of inflammation in hepatic encephalopathy.- III. Brain imging in HE research.- Altered neural oscillations and synchronization: a pathophysiological hallmark of hepatic encephalopathy.- Neurotransmitter receptor alterations in hepatic encephalopathy.- Quantitiative T1 and water content mapping in hepatic encephalopathy.- Central nervous system involvement in Hepatitis C virus infection: what to measure?.- Localized MR-correlated spectroscopy using two spectral dimensions: Theoretical description and pilot investigation in hepatic encephalopathy.- IV. New aspects of nitrogen metabolism.- Urea cycle disorders.-The pathophysiology of citrin deficiency.- Hepatic encephalopathy in organic acidurias - Hyperammonemia and energy failure.- Glutamine synthetase deficiency in the human.- Hepatic dysfunction and encephalopathy in inborn errors of metabolism.- Amino acids and mTOR-dependent signalling.- Ceramide is a negative regulator of insulin action, nutrient uptake and protein synthesis in cultured rat skeletal muscle cells.- Ammonia transport in aquaporins - Molecular mechanicsm and clinical relevance.- Glutamine synthetase as a target of ss-catenin: New insights into hepatic heterogeneity.- Ammonia metabolism in liver cirrhosis.- V. Clinical assissment of HE.- Definition and assessment of low grade hepatic encephalopathy.- Hepatic encephalopathy in acute liver failure.- Quality of life in chronic liver diseases and in hepatic encephalopathy.- Hyponatremia: A risk factor of hepatic encephalopathy in cirrhosis.- Neuropsychological dysfunction in minimal hepatic encephalopathy: a review compared with own experience.- Computerised or paper and pencil psychometric tests for the assessment of hepatic encephalopathy?.- Neuropsychological basis of critical flicker frequency analysis (CFF).- Critical assessment of the PHES-test in patients with low grade hepatic encephalopathy.- VI. Treatment.- Evidence-based medicine and treatment of hepatic encephalopathy.- Brain acute liver failure edema in: Mechanisms and therapeutical options.- Hepatic encephalopathy and artificial liver support.- Prometheus(R) - A new extracorporeal system for the treatment of liver failure.- Modulation of intestinal flora for the treatment of hepatic encephalopahty in cirrhosis.- Prevention of post-TIPS hepatic encephalopathy.- Milestones in HE research.- Group picture.- Index

Interorgan ammonia, glutamate, and glutamine trafficking in pigs with acute liver failure

Abstract: Ammonia reduction is the target for therapy of hepatic encephalopathy, but lack of quantitative data about how the individual organs handle ammonia limits our ability to develop novel therapeutic strategies. The study aims were to evaluate interorgan ammonia metabolism quantitatively in a devascularized pig model of acute liver failure (ALF). Ammonia and amino acid fluxes were measured across the portal drained viscera (PDV), kidneys, hind leg, and lungs in ALF pigs. ALF pigs developed hyperammonemia and increased glutamine levels, whereas glutamate levels were decreased. PDV contributed to the hyperammonemic state mainly through increased shunting and not as a result of increased glutamine breakdown. The kidneys were quantitatively as important as PDV in systemic ammonia release, whereas muscle took up ammonia. Data suggest that the lungs are able to remove ammonia from the circulation during the initial stage of ALF. Our study provides new data supporting the concept of glutamate deficiency in a pig model of ALF. Furthermore, the kidneys are quantitatively as important as PDV in ammonia production, and the muscles play an important role in ammonia removal.

Amino acid, ammonia and urea concentrations in human pre-ovulatory ovarian follicular fluid

Abstract: BACKGROUND: This study aimed to determine amino acid (AA), ammonia and urea concentrations in human ovarian follicular fluid and to compare these concentrations with those in the circulation METHODS: Samples of pre-ovulatory follicular fluid and peripheral venous blood were obtained from 14 IVF patients High-performance liquid chromatography (HPLC) measurements of 25 AAs were the main outcome measures RESULTS: There was a significant gradient of most AAs from plasma to follicular fluid, with the exception of glutamate, which demonstrated a three-fold increase in follicular fluid concentration (700 ± 380 mM) compared with plasma (2318 ± 220 mM; P < 0001) The plasma-to-follicular fluid concentration difference for glutamine (8183 ± 92 mM) was greatest among all AAs Among essential AAs, this difference was greatest for the branched-chain AAs, isoleucine, leucine and valine Ammonia concentrations in follicular fluid and blood were 3887 ± 223 and 2211 ± 196 mM, respectively (P < 0001) Urea concentration in follicular fluid was 337 ± 018 mM, a value not significantly different from plasma concentration (336 ± 022 mM; P = 0911) CONCLUSIONS: These plasma–follicular fluid differences may reflect both the utilization of AAs and the transport characteristics of the follicular cells There is accumulation of glutamate and ammonia in pre-ovulatory follicular fluid The data for urea are consistent with transport by passive diffusion, with no evidence of an active urea cycle in the cells of the follicle

Global effects of vitamin A deficiency on gene expression in rat liver: evidence for hypoandrogenism.

Abstract: Vitamin A (retinol) metabolites are ligands for transcription factors that regulate many genes. The liver is the main storage depot for retinol and plays a role in vitamin A homeostasis. To better understand the effects of vitamin A deficiency on liver gene expression, we produced retinol deficiency in male rats by feeding a diet low in retinol for 53 days after weaning and examined the effects on gene expression in liver using Affymetrix oligonucleotide microarrays. We detected expression of 41% of the 8799 probe sets represented on the RGU-34A GeneChips. Vitamin A deficiency resulted in major changes in liver gene expression: 805 genes (22% of all genes detected) differed at P≤.05 (false discovery rate <0.143). Genes involved in fatty acid metabolism, peroxisomal function, glycolysis, glutamate metabolism and the urea cycle were altered. The expression of many sexually dimorphic genes was altered toward a feminized or senescent pattern of gene expression in the liver. Retinol deficiency also produces a shift toward increased protein and fat catabolism and decreased fatty acid synthesis.

Disorders of the Urea Cycle and Related Enzymes

Abstract: Six inherited disorders of the urea cycle are well described (Fig. 20.1). These are the deficiencies of carbamoyl phosphate synthetase (CPS), ornithine transcarbamoylase (OTC), argininosuccinate synthetase, argininosuccinate lyase, arginase, and N-acetylglutamate synthetase (NAGS). Deficiencies of glutamine synthetase and of citrin have also been described. All these defects are characterised by hyperammonaemia and disordered amino acid metabolism. The presentation is highly variable: those presenting in the newborn period usually have an overwhelming illness that rapidly progresses from poor feeding, vomiting, lethargy or irritability and tachypnoea to fits, coma and respiratory failure. In infancy, the symptoms are less severe and more variable. Poor developmental progress, behavioural problems, hepatomegaly and gastrointestinal symptoms are common. Children and adults frequently have a chronic neurological illness that is characterised by variable behavioural problems, confusion, irritability and episodic vomiting. However, during any metabolic stress the patients may become acutely unwell. Arginase deficiency has more specific symptoms, such as spastic diplegia, dystonia, ataxia and fits. All these disorders have autosomal- recessive inheritance except ornithine transcarbamoylase deficiency, which is X-linked.

Effects of arginine treatment on nutrition, growth and urea cycle function in seven Japanese boys with late-onset ornithine transcarbamylase deficiency

Abstract: The aim of this study was to investigate the effects of arginine on nutrition, growth and urea cycle function in boys with late-onset ornithine transcarbamylase deficiency (OTCD). Seven Japanese boys with late-onset OTCD enrolled in this study resumed arginine treatment after the cessation of this therapy for a few years. Clinical presentations such as vomiting and unconsciousness, plasma amino acids and urinary orotate excretion were followed chronologically to evaluate urea cycle function and protein synthesis with and without this therapy. In addition to height and body weight, blood levels of proteins, lipids, growth hormone (GH), insulin-like growth factor-I (IGF-I) and IGF-binding protein -3 (IGFBP-3) were monitored. The frequency of hyperammonemic attacks and urinary orotate excretion decreased significantly following the resumption of arginine treatment. Despite showing no marked change in body weight, height increased gradually. Extremely low plasma arginine increased to normal levels, while plasma glutamine and alanine levels decreased considerably. Except for a slight increase in high-density lipoprotein cholesterol level, blood levels of markers for nutrition did not change. In contrast, low serum IGF-I and IGFBP-3 levels increased to age-matched control levels, and normal urinary GH secretion became greater than the level observed in the controls. Arginine treatment is able to reduces attacks of hyperammonemia in boys with late-onset OTCD and to increase their growth.

Clinical and Functional Characterization of a Human ORNT1 Mutation (T32R) in the Hyperornithinemia-Hyperammonemia-Homocitrullinuria (HHH) Syndrome

Abstract: We studied two related families (HHH013 and HHH015) with the hyperornithinemia-hyperammonemia-homocitrullinuria (HHH) syndrome, a disorder of the urea cycle and ornithine degradation pathway, who have the same novel ornithine transporter (ORNT1) genotype (T32R) but a variable phenotype. Both HHH015 patients are doing well in school and are clinically stable; conversely, the three affected HHH013 siblings had academic difficulties and one suffered recurrent episodes of hyperammonemia and ultimately died. Overexpression studies revealed that the product of the ORNT1-T32R allele has residual function. Ornithine transport studies in HHH015 fibroblasts, however, showed basal activity similar to fibroblasts carrying nonfunctional ORNT1 alleles. We also examined two potential modifying factors, the ORNT2 gene and the mitochondrial DNA lineage (haplogroup). Haplogroups, associated with specific diseases, are hypothesized to influence mitochondrial function. Results demonstrated that both HHH015 patients are heterozygous for an ORNT2 gain of function polymorphism and belong to haplogroup A whereas the HHH013 siblings carry the wild-type ORNT2 and are haplogroup H. These observations suggest that the ORNT1 genotype cannot predict the phenotype of HHH patients. The reason for the phenotypic variability is unknown, but factors such as redundant transporters and mitochondrial lineage may contribute to the neuropathophysiology of HHH patients.

Nitrogen metabolism and excretion in the aquatic chinese soft‐shelled turtle, Pelodiscus sinensis, exposed to a progressive increase in ambient salinity

Abstract: This study aimed to determine effects of 6-day progressive increase in salinity from 1 per thousand to 15 per thousand on nitrogen metabolism and excretion in the soft-shelled turtle, Pelodiscus sinensis. For turtles exposed to 15 per thousand water on day 6, the plasma osmolality and concentrations of Na+, Cl- and urea increased significantly, which presumably decreased the osmotic loss of water. Simultaneously, there were significant increases in contents of urea, certain free amino acids (FAAs) and water-soluble proteins that were involved in cell volume regulation in various tissues. There was an apparent increase in proteolysis, releasing FAAs as osmolytes. In addition, there might be an increase in catabolism of certain amino acids, producing more ammonia. The excess ammonia was retained as indicated by a significant decrease in the rate of ammonia excretion on day 4 in 15 per thousand water, and a major portion of it was converted to urea. The rate of urea synthesis increased 1.4-fold during the 6-day period, although the capacity of the hepatic ornithine urea cycle remained unchanged. Urea was retained for osmoregulation because there was a significant decrease in urea excretion on day 4. Increased protein degradation and urea synthesis implies greater metabolic demands, and indeed turtles exposed to 15 per thousand water had significantly higher O2 consumption rate than the freshwater (FW) control. When turtles were returned from 15 per thousand water to FW on day 7, there were significant increases in ammonia (probably released through increased amino acid catabolism) and urea excretion, confirming that FAAs and urea were retained for osmoregulatory purposes in brackish water.

In Vivo Urea Kinetic Studies in Conscious Mice

Abstract: Stable isotope studies in conscious mice have been limited by the invasive catheterization procedures and relatively large sample size required. We developed minimally invasive catheterization protocols that together with the ability to analyze small samples have allowed for the study of urea kinetics in conscious mice. A single dose of 15N15N-urea followed by multiple sampling in mice (n = 6) showed that a primary pool of urea exchanged rapidly [70.65 +/- 14.96 mmol/(kg x h)] with a secondary pool. The urea entry rate determined with this protocol was 3.36 +/- 0.30 mmol/(kg x h). Continuous infusion of 15N15N-urea (n = 6) achieved plateau enrichment values at 3.3 +/- 0.2.h from which the urea entry rate was determined by isotope dilution [3.24 +/- 0.23 mmol/(kg x h)]. The urea entry rate measured by the single dose or continuous infusion protocol did not differ (P = 0.76). The minimally invasive methods described allow us to study not only ureagenesis and urea cycle disorders in vivo, but also urea transport and transporter function and nitrogen metabolism in general in mouse models. This is especially relevant because mouse targeting technologies will likely facilitate the generation of organ and tissue specific nulls of the various urea cycle enzymes.

Influence of sodium fluoride and caffeine on the concentration of fluoride ions, glucose, and urea in blood serum and activity of protein metabolism enzymes in rat liver.

Abstract: The aim of the study was examining the effect of fluoride ions and caffeine administration on glucose and urea concentration in blood serum and the activity of protein metabolism enzymes and selected enzymes of the urea cycle in rat liver. The study was carried out using 18 male Sprague-Daowley rats (4.5 mo old). Rats were divided into three groups. Group I received distilled water ad libitum. Group II received 4.9 mg F-/kg body mass/d of sodium fluoride in the water, and group III received sodium fluoride (in the above-mentioned dose) and 3 mg/kg body mass/d of caffeine in the water. After 50 d, the rats were anesthetized with thiopental and fluoride ions, glucose, and urea concentration in blood serum were determined. Also determined were the activities of aspartate aminotransferase, alanine aminotransferase glutamate dehydrogenase, ornithine carbamoylotransferase and arginase in liver homogenates. Liver was taken for pathomorphological examinations. The applied doses of F- (4.9 mg/kg body mass/d) and F- + caffeine (4.9 mg F-/kg body mass/d + 3 mg caffeine/kg body mass/d) resulted in a statistically significant increase of fluoride ion concentration in blood serum, a slight increase of the glucose concentration, and no changes in the concentration of urea in blood serum. This might testify to the absence of kidney lesions for the applied concentrations of F-. No change in the functioning of hepatocytes was observed; however, slight disturbances have been noted in the functioning of the liver, connected with the activation of urea cycle, increase of arginase activity, and accumulation of F- in this organ. There was no observed significant influence of caffeine supplementation on the obtained results.

A proteomics approach to identify protein expression changes in rat liver following administration of 3,5,3'-triiodo-L-thyronine.

Abstract: We analyzed whole cell protein content of rat liver following T3 administration. Fourteen differentially expressed proteins were unambiguously identified and were involved in substrates and lipid metabolism, energy metabolism, detoxification of cytotoxic products, calcium homeostasis, amino acid catabolism, and the urea cycle. This study represents the first systematic identification of T3-induced changes in liver protein expression profile and provides novel information at the molecular, cellular, and tissue level of T3 action.

Postprandial increases in nitrogenous excretion and urea synthesis in the Chinese soft-shelled turtle, Pelodiscus sinensis.

Abstract: The objective of this study was to determine the effects of feeding on the excretory nitrogen (N) metabolism of the aquatic Chinese soft-shelled turtle, Pelodiscus sinensis, with a special emphasis on the role of urea synthesis in ammonia detoxification. P. sinensis is ureogenic and possesses a full complement of ornithine-urea cycle enzymes in its liver. It is primarily ureotelic in water, and the estimated rate of urea synthesis in unfed animals was equivalent to only 1.5% of the maximal capacity of carbamoyl phosphate synthetase I (CPS I) in its liver. Approximately 72 h was required for P. sinensis to completely digest a meal of prawn meat. During this period, there were significant increases in ammonia contents in the stomach at hour 24 and in the intestine between hours 12 and 36, which could be a result of bacterial activities in the intestinal tract. However, ammonia contents in the liver, muscle, brain and plasma remained unchanged throughout the 72-h post-feeding. In contrast, at hour 24, urea contents in the stomach, intestine, liver, muscle, brain and plasma increased significantly by 2.9−, 3.5−, 2.6−, 2.9−, 3.4 and 3.0-fold, respectively. In addition, there was a 3.3- to 8.0−fold increase in the urea excretion rate between hours 0 and 36 post-feeding, which preceded the increase in ammonia excretion between hours 12 and 48. By hour 48, 68% of the assimilated N from the feed was excreted, 54% of which was excreted as urea-N. The rate of urea synthesis apparently increased sevenfold during the initial 24 h after feeding, which demanded only 10% of the maximal CPS I capacity in P. sinensis. The postprandial detoxification of ammonia to urea in P. sinensis effectively prevented postprandial surges in ammonia contents in the plasma and other tissues, as observed in other animals, during the 72-h period post-feeding. In addition, postprandial ammonia toxicity was ameliorated by increased transamination and synthesis of certain amino acids in the liver and muscle of P. sinensis. After feeding, a slight but significant increase in the glutamine content occurred in the brain at hour 24, indicating that the brain might experience a transient increase in ammonia and ammonia was detoxified to glutamine.

Identification of novel mutations in the human ornithine transcarbamylase (OTC) gene of Korean patients with OTC deficiency and transient expression of the mutant proteins in vitro.

Abstract: The urea cycle plays key roles to prevent the accumulation of toxic nitrogenous compound and synthesize arginine de novo. Ornithine transcarbamylase (OTC) deficiency is the most common inborn error of urea cycle, which is inherited in an X-linked manner. This study was undertaken to characterize molecular defects in Korean patients with OTC deficiency. With direct sequence analysis of OTC gene of 26 unrelated Korean patients with OTC deficiency, 23 different mutations were identified. Among these mutations, eleven were novel mutations. The novel mutations were p.Leu9X, p.Arg26Pro, p.Gly100Arg, p.Met205Thr, p.Lys221Asn, p.Asp249Gly, p.Phe281Ser, p.Val323Met, c.571delC, c.853delC, and c.796-805del. All the novel mutations in this study were tested in 100 normal alleles. In vitro expression study of some of novel missense mutations elucidated the correlation of genotype and phenotype of the OTC deficiency. © 2006 Wiley-Liss, Inc.

Ornithine Restores Ureagenesis Capacity and Mitigates Hyperammonemia in Otcspf-ash Mice

Abstract: We showed that Otc(spf-ash) mice, a model of ornithine transcarbamylase deficiency, were able to sustain ureagenesis at the same rate as control mice, despite reduced enzyme activity, when a complete mixture of amino acids was provided. An unbalanced amino acid mixture, however, resulted in reduced ureagenesis and hyperammonemia. To study the effect of ornithine supplementation [316 micromol/(kg.h)] on urea and glutamine kinetics in conscious Otc(spf-ash) mice under a glycine-alanine load [6.06 mmol/(kg.h)], a multiple tracer infusion protocol ([(13)C(18)O]urea, [5-(15)N]glutamine, [2,3,3,4,4 D(5)]glutamine and [ring-D(5)] phenylalanine) was conducted. Ornithine supplementation increased ureagenesis [3.18 +/- 0.88 vs. 4.56 +/- 0.51 mmol/(kg.h), P < 0.001], reduced plasma ammonia concentration (1125 +/- 621 vs. 193 +/- 94 micromol/L, P < 0.001), and prevented acute hepatic enlargement (P < 0.006) in Otc(spf-ash) mice. Ornithine supplementation also increased [96 +/- 20 vs. 120 +/- 16 micromol/(kg.h), P < 0.001] the transfer of (15)N from glutamine to urea, to values observed in the control mice [123 +/- 17 micromol/(kg.h)]. De novo amido-N glutamine flux was higher [1.57 +/- 0.37 vs. 3.04 +/- 0.86 mmol/(kg.h); P < 0.001] in Otc(spf-ash) mice, but ornithine supplementation had no effect (P < 0.56). The flux of glutamine carbon skeleton was affected by both genotype (P < 0.0001) and by ornithine (P 0. 036). In conclusion, ornithine supplementation restored ureagenesis, mitigated hyperammonemia, prevented liver enlargement, and normalized the transfer of (15)N from glutamine to urea. These data strongly suggest that ornithine has the potential for the biochemical correction of OTCD in Otc(spf-ash) mice.

Enzymatic evidence for the key role of arginine in nitrogen translocation by arbuscular mycorrhizal fungi

Abstract: Key enzymes of the urea cycle and (15)N-labeling patterns of arginine (Arg) were measured to elucidate the involvement of Arg in nitrogen translocation by arbuscular mycorrhizal (AM) fungi. Mycorrhiza was established between transformed carrot (Daucus carota) roots and Glomus intraradices in two-compartment petri dishes and three ammonium levels were supplied to the compartment containing the extraradical mycelium (ERM), but no roots. Time courses of specific enzyme activity were obtained for glutamine synthetase, argininosuccinate synthetase, arginase, and urease in the ERM and AM roots. (15)NH(4)(+) was used to follow the dynamics of nitrogen incorporation into and turnover of Arg. Both the absence of external nitrogen and the presence of L-norvaline, an inhibitor of Arg synthesis, prevented the synthesis of Arg in the ERM and resulted in decreased activity of arginase and urease in the AM root. The catabolic activity of the urea cycle in the roots therefore depends on Arg translocation from the ERM. (15)N labeling of Arg in the ERM was very fast and analysis of its time course and isotopomer pattern allowed estimation of the translocation rate of Arg along the mycelium as 0.13 microg Arg mg(-1) fresh weight h(-1). The results highlight the synchronization of the spatially separated reactions involved in the anabolic and catabolic arms of the urea cycle. This synchronization is a prerequisite for Arg to be a key component in nitrogen translocation in the AM mycelium.

Pharmacology Review: Sodium Phenylacetate and Sodium Benzoate in the Treatment of Neonatal Hyperammonemia

Abstract: Ammonia is present in all body fluids and exists primarily as ammonium ion at physiologic pH. Hyperammonemia is defined as a blood ammonia concentration greater than about 100 mcmol/L in neonates or 50 mcmol/L in children and adults (precise cut-offs vary, depending on individual laboratory normative ranges). The concentration of ammonia is 10 times higher in tissue than in blood. A 5- to 10-fold increase in blood ammonia concentration usually is toxic to the nervous system. Hyperammonemia in the neonatal period, especially when due to inborn errors of metabolism, can progress rapidly and cause severe neurologic damage or early death. Hyperammonemia can be caused by inborn errors of metabolism as well as by a variety of acquired conditions (Tables 1 and 2). Urgent treatment is required because of the potential for irreversible neurologic sequelae that can, in many cases, be prevented by prompt diagnosis and institution of therapy. | Urea cycle defects | || | Amino acid transporter deficiencies | | Organic acidemias | | Fatty acid oxidation defects | | Pyruvate carboxylase deficiency | | Mitochondrial disorders | | Hyperinsulinism/hyperammonemia syndrome (glutamate dehydrogenase mutations) | | Delta1-pyrroline-5-carboxylate synthase deficiency | Table 1. Inborn Errors of Metabolism Associated With Hyperammonemia | Sampling artifact | || | Cardiovascular | | Perinatal asphyxia | | Liver failure | | Bacterial colonization (urease-positive organisms) | | Iatrogenic | Table 2. Causes of Acquired Hyperammonemia The combination of sodium phenylacetate (NAPA) and sodium benzoate (NABZ) in a 10%/10% solution is an intravenously administered United States Food and Drug Administration (FDA)-approved drug used as adjunctive therapy for …

Reduced Ornithine Transcarbamylase Activity Does Not Impair Ureagenesis in Otcspf-ash Mice

Abstract: Mouse models for urea cycle disorders have been available for the past 30 y; however, until now, no measurements of urea production in vivo have been conducted. Urea entry rate was determined in Otc spf-ash and littermate controls employing a primed-continuous infusion of 15 N 15 N urea. A saline infusion control, a complete mixture of amino acids (AA), or a glycine-alanine (GA) mixture was infused at 86 (AA1 and GA1) and 172 mg Nkg � 1 � h � 1 (AA2 and GA2) to impose a defined nitrogen load on the urea cycle. Urea entry rate and plasma urea concentration increased (P , 0.001) as a consequence of the increase in the infusion rate of the complete mixture of amino acids, but the 2 genotypes did not differ (P ¼ 0.96 and P ¼ 0.44, respectively). The infusion of the GA mixture, however, decreased (P , 0.001) the plasma urea concentration and urea entry rate in Otc spf-ash mice compared with controls. At the highest level (GA2), urea entry rate was further depressed (P , 0.001), Otc spf-ash mice became hyperammonemic (1701 6 150 mmol/L), and hyperammonemic symptoms were evident. An acute hepatic enlargement (P , 0.001) was also evident in Otc spf-ash mice infused with GA2. These results show that despite vestigial OTC activity, Otc spf-ash mice were able to maintain ureagenesis at the same rate of control animals when a complete mixture of amino acids was infused. This implies that Otc spf-ash mice are able to dispose of ammonia, without apparent adverse effects, when a balance mixture of amino acids is provided, despite reduced enzyme activity. J. Nutr. 136: 1017-1020, 2006.

Citrulline and Ammonia Accumulating in Citrullinemia Reduces Antioxidant Capacity of Rat Brain In Vitro

Abstract: Citrullinemia is an inborn error of the urea cycle caused by deficient argininosuccinate synthetase, which leads to accumulation of L-citrulline and ammonia in tissues and body fluids. The main symptoms include convulsions, tremor, seizures, coma, and brain edema. The pathophysiology of the neurological signs of citrullinemia remains unclear. In this context, we investigated the in vitro effects of L-citrulline and ammonia in cerebral cortex from 30-day-old rats on oxidative stress parameters, namely thiobarbituric acid-reactive substances (TBA-RS), chemiluminescence, mitochondrial membrane protein thiol content, intracellular content of hydrogen peroxide, total radical-trapping antioxidant potential (TRAP), total antioxidant reactivity (TAR) as well as on the activities of the antioxidant enzymes (catalase, superoxide dismutase, and glutathione peroxidase). L-Citrulline significantly diminished TRAP (26%) and TAR (37%), while ammonia decreased TAR (30%). Ammonia increased SOD activity (65%) and L-citrulline did not affect the activities of any antioxidant enzymes. We also observed that L-citrulline and ammonia did not alter lipid peroxidation parameters, levels of hydrogen peroxide, and mitochondrial membrane protein thiol content. Taken together, these results may indicate that L-citrulline and ammonia decreased the antioxidant capacity of the brain, which may reflect a possible involvement of oxidative stress in the neuropathology of citrullinemia.