Showing papers on "Urea cycle published in 1981"

Adaptation of nitrogen metabolism to hyperosmotic environment in Amphibia

Abstract: A large number of species of Amphibia, both Anura and Urodela, are capable of tolerating a moderately saline environment. Rana cancrivora, Bufo viridis, and Xenopus laevis are among the most euryhaline frogs so far studied. Rana cancrivora can tolerate undiluted seawater. In a saline environment, Amphibia show raised plasma sodium and chloride and raised intracellular potassium and chloride. In the adults, plasma and tissue urea are elevated, especially in the more euryhaline species. Free amino acids contribute negligibly to plasma osmolarity, but are very important in intracellular fluids. By these various means, the osmotic pressure of body fluids is always maintained at a higher level than that of the surroundings. Larval Amphibia, however, do not make urea; Rana cancrivora tadpoles can live in saltwater, but maintain the osmolarity of their body fluids below that of their surroundings. Adaptive responses to hyperosmolar environment include decreased skin sodium transport, greatly reduced urine flow, and release of posterior pituitary hormones. After the initial response, the release of these hormones declines, and urine flow increases. Accumulation of urea occurs slowly, but this substance plays an increasingly important role as adaptation proceeds. Accumulation is due initially to urea retention, and possibly to greater synthesis due to a high concentration of precursors of the urea cycle. In later stages of adaptation, increased urea synthesis is due to elevated levels of urea cycle enzymes, especially those that appear to have been rate-limiting. Liver glutamate dehydrogenase is also elevated. In animals subjected to pure osmotic stress, in solutions not containing sodium, responses are similar to, but not identical with, those caused by a medium containing sodium chloride.

Effects of arginine-free meals on ureagenesis in cats

Abstract: Cats given a single arginine-free meal have been reported to develop severe hyperammonemia, attributed to impaired function of ornithine aminotransferase (OAT). We found that cats that developed hyperammonemia following an arginine-free meal had low hepatic ornithine levels. However, the average sum of hepatic ornithine plus arginine plus citrulline rose, indicating that some ornithine synthesis via OAT took place, and hyperammonemia failed to occur in cats with higher hepatic ornithine levels. OAT activity and kinetic constants were comparable to values reported in the rat. Furthermore, dietary supplementation with ornithine caused only occasional and transient hyperornithinemia. Thus, OAT can function in the cat. The Ka of N-acetylglutamate (AGA) synthetase for arginine was 5 times higher in cats than in rats, but AGA content and citrullinogenesis by intact mitochondria were the same following arginine-free or arginine-containing meals. Other kinetic parameters of AGA synthetase and carbamoylphosphate synthetase were similar to values in the rat. We conclude that low levels of hepatic ornithine are probably responsible for making some cats susceptible to hyperammonemia following this stimulus.

Effect of casein and starch infusion in the large intestine on nitrogen metabolism of growing swine.

Abstract: Eight crossbred female pigs (40 kg) with cannulae placed in the terminal ileum were used to evaluate the effect of infusions of casein starch and casein plus starch on organic matter fermentation and microbial protein synthesis in the large intestine, and their effect on urinary urea and orotic acid excretion, and on nitrogen (N) retention. Infused casein and starch were both totally digested. Nitrogen retention was increased (P greater than 0.05) when casein was infused. Starch infusion resulted in an increase (P greater than 0.05) in fecal N in the form of total protein (amino acids). The high correlation (P greater than 0.01) between fecal total protein and RNA indicates that the increase in fecal N resulted from an increase in microbial protein synthesis. About 5.2 g of bacterial protein was synthesized per 100 g of cornstarch fermented in the large intestine. Casein infusion increase (P greater than 0.05) total urinary N. Differences between treatments for urinary N were entirely because of changes in urinary urea. Urinary ammonia and unaccounted N were not affected by treatments. Urinary orotic acid was a good indicator of the urea cycle activity because of its hig correlation (P greater than 0.0) with urinary urea. Plasma urea N concentration was increased (P greater than 0.05) only when casein plus starch was infused.

Orotic Aciduria in Lysinuric Protein Intolerance: Dependence on the Urea Cycle Intermediates

Abstract: Orotic Aciduria in Lysinuric Protein Intolerance: Dependence on the Urea Cycle Intermediates

Dietary and hormonal regulation of urea cycle enzymes in rat liver.

Abstract: When 6-week-old rats fed normal diet (22% protein) were transferred to 10 and 75% protein diet, the levels of urea cycle enzymes showed decreases and increases respectively. The activities expressed as units per gram wet weight of liver had not stabilized after 7 days on the new diet; the corresponding specific activities were closer to leveling off. Four daily injections of cortisol raised CPS, ASS, and arginase. The percentage increases were greater on a 10 than on a 75% protein diet. Adrenalectomy of rats fed 10% protein decreased all urea cycle enzymes; on 75% protein, only arginase decreased. All enzymes could be raised to control levels within 24 h by three injections of cortisol. Thyroxine produced only slight increases in urea cycle enzymes. On a 10% protein diet, all five enzymes were raised by thyroidectomy, and further raised by injection of thyroxine.

Ultrastructural pathology in congenital defects of the urea cycle: Ornithine transcarbamylase and carbamylphosphate synthetase deficiency

Abstract: Inborn defects of urea synthesis, leading to hyperammonemia, are complex inherited disorders, whose structural sequelae in different tissues and organs have not yet been studied in detail. Ultrastructural investigations have been performed on two cases of deficiencies of two consecutive enzymes of the urea cycle, carbamylphosphate synthetase and ornithine transcarbamylase, and the findings are compared with previously reported results. With regard to liver pathology it appeared that 1). Hepatocytes in CPS deficiency mainly exhibited changes of SER and mitochondrial compartments, whereas 2). OTC deficiency was characterized by regressive liver cell change, with abnormal configuration of the RER, formation of telolysosomes and peribiliary vesiculation. It is suggested that the mitochondrial disorder in the CPS defect is directly related to the lack of a major enzyme protein in this organelle, resulting in structural damage. The leading renal change in CPS deficiency is foot process fusion of glomerular podocytes. Brain alteration in this disorder is similar to that reported for other hyperammonaemic urea cycle defects.

Biochemical Aspects of Urea Cycle Disorders

Abstract: In this paper I will review the biochemistry of the urea cycle of mammalian liver and point out the important clinical implications of this biochemistry. Second, I will consider some problems that arise in the use of human liver enzyme assays for evaluating deficiencies of urea cycle enzymes. Finally, I will comment on our own research concerning enzyme induction of the urea cycle of mammalian liver. In Fig 1 the urea cycle is reviewed. Within the mitochondrion of the liver cell, ammonium cation combines with bicarbonate anion and 2 moles of adenosine 59-triphosphate (ATP) in the presence of an activator, N-acetylglutamate, to form carbamylphosphate. Enzyme 1, which carries out this reaction is carbamylphosphate synthetase I (CPS1). There is also another enzyme, CPS2, in the cytoplasm of liver cells about which more will be said later. Once carbamylphosphate is formed, the carbamyl group is transferred to the δ-amino group of ornithine to form citrulline. The enzyme in this reaction, enzyme 2, is ornithine transcarbamylase (OTC). Citrulline escapes, by a process other than active transport, into the cytoplasm where it is conjugated with aspartate to form argininosuccinate; this is accomplished by enzyme 3, argininosuccinate synthetase (AS). Argininosuccinate, a double amino acid, is then cleaved to yield fumarate, and the nitrogen of aspartate moves to arginine. The enzyme in this reaction, enzyme 4, is argininosuccinate lyase (AL); its trivial name is argininosuccinase. Finally, arginine undergoes cleavage of its ureido group by enzyme 5, arginase, to form urea and regenerate ornithine. Ornithine is then taken up into the mitochondria by an active transport process to combine again with carbamylphosphate.

Evidence of lack of toxicity of sodium phenylacetate and sodium benzoate in treating urea cycle enzymopathies.

Abstract: EVIDENCE OF LACK OF TOXICITY OF SODIUM PHENYLACETATE AND SODIUM BENZOATE IN TREATING UREA CYCLE ENZYMOPATHIES A question concerning the potential toxicity of sodium phenylacetate and sodium benzoate has been recently raised by Dr Woolf (J. bTher. Metab. Dis. 3 (1980) 61). He cited Sherwin and Kennard's work in 1919 where they described 12 adults who exhibited clinical symptoms following ingestion of a single dose of 5g of sodium phenylacetate, and one who ingested 16 g. With regard to sodium phenylacetate, it should be noted that Ambrose et aL (1933) gave 5-10g phenylacetate daily to one man for 28 days. They gave smaller doses to 34 other subjects. No side-effects were noted. We have treated two patients with urea-cycle enzymopathies for 4-26 weeks using phenylacetate at a dose of 250 mg/kg daily without side-effects. In one, a 3-yearold girl with ornithine transcarbamylase deficiency, plasma ammonia concentra t ions fell from 120 to 30gmol/l, and glutamine from 2000 to I100gmol/l during therapy, Tests of hepatic, renal and haematologic function remained within normal limits. The child tolerated the medication without abdominal distress, lethargy or irritability. The mother did note the rather unpleasant odour of this drug. A second patient, a 17-year-old girl with partial carbamyl phosphate synthetase deficiency, showed a similar benign course. She received from 6 to 10g phenylacetate daily. Plasma ammonia concentrations were within normal limits during therapy. We agree with Dr Woolf that sodium benzoate may not be the best choice in treating patients with hyperammonaemia secondary to impaired liver function. Phenylacetate may be more effective in such cases, since the acetylation enzyme activity is greater in human kidney than in human liver (Moldave and Meister, 1957). We have used phenylacetate in the short-term management of hyperammonaemia in Reye's syndrome and hepatic failure. Two children with each condition received 250mg/kg phenylacetate daily intravenously for 1-3 days. Plasma ammonia concentrations fell in each instance. There was no change in acid-base balance, hepatic, renal or haematological function. The children were comatose at the time of therapy, and there was no change in clinical status observed during therapy. As for sodium benzoate, this compound has been used safely as a food preservative for many years, and has been used to treat infants with non-ketotic hyperglycinaemia for as long as 3 years at a dose of 300 mg/kg without reported toxicity (Trijbels et aL, 1974). We have used it to treat infants with carbamyl phosphate synthetase deficiency of neonatal onset, ornithine transcarbamyIase deficiency, and argininosuccinic acid synthetase deficiency for 2-30 months without evidence of toxicity (Batshaw et al., 1981). Two children have had transient increases in SGOT levels during benzoate therapy, but this also occurs during untreated hyperammonaemia. Sodium benzoate has been well tolerated and effective in maintaining normal ammonia concentrations in these children. Finally, Dr Woolf asks if the coenzyme A-esters of these compounds inhibit N-acetylglutamate synthetase, thereby worsening hyperammonaemia. Although it is known that CoA esters of organic acids are competitive inhibitors of N-acetylglutamate synthetase (Coude et al., 1979), it should be noted that there may be striking quantitative differences for this effect of aliphatic CoA esters. Perhaps the best evidence that such inhibition does not occur h~ vivo is the consistent fall in ammonia concentrations following infusion of sodium benzoate in hyperammonaemic patients. We believe that these preliminary data suggest that continued trials of sodium benzoate and sodium phenylacetate are warranted. There is also a need for additional pharmacological and toxicological information, particularly in infants.

Urinary pyrimidine excretion in arginase deficiency.

Abstract: A high-performance liquid-chromatographic method was used to separate and identify uracil, uridine, pseudouridine and orotic acid after preliminary extraction in two patients (McKusick 20780). Urinary uracil excretion was 10-35 times normal in both patients with arginase deficiency. Uridine and orotic acid, not normally detected, were excreted in large amounts and were directly influenced by protein intake. Their excretions were correlated with urinary arginine excretion. Urinary uracil levels remained consistently high and showed minimal variations with increased protein intake or urinary arginine levels. The measurement of urinary pyrimidines appears to be useful for the detection, differential diagnosis and dietary monitoring of patients with urea cycle disorders. The data presented extends this observation to include patients with arginase deficiency.

Clinical Aspects of Disorders of the Urea Cycle

Abstract: SYMPTOMS All of the defects of the urea cycle may result in the accumulation of ammonia. This is manifested clinically by episodes of vomiting, lethargy, seizures, coma, and ultimately in death if the hyperammonemia is not controlled. In older children these symptoms may be preceded by a period of irritability and hyperactivity. It has usually been considered that the severity of the hyperammonemia depends on the site of the block; the nearer it is to ammonia, the greater the accumulation. There are, however, exceptions, as hyperammonemia leading to neonatal death has been observed in arginino-succinate lyase deficiency,39,40 the fourth step in the cycle. Death due to hyperammonemia has been observed in all the defects except arginase deficiency.41-43 There are variations in the severity of the clinical course of all of these syndromes; the exact reason for this has not been ascertained, although it is quite possible that this is related to the degree of residual enzyme activity. Thus, carbamylphosphate synthetase deficiency has been associated both with death in the newborn period or survival without specific treatment but with physical and mental retardation for a number of years.42-44 The clinical features of ornithine transcarbamylase deficiency have been the most completely described.45,46 As this anomaly is inherited as a sexlinked dominant, males are most severely affected, and neonatal death has been almost universal.13 Males who have survived without the most vigorous treatment have had a variant of the disease,47,48 the enzyme having different kinetic properties. Females who have various degrees of residual enzyme activity usually do not manifest symptoms until sometime after the neonatal period, often in response to some stress.

Carrier Detection of Urea Cycle Disorders

Abstract: Published reports100,102,108 indicate that the degree of success attained in the treatment of infants with urea cycle disorders has not been completely satisfactory. Some new approaches in dietary management, using a combination of arginine, benzoate, and phenylacetate may be useful,67,72,76 but more studies are required. In many instances, clinical problems reflect delays in biochemical diagnosis. Families at risk need to be identified and the genotype of the fetus should be established as soon as feasible after conception, so that appropriate management can be provided during pregnancy and after delivery. Availability of effective methods for carrier detection is a critical component of this process. With any metabolic disorder, a number of factors should be considered for carrier detection. These include: (1) frequency of occurrence of affected patients in the available population, (2) mode of inheritance, (3) cost-benefit ratio, (4) soundness of the methodology used, and (5) feasibility of prenatal diagnosis. Interestingly, the Tay-Sachs prevention program with the major objective of carrier identification is the only accepted program that has been widely discussed and evaluated.109 The frequency of occurrence of each of the five urea cycle enzyme disorders is uncertain, but ornithine transcarbamylase (OTC) deficiency, the only sex-linked disorder, is considered to be the most common. Not only the hemizygotes, but also many of the heterozygotes of OTC deficiency are clinically affected. The frequency of argininosuccinic aciduria has been estimated to be 1 in 60,000, based on results of a newborn screening program carried out in Massachusetts.110 Carrier identification of urea cycle disorders in the general population is not practical because of cost considerations, but it should be applied to members of affected families.

Regulation of pathways of ornithine metabolism. Effects of thyroid hormone and diabetes on the activity of enzymes at the "ornithine crossroads' in rat liver.

Abstract: There is a significant increase in four of the urea cycle enzymes in liver from hypothyroid rats, arginase alone showed an opposite trend; these changes are reversed by physiological doses of thyroxine; hyperthyroidism results in a significant decrease in ornithine transcarbamoylase activity. The pattern of change in thyroidectomized and alloxan-diabetic rats showed marked similarities in respect to urea cycle and ornithine-metabolizing enzymes which are discussed in the light of the common feature of hypoinsulinism of diabetes and depressed response to insulin in hypothyroidism. The profile of ornithine-metabolizing enzymes is consonant with the decreased protein synthesis and turnover in hypothyroidism.

Aminoacid parenteral infusion soln. - for hepatic coma treatment contg. relatively high concns. of arginine, ornithine, glutamic acid and aspartic acid

Abstract: New aminoacid soln. for parenteral infusion contains an amino acid mixt. of the following compsn.: 3.0+/-1.6 wt.parts L-aspartic acid; 4.6+/-0.8 wt.parts L-threonine; 2.7+/-1.1 wt.parts L-serine; 7.1+/-2.5 wt.parts L-proline; 5.7+/-2.8 wt.parts L-glutamic acid; 6.3+/-1.5 wt.parts L-glycine; 8.3+/-2.2 wt.parts L-alanine; 10.4+/-1.5 wt.parts L-valine; 2.0+/-1.0 wt.parts L-cysteine; 0.8+/-0.4 wt.parts L-methionine; 8.6+/-2.9 wt.parts L-isoleucine; 13.4+/-2.4 wt.parts L-leucine; 0.7+/-0.4 wt.parts L-tyrosine; 1.6+/-1.0 wt.parts L-phenylalanine; 7.5+/-2.0 wt.parts L-lysine; 4.7+/-1.4 parts L-histidine; 1.5+/-0.9 wt.parts L-tryptophan; 8.8+/-2.7 wt.parts L-arginine; and 1.3+/-0.3 wt.parts L-ornithine. Used for treatment of cerebral function disorders caused by hepatic insufficiency (i.e. hepatic coma). The onset of clinical effects is faster and the effect is longer lasting than with known parenteral amino acid compsns. for the treatment of hepatic coma. The effect on hepatic coma is accompanied by a lowering of elevated blood ammonia levels, attributed to the presence in the amino acid mixt. of relatively high concns. of amino acids of the urea cycle (e.g. arginine and ornithine), as well as glutamic and aspartic acids.

Organic acidaemia and Hyperammonaemia: review.

Abstract: Recent clinical and experimental evidence on the effects of organic acids in producing or ameliorating hyperammonaemia is reviewed. The importance of hepatic mitochondrialN-acetylglutamate and its precursors, glutamate and acetyl-CoA, in the control of ureagenesis and thus blood ammonia levels is emphasized by recent work. The hypothesis is proposed that protein loads stimulate urea cycle activity via glutamate-induced changes inN-acetylglutamate concentration, while the effects of organic acids on ureagenesis are related in a predictable way to their effects on hepatic concentrations of acetyl-CoA.

Computer simulation of the urea cycle: trials for an appropriate model.

Abstract: Several models of the urea cycle with normal and deficient enzymes were simulated on a desk-top computer in order to test if a model fitting data of patients suffering from deficiencies of urea cycle enzymes could be obtained. Discrepancies were notable if only the four enzymes of the urea cycle were used. The effect of adding either n-acetylglutamate synthetase and CPS or the mitochondrial transport system of ornithine and its catabolism to the model were tested singly or combined. The use of the combined model with the rate constants adjusted to recent data of the human enzymes together with a transport system for N-acetylglutamate out of the mitochondrium improved the results. Computer simulations are thus a useful tool for testing concepts of the functioning of the urea cycle.

Control of urea cycle enzymes in rat liver by glucagon.

Abstract: A single injection of glucagon increased the levels of all five urea cycle enzymes in 28-day-old rats, but had no significant effect in 35-day-old animals. Rats weaned onto diets varying in protein content showed rapid adaptation in the levels of carbamoylphosphate synthetase, which were complete 4 days after weaning. Induction of the enzyme by glucagon was independent of the diet. In animals subjected to a controlled feeding regime, no circadian variation in the levels of urea cycle enzymes could be observed. However, the inducibility of the enzyme by glucagon was greatest if the hormone was given near the beginning of the period of darkness and feeding.

Regulation of Urea Cycle Enzymes in Transplantable Hepatomas and in the Livers of Tumor-bearing Rats and Humans

Abstract: The levels of the five enzymes of the urea cycle were measured in normal 5-week-old rats, in a transplantable hepatoma, and in the livers of tumor-bearing rats (host livers). The levels of all five enzymes were much lower in the hepatoma, although there was no exact correlation of the decrease in levels. In host livers, the levels were higher than in the tumors, but lower than in normal liver. The levels of all five urea cycle enzymes were positively correlated with dietary protein content in normal livers, in hepatomas, and in host livers. In fact, the hepatomas showed the greatest changes in response to diet. On all diets, the levels in host liver remained below those in normal liver, indicating that the decreased level was probably not due to preferential utilization of nutrients by the tumor. The levels of urea cycle enzymes in normal liver were not altered by a single injection of glucocorticoid, glucagon, or dibutyryl cyclic adenosine 3':5'-monophosphate. By contrast, in hepatoma, the levels were usually significantly elevated by the same treatment. In addition, the levels in host livers were always significantly elevated and were usually above those in normal animals, whether the latter were hormone treated or not. Injection of plasma from tumor-bearing rats into normal animals produced a decrease in the levels of all five enzymes; if glucagon was injected together with the plasma, large increases in levels were observed. This result supports the concept of a humoral factor produced by the tumor which affects the levels and the inducibility of urea cycle enzymes in host livers. Autopsied human primary hepatomas also showed levels of urea cycle enzymes below those in normal livers with host livers having intermediate values. A cell line derived from a human hepatoma showed induction of arginase by glucocorticoid in culture; in this, it resembled a cell line of the rat hepatoma. Tyrosine aminotransferase in human hepatoma cells was not induced by glucocorticoid; in this, it differed from the rat hepatoma cells where induction of this enzyme was observed.

[Variations in the plasma concentration of ornithine, citrulline and arginine in acute experimental liver failure].

Abstract: In pigs with acute hepatic failure, induced by one stage hepatic devascularization, plasma amino acids pattern changes markedly. Besides an increase in aromatic amino acid and a decrease in branched chain amino acid levels, urea cycle amino acids levels change. The content of ornithine and citrulline is enhanced, while arginine level drops down dramatically within six hours after surgery. These observations suggest that an enrichment of the arginine content of amino acids solutions recently proposed for the therapy of the hepatic encephalopathy might be useful.

Introduction to urea cycle symposium.

Abstract: When Donough O9Brien reviewed the field of metabolic diseases in 1965, he discussed disorders implicating carbamylphosphate synthetase (CPS), ornithine transcarbamylase (OTC), argininosuccinic acid (ASA) synthetase, and argininosuccinic acid lyase. Arginase defects had not yet been encountered. Several cases of argininosuccinic aciduria had been reported, but only single cases of CPS, OTC, and ASA synthetase defects had been described. The succeeding 15 years have seen much progress in this field, much of it reflecting the efforts of individuals participating in this program. My first contact with disorders of the urea cycle was in 1976 when a suspected OTC deficiency in a female infant was identified by Dr Kenneth Shaw, for a colleague of ours at the University of Southern California Medical Center. I was asked to help with the care of that infant, who died many months later of meningitis. Largely due to Ken9s indefatigable efforts in screening high-risk infants for metabolic diseases, we have seen various urea cycle defects identified in several other infants. However, we have sometimes encountered the unexpected in attempting to confirm the clinical diagnosis by enzyme analysis. The following comments on two patients exemplify the situation. Patient C.S. was in a coma on admission; his blood ammonia level was 895 µg/100 ml. Urinary orotic acid levels were elevated and our clinical impression was that the patient had a variant of an OTC defect. Analysis of the liver biopsy specimen revealed 35% CPS, 42% OTC, 24% ASA synthetase, 100% ASA lyase, and 31% arginase. Patient N.G. was born in a nearby medical center and shortly thereafter became lethargic, vomited, and lapsed into a coma.

The Production of Urea from Ornithine in Rat Hepatoma Cells Continuously Cultured in a Chemically Defined Medium

Abstract: R-Y121B cells derived from a rat Reuber hepatoma cell line have been grown serially in arginine-and glutamine-deprived, ornithine-supplemented Eagle's minimum essential medium. This cell line has the biochemical machinery to synthesize urea. It produces urea from ornithine via the urea cycle pathway. The synthesis of urea depends on the concentration of ornithine and acetylglutamate. There was no definite amino acid found to be the urea nitrogen source in R-Y121B cells.