I
`I
`
`I
`
`I
`
`SECOND EDITION
`
`Editors
`
`FREDERICK J .. SUCHY, M .. D.
`Professor and Chair
`Department of Pediatrics
`Mount Sinai School of Medicine
`Pediatrician-in-Chief
`Department of Pediatrics
`Mount Sinai Hospital
`New York, New York
`
`J. SOKOL, M.D.
`Professor
`Pediatric Liver Center and Liver Transplantation Program
`Department of Pediatrics
`University of Colorado Health Sciences Center
`Program Director
`Pediatric General Clinicial Research Center
`The Children's Hospital
`Denver, Colorado
`
`BALISTRERI, M.D ..
`WILLIAM
`Dorothy MM. Kersten Professor
`Department of Pediatrics
`University of Cincinnati College of Medicine
`Director
`Division of Pediatric Gastroenterology, Hepatology, and Nutrition
`and The Pediatric Liver Care Center
`Children's Hospital Medical Center
`Cincinnati, Ohio
`
`~~LIPPINCOTT WILLIAMS & WILKINS
`
`•
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`A Wolters Kluwer Company
`Philadelphia • Baltimore • New York • London
`Buenos Aires • Hong Kong • Sydney • Tokyo
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`Developmental Editor: Alexandra T. Anderson
`Production Editor: Donna J. Carty
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`
`© 2001 by LIPPINCOTT WILLIAMS & WILKINS
`530 Walnut Street
`Philadelphia, PA 19106 USA
`LWW.com
`
`All rights reserved. This book is protected by copyright. No part of this book may be
`reproduced in any form or by any means, including photocopying, or utilized by any
`information storage and retrieval system without written permission from the copyright
`owner, except for brief quotations embodied in critical articles and reviews. Materials
`appearing in this book prepared by individuals as part of their official duties as U.S.
`government employees are not covered by the above-mentioned copyright.
`
`Printed in the USA
`
`Library of Congress Cataloging-in-Publication Data
`
`Liver disease in children I edited by Frederick J. Suchy, Ronald J. Sokol,
`William F. Balistreri. -2nd ed.
`p.; em.
`Includes bibliographical references and index.
`ISBN 0-7817-2098-2
`1. Liver-Diseases. 2. Pediatric gastroenterology. I. Suchy, Frederick J.
`II. Sokol Ronald J. III. Balistreri, William F.
`[DNLM: 1. Liver Deseases-diagnosis-Child. 2. Liver Disease-physiopathology(cid:173)
`Child. 3. Liver Diseases-therapy-Child. WS 310L784 2001]
`RJ456.L5 L575 2001
`618.92'362-dc21
`
`00-056914
`
`Care has been taken to confirm the accuracy of the information presented and to
`describe generally accepted practices. However, the authors, editors, and publisher are
`not responsible for errors or omissions or for any consequences from application of the
`information in this book and make no warranty, expressed or implied, with respect to
`the currency, completeness, or accuracy of the contents of the publication. Application
`of this information in a particular situation remains the professional responsibility of
`the practitioner.
`The authors, editors, and publisher have exerted every effort to ensure that drug
`selection and dosage set forth in this text are in accordance with current
`recommendations and practice at the time of publication. However, in view of ongoing
`research, changes in government regulations, and the constant flow of information
`relating to drug therapy and drug reactions, the reader is urged to check the package
`insert for each drug for any change in indications and dosage and for added warnings
`and precautions. This is particularly important when the recommended agent is a new
`or infrequently employed drug.
`Some drugs and medical devices presented in this publication have Food and Drug
`Administration (FDA) clearance for limited use in restricted research settings. It is the
`responsibility of the health care provider to ascertain the FDA status of each drug or
`device planned for use in their clinical practice.
`
`10 9 8 7 6 5 4 3 2 1
`
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`

`
`ISORDERS OF TH UREA
`
`MICHAEL T. GERAGHTY
`SAUL W. BRUSU.OW
`
`The urea cycle serves to incorporate waste nitrogen atoms
`into urea (the major waste nitrogen product), and it is
`required for the de novo biosynthesis and degradation of
`argmme.
`A defect in the urea cycle has two consequences. First, it
`leads to hyperammonemia with the accumulation of nitro(cid:173)
`gen atoms in a variety of molecules, the pattern of which
`varies according to the specific enzymatic defect. Secondly,
`the amino acid arginine becomes an essential amino acid in
`all conditions except arginase deficiency.
`
`BIOCHEMISTRY
`
`The enzymes, substrates, and cofactors required for ureage(cid:173)
`nesis are shown in Fig. 3 5 .1. The urea cycle consists of five
`enzymes, and normal functioning depends on the presence
`of a number of mitochondrial membrane transporters (1).
`Carbamyl phosphate synthetase (CPS1) is a mitochondrial
`matrix enzyme that catalyzes the biosynthesis of carbamyl
`phosphate (CP) from ammonium and bicarbonate. N(cid:173)
`acetylglutamate synthetase (NAGS) catalyzes the formation
`of NAG from glutamate and acetyl-CoA. NAG is an essen(cid:173)
`tial cofactor for CPS 1 and may be an important regulator of
`ureagenesis. Ornithine transcarbamylase (OTC) is a mito(cid:173)
`chondrial matrix enzyme that catalyzes the biosynthesis of
`citrulline from ornithine and CP. Citrulline is then exported
`to the cytosol, where it condenses with aspartate via argini(cid:173)
`nosuccinic acid synthase (AS) to form argininosuccinate.
`This in turn is cleaved to arginine and fumarate by argini(cid:173)
`is subsequently hydrolyzed by
`nosuccinase. Arginine
`arginase (ARG I) to urea and ornithine. Ornithine can then
`be again transcarbamylated to citrulline. In addition to the
`enzymes mentioned above, ORNT1, a mitochondrial mem-
`
`M. T. Geraghty: Department of Pediatrics, Johns Hopkins University
`School of Medicine; and McKusick-Nathans Institute of Genetic Medicine,
`Johns Hopkins Hospital, Baltimore, Maryland 21287-4922
`S. W. Brusilow: Department of Pediatrics, Johns Hopkins University
`School of Medicine; and Department of Pediatrics, Johns Hopkins Hospital,
`Baltimore, Maryland 21287-4922.
`
`brane ornithine transporter, and Citrin, a mitochondrial
`membrane citrulline transporter, are required for normal
`functioning of the urea cycle (2,3).
`
`NITROGEN SOURCES FOR UREA
`
`Waste nitrogen disposal is a complex process reqmnng
`interorgan, intrahepatic, and intracellular compartmenta(cid:173)
`tion for the conversion of nitrogen not used for synthetic
`purposes to urea (Fig. 35.2). Free ammonium and aspar(cid:173)
`tic acid are the sole sources of nitrogen for ureagenesis.
`However, the pathways from amino acid nitrogen to
`ammonium and aspartate are less clear. Within the liver,
`both alanine and glutamate are transaminated to aspartate
`and incorporated into urea ( 4,5). These reactions take
`place predominantly
`in
`the periportal hepatocytes,
`emphasizing the role of metabolic zonation within the
`liver (6). Perivenous hepatocytes predominantly contain
`enzymes that catalyze the amidation of glutamate to glut(cid:173)
`amine or, alternatively, the deamination of glutamate to
`ammonium and ketoglutarate. A number of other amino
`acids may provide ammonium for ureagenesis by deami(cid:173)
`nation including histidine, tryptophan, threonine, and
`lysine.
`Extrahepatic sources of nitrogen for ureagenesis are
`derived from the gastrointestinal tract, the kidney, and
`muscle. Within the intestines, glutamine is converted to
`ammonium, citrulline, and alanine, all of which are
`released into the portal circulation (7,8). Ammonium and
`alanine are taken up by the liver, whereas citrulline is not,
`but rather is transported to the kidney for conversion to
`arginine (9). The kidney provides a waste nitrogen atom
`by catalyzing the synthesis of arginine from citrulline and
`aspartate via renal AS activity (10-13). Renal glutaminase
`also may supply ammonium directly for the CPS reaction.
`Within the muscle, alanine production via transamination
`of pyruvate represents an important nitrogen precursor for
`ureagenesis. The muscle is also a major source of gluta(cid:173)
`mine production (I 4-16).
`
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`

`
`828
`
`Liver Disease in Children
`
`*Ammonium +Bicarbonate
`
`¢CPS!
`
`Glutamate + acetyl CoA
`
`NAGS t
`
`Carbamyl phosphate
`
`-c~~'(IIE---
`
`N -acetylglutamate
`
`Mitochondrial matrix 8
`
`Ornithine
`
`8
`
`Citrulline
`
`------------;1 ORNT1 r-l --------------\(CITRIN ! - - - - - - - - - - (cid:173)
`Cytosol
`
`Ornithine
`
`Citrulline
`
`**UREA ~ ARGI
`
`AS ~ *Aspartate
`
`Arginine
`
`Argininosuccinate
`
`Fumarate
`
`FIGURE 35.1. Substrate and products involved in the urea cycle. The asterisks denote the waste
`nitrogen atoms. CPS1, mitochondrial carbamyl phosphate synthetase; NAGS, N-acetylglutamate
`synthetase; OTC, ornithine transcarbamylase; CITRIN, presumed citrulline mitochondrial carrier
`protein; AS, argininosuccinic acid synthetase; AL, argininosuccinic acid lyase; ARG1, hepatic
`arginase; ORNT1, mitochondrial ornithine transporter.
`
`FIGURE 35.2. Pathways of waste nitrogen
`synthesis from amino acids. Muscle is the
`major source of nitrogen for ureagenesis
`through its production of alanine and gluta(cid:173)
`mine. NH4, ammonium; Gin, glutamine; Ala,
`alanine; Asp, aspartic acid; Cit, citrulline; Glu,
`glutamic acid.
`
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`
`Disorders of the Urea Cycle
`
`829
`
`GENETICS
`
`Arginase
`
`Carbamyl Phosphate Synthtase
`
`The gene encoding the CPS 1 gene in humans has been par(cid:173)
`tially characterized. A full-length complementary DNA
`(eDNA) of 5,215 bp has been cloned and encodes an open
`reading frame predicted to encode a protein of 1,500 amino
`acids, including a mitochondrial leader sequence (17,18).
`Although the genomic structure has not been reported, the
`gene has been localized to chromosome 2q35 (19,20). Several
`mutations in affected individuals have been described (17 ,21).
`
`Ornithine Transcarbamylase
`
`The gene encoding ornithine transcarbamylase activity has
`been characterized in humans (22,23). It is located at chro(cid:173)
`mosome Xp21.1 (24). It spans 8 5 kb, contains 10 exons, and
`generates a 1 ,600-bp messenger RNA (mRNA). The pre(cid:173)
`dicted protein has 354 amino acids, the first 32 of which are
`a mitochondrial leader sequence. The protein is active as a
`homotrimer. The OTC gene is subject to X chromosome
`inactivation in females, contributing to the wide variation
`and severity seen in carrier females. Molecular analysis of
`affected individuals has demonstrated a wide variety of muta(cid:173)
`tions (25-28). Direct mutation analysis is accomplished by
`Southern blot analysis (,.., 10% of patients have deletions) or
`polymerase chain reaction amplification of the 10 exons and
`their intron boundaries followed by sequencing. Linkage
`analysis can be used to exclude or include the inheritance of
`the mutant gene where direct mutation analysis is unsuccess(cid:173)
`ful and where the pedigree structure is suitable.
`
`Arginosucdnic Add Synthase
`
`The gene encoding arginosuccinic acid synthase activity
`maps to chromosome 9q34. It covers 63 kb, has 16 exons,
`and generates an mRNA of 1,600 bp (29,30). The pre(cid:173)
`dicted protein has 412 amino acids and is active as the
`homotetrameric form. There are a large number of
`processed pseudogenes (i.e., homologous, intronless, and
`inactive sequences) mapped to several loci, including chro(cid:173)
`mosomes 2, 3, 4, 5, 6, 7, 9, 11, 12, X, andY (31). These
`probably arose by reverse transcription of mRNA followed
`by integration of copied sequences into genomic DNA. A
`large number of mutations have been described in the struc(cid:173)
`tural gene, the majority of which produce mRNA and no
`immunoreactive protein, suggesting most mutations in this
`disorder produce unstable protein products (32-34).
`
`Arginosucdnase
`
`The gene encoding arginosuccinase maps to chromosome
`7cen-q11.2. It is 35 kb long, divided to 16 exons, and
`encodes a predicted protein of 463 amino acids (35,36). The
`active enzyme is the homotetrameric form. Disease causing
`mutations have been described in affected patients (37,38).
`
`There are two forms of arginase activity. ARG 1 is expressed
`in the liver and red cells and is deficient in arginemia. The
`gene encoding ARG 1 maps to chromosome 6q23, is 11.5 kb
`in size, and contains 8 exons (39-42). The mRNA is 1.4 kb
`and encodes a predicted protein of 312 amino acids. Molec(cid:173)
`ular studies have shown that arginase deficiency is produced
`by a variety of mutations (43). A second arginase (ARG2) is
`expressed in a wide variety of tissues, including the kidney,
`brain, and gastrointestinal tract. ARG2 maps to chromosome
`14q24.1-24.3 and encodes a mitochondrial protein of 355
`amino acids (44,45). ARG2 appears to play a role in nitric
`oxide (NO) biosynthesis, as well as in ornithine, proline, and
`polyamine metabolism (46). To date, disorders due to defi(cid:173)
`ciency of this enzyme have not been described.
`
`N-Acetylglutamate Synthetase
`
`To date, the gene encoding this enzyme has not been iden(cid:173)
`tified or characterized.
`
`CLINICAL FEATURES
`
`The clinical presentation of patients with CPS, OTC, AS,
`and argininosuccinase deficiencies is very similar in that they
`all present with hyperammonemia. However, considerable
`variability occurs within and among each of these diseases.
`This variability relates to the different mutations found in
`each of the genes encoding these enzymes, to the pattern of
`inheritance (OTC is X linked), and finally to the relative tox(cid:173)
`icity of accumulated substrates or deficient products (Table
`35.1). It is convenient to divide the presentation of these dis(cid:173)
`orders into neonatal presentation and late presentation.
`
`NEONATAL ONSET
`
`In the neonatal period the presentation of these disorders is
`stereotypic and characterized by encephalopathy, respiratory
`alkalosis, and hyperammonemia. Infants are usually the
`product of a full-term normal pregnancy with no known pre(cid:173)
`natal or perinatal risk factors. Labor and delivery are normal,
`and the patient appears to be normal for at least 24 hours.
`Between 24 and 72 hours, the patient shows decreased feed(cid:173)
`ing, which usually proceeds to vomiting, increasing lethargy,
`hypothermia, and hyperventilation. The latter often leads to
`a suspicion of pulmonary disease. However, chest X-rays are
`often normal. Routine laboratory data generally show a res(cid:173)
`piratory alkalosis. Electrolytes are uninformative, and their
`very normality serves to exclude other causes of hyperam(cid:173)
`monemia. Serum urea nitrogen may be very helpful because
`it is often either absent or as low as 1 mg/dL. Without treat(cid:173)
`ment, the infant becomes comatose, requiring mechanical
`ventilation. Computed tomography scan of the brain often
`reveals only cerebral edema. If plasma ammonium is not
`
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`TABLE 35.1. ENZYMATIC AND BIOCHEMICAL ABNORMALITIES IN UREA CYCLE DEFECTS
`
`Enzyme Disorder
`
`Tissue
`
`Compartment
`
`Diagnostic Findings
`
`Diagnostic Tissue
`
`Gene (OMIM No.)
`
`N-acetyl glutamate
`synthetase
`
`NAG deficiency
`
`Liver, intestine,
`kidney (trace}, spleen
`
`Ornithine
`transcarbamylase
`(OTC)
`
`OTC deficiency
`
`Liver, intestine,
`kidney (trace)
`
`Carbamyl phosphate
`synthetase (CPS 1)
`
`CPS 1 deficiency
`
`Liver, intestine,
`kidney (trace)
`
`Argininosuccinic acid
`synthetase (AS)
`
`Citrullinemia
`
`Argininosuccinic
`acid lyase (AL)
`
`Argininosuccinic
`aciduria
`
`Liver, kidney, fibroblasts, Cytosol
`brain (trace)
`
`Liver, kidney, fibroblasts, Cytosol
`brain
`
`Mitochondrial matrix Citrulline: absent/trace
`Arginine, ornithine: low
`Glutamine, alanine: increased
`No orotic aciduria
`Mitochondrial matrix Citrulline: absent/trace
`Arginine, ornithine: low
`Glutamine, alanine: increased
`Orotic aciduria present
`Mitochondrial matrix Citrulline: absent/trace
`Arginine, ornithine: low
`Glutamine, alanine: increased
`Orotic aciduria absent
`Citrulline: >1,000 j.tM
`Arginine, ornithine: low
`Glutamine, alanine: increased
`Citrulline: > 100 j.tM
`Argininosuccinic acid
`and anhydrides present
`Arginine, ornithine: low
`Glutamine, alanine: increased
`Arginine markedly increased
`Glutamine elevated
`Diamino aciduria (arginuria, lysinuria,
`cystinuria, and ornithinuria)
`Orotic aciduria
`
`t--<
`;:;·
`"'
`.....
`t;
`~-
`"'
`~ ....
`"'
`~-
`Q
`~
`~ ;:s
`
`Liver biopsy
`
`Not known
`(237310)
`
`Liver biopsy
`
`Liver biopsy
`
`Fibroblasts
`
`Fibroblasts
`
`Xp21
`(311250)
`
`2q35
`(237300)
`
`9q34
`(215700}
`
`7cen-q11.2
`(207900}
`
`Erythrocytes
`
`6q23
`(207800}
`
`Arginase (ARG1)
`
`Arginemia
`
`Liver, erythrocytes
`
`Cytosol
`
`Transporters
`Solutecarrier family 7
`member 7 (SLC7 A7)
`
`Lysinuric protein
`intolerance
`
`Mitochondrial
`ornithine transporter
`(ORNT1)
`
`Citrin (SLC25A 13}
`
`HHH syndrome
`(hyperammonemia,
`hyperornithinemia,
`homocitru II in em ia)
`Adult onset type II
`citrullinemia (CTLN2)
`
`Kidney, leukocytes, small Plasma membrane
`intestine; trace in
`other tissues including
`liver and heart
`Liver, pancreas, low
`levels in other tissues
`including kidney
`
`Mitochondrial
`membrane
`
`Lysinuria, arginuria, ornithinuria
`Low/normal levels of lysine, ornithine
`1 arginine in plasma
`High glutamine serine threonine
`Hyperornth i nem ia, homocitru IIi nem ia Fibroblasts
`(may comigrate with methionine)
`
`14q11.2
`(222700}
`
`13q14
`(238970)
`
`Most tissues including
`liver and kidney
`
`Mitochondrial
`membrane
`
`Citrullinemia
`
`Liver enzyme
`abnormal, other
`tissues normal
`
`7q21.3
`(603471)
`
`
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`

`
`Disorders of the Urea Cycle
`
`831
`
`measured, the infant's death is often ascribed to sepsis,
`intracranial hemorrhage, or some other disease commonly
`associated with prematurity, despite the patient being a full(cid:173)
`term infant. Family history is often neglected, a history of
`consanguinity, neonatal sibling deaths, or neonatal male
`deaths on pedigree analysis may be noted.
`The finding on increased plasma ammonium level should
`direct diagnostic efforts toward an inborn error of metabo(cid:173)
`lism. The differential diagnosis of hyperammonemia in the
`neonate includes disorders of the urea cycle, organic acidurias,
`fatty acid oxidation defects, lactic acidosis, respiratory chain
`disorders, herpes simplex infection, and transient hyperam(cid:173)
`monemia of the newborn (a poorly understood disease, char(cid:173)
`acterized by symptomatic pulmonary disease within the first
`24 hours oflife and severe hyperammonemia) (47-50). Care(cid:173)
`ful analysis of the basic biochemical laboratory indices can
`help distinguish between these varying causes. Organic
`acidurias are frequently associated with an increased anion gap
`metabolic acidosis. There may or may not be hypo- or hyper(cid:173)
`glycemia. Fatty acid oxidation defects are often associated with
`hypoglycemia, a normal anion gap, hyperchloremic acidosis,
`and abnormal liver function tests. There may be an associated
`myopathy or cardiomyopathy. Respiratory chain disorders
`and other causes of lactic acidemia are associated with a meta(cid:173)
`bolic acidosis and an increased anion gap, in contrast to the
`classical presentation of primary urea cycle defects with respi(cid:173)
`ratory alkalosis and normal electrolytes. A definitive diagnosis
`of the above conditions can generally be made on the basis of
`plasma amino acid analysis and urine organic acid analysis,
`together with quantitative and qualitative carnitine/acylcarni(cid:173)
`tine profiles or an acylglycine profile.
`Quantitative plasma amino acids allow the differentiation
`between the primary urea cycle defects (51). AS deficiency is
`characterized by plasma citrulline levels of 1,000 to 5,000 ~
`(normal level 10-20). In argininosuccinase deficiency, cit(cid:173)
`rulline levels usually range from 100 to 300 ~and are asso(cid:173)
`ciated with the finding of high concentrations of argininosuc(cid:173)
`cinic acid and its anhydrides, not· found in normal plasma.
`Argininosuccinic acid may co migrate with isoleucine and thus
`may be missed by the unwary chromatographer. Citrulline is
`the product of CPS and OTC, and thus it is undetectable or
`nearly so in plasma of hyperammonemic patients suffering a
`deficiency in one of these enzymes. CPS and OTC deficien(cid:173)
`cies can be distinguished from each other by the level of orotic
`acid in the urine. High levels occur in OTC deficiency as a
`consequence of diversion of accumulated mitochondrial CP
`to the cytosolic pyrimidine synthetic pathways. Orotic acid is
`absent in patients with CPS deficiency. High levels of plasma
`glutamine and alanine are frequently found in patients with
`hyperammonemia due to urea cycle defects, whereas there is a
`reduction in the levels of ornithine and arginine. Finally, the
`diagnosis of CPS deficiency is usually made by exclusion of
`the above findings. CPS and OTC or NAGS deficiencies can
`be confirmed on liver biopsy, whereas enzyme assays on fibro(cid:173)
`blasts or red cells confirm the diagnosis in AS, argininosucci(cid:173)
`nase, and arginase deficiencies, respectively.
`
`LATE ONSET
`
`All of the four diseases, CPS, OTC, AS, and argininosuccinase
`deficiencies may present later in life (52-60). The clinical fea(cid:173)
`tures relate to episodic hyperammonemia. In. infancy, it may
`be associated with weaning from breast milk or changing from
`low protein milk formula to cow's milk. In older children and
`adults, the symptoms are often related to high-protein meals
`or infection. Frequently no obvious cause is found. Protein
`avoidance is often a notable feature in the history, and patients
`often select a low protein diet. The major symptoms of these
`episodes of hyperammonemia include episodic vomiting and
`abnormal mental status. The latter is manifested by lethargy,
`somnolence often progressing to coma, irritability, agitation,
`combativeness, disorientation, and ataxia. Seizures, delayed
`physical growth, and developmental delay are common,
`although there are reports of normal development. The
`episodes often disappear with cessation of protein intake or
`intravenous infusion of glucose. Laboratory tests often reveal
`a respiratory alkalosis, the presence of which should always
`prompt a search for hyperammonemia. Hepatomegaly and
`hair abnormality (trichorrhexis nodosa) are common findings
`in argininosuccinase deficiency. Diagnostic delay and errors
`are common in the late-onset group. Symptoms have been
`attributed to colic, gastroenteritis, cyclical vomiting, hyperac(cid:173)
`tivity, encephalitis, Reye's syndrome, epilepsy, hepatitis, drug
`toxicity, brain tumor, and child abuse ( 61). The major symp(cid:173)
`toms of arginase deficiency include progressive spastic
`tetraplegia, seizures, psychomotor retardation, and growth
`failure (62-70). Symptoms may occur early in infancy, includ(cid:173)
`ing irritability, unconsolable crying, anorexia, vomiting, pro(cid:173)
`tein avoidance, and delayed developmental milestones. Symp(cid:173)
`tomatic hyperammonemia may occur, often with severe
`encephalopathy. Laboratory abnormalities
`include mild
`hyperammonemia, hyperarginemia as high as 1,500 ~(nor­
`mal 15-115), dibasic aminoaciduria, and orotic aciduria.
`Plasma glutamine levels may be increased.
`The large number of girls in the late-onset group is a con(cid:173)
`sequence of symptomatic OTC deficiency in females. OTC
`deficiency is an X-linked disorder with a wide variability of
`phenotypic expression (71,72). Asymptomatic carriers of the
`OTC mutation present a special at-risk group. Approximately
`two thirds of these women have a disorder of nitrogen home(cid:173)
`ostasis as shown by abnormally high plasma glutamine levels
`and a mean plasma ammonium level significantly higher than
`control women. Carrier females appear to be at some small
`risk for postpartum hyperammonemic encephalopathy, some(cid:173)
`times mistaken for postpartum psychosis. However, contrary
`to anecdotal reports, women OTC carriers do not have an
`increased incidence of migraine headaches (73). It is unclear
`how this group responds to other common late-onset diseases
`such as coronary artery disease or diabetes mellitus, which
`may cause significant catabolic stress.
`The differential diagnosis for late-onset hyperammonemic
`encephalopathy includes CPS, OTC, AS, and argininosucci(cid:173)
`nase deficiencies, the HHH (hyperammonemia, hyperor-
`
`
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`832
`
`Liver Disease in Children
`
`nithinemia, and homocitrullinuria) syndrome, lysinuric pro(cid:173)
`tein intolerance (LPI), adult -onset citrullinemia, organic
`acidurias, fatty acid oxidation defects, Reye's syndrome, liver
`disease, general malignancy, and ammoniagenesis in an
`infected bladder or ureter (74-76). Again, careful analysis of
`basic biochemical data will help in reaching a diagnosis.
`Organic acidurias are suggested by the presence of an increased
`anion gap metabolic acidosis, with or without hypo- or hyper(cid:173)
`glycemia. Analysis of urine by gas chromatography/ mass spec(cid:173)
`trometry will usually result in the definitive specific diagnosis.
`Likewise, fatty acid oxidation defects are most often associated
`with a hypoketotic hypoglycemia and a normal anion gap, and
`analysis of an acylcarnitine or acylglycine profile will usually
`result in a specific diagnosis. As before, plasma amino acids,
`particularly the citrulline level, are crucial in distinguishing
`between the primary urea cycle defects. Citrulline levels are
`usually greater than 1, 000 ~ in AS deficiency, and between
`100 and 300 ~ in argininosuccinase deficiency, where they
`are associated with increased concentrations of argininosuc(cid:173)
`cinic acid and its anhydrides. Citrulline is also mildly elevated
`in the HHH syndrome, where there are high concentrations
`of ornithine and homocitrulline. The latter compound may
`co migrate with the methionine and be mistaken for such. CPS
`deficiency and OTC deficiency cannot be distinguished by
`amino acid analysis. However, OTC deficiency is character(cid:173)
`ized by large increases in orotic acid. CPS deficiency is sug(cid:173)
`gested by exclusion, although family history may dearly dis(cid:173)
`tinguish between the two disorders because OTC deficiency is
`an X-linked condition. Measurement of hepatic CPS and
`OTC deficiency should be definitive for CPS deficiency but
`may be ambiguous for OTC deficiency. Because this disorder
`is X linked, Lyonization results in a mosaic of hepatocytes
`within the liver, giving a wide range ofOTC activity from nor(cid:173)
`mal to completely affected in female carriers (77).
`Female carriers of OTC deficiency represent a particular
`diagnostic problem. These can be identified on the basis of
`pedigree analysis. The birth of two affected children or one
`affected child and the presence of an affected first-degree
`relative imply obligate heterozygosity.
`A recent study showed that the birth of a single affected
`male results from a new mutation 7% of the time (78). Con(cid:173)
`versely, the birth of a symptomatic daughter resulted from a
`new mutation 80% of the time. This deviation from Hal(cid:173)
`dane's rule was suggested to result from a greater mutation rate
`for point mutations in sperm. A second study confirmed these
`results among singleton affected females and showed that
`33% of mothers of singleton affected males did not carry the
`mutation (73). Biochemically, two thirds of clinically normal
`obligate heterozygotes have plasma glutamine levels greater
`than 1,000 ~ (71). The finding of orotic aciduria is another
`useful indication of heterozygosity. Finally, DNA analysis,
`either by direct mutation detection or by linkage analysis, is
`increasingly used and is the method of choice. The allopuri(cid:173)
`nol test is a simple, safe, and sensitive test for determination
`of carrier status in postpubertal women that can be used if
`DNA analysis is unrevealing or ambiguous (79). Oxypurinol
`
`monophosphate (a reaction product of allopurinol) inhibits
`orotidine monophosphate decarboxylase and allows easier
`detection of orotidine. The production of this compound is
`already increased in heterozygotes for OTC deficiency as a
`consequence of CP accumulation and shunting into pyrimi(cid:173)
`dine biosynthesis. Protein and alanine loading tests carry the
`risk ofhyperammonemia and are not advisable (80-83).
`
`PATHOPHYSIOLOGY
`
`Ammonium appears to be the major cause of the acute
`encephalopathy seen in patients with urea cycle defects. In a
`primate animal model, increasing hyperammonemia was asso(cid:173)
`ciated with decreased spontaneous activity, disinterest in sur(cid:173)
`roundings, vomiting, somnolence progressing
`to coma,
`seizures, absence of corneal reflexes, apnea, progressive
`increase in intracranial pressure, and respiratory alkalosis (84).
`Electroencephalographic recordings showed a slow-wave
`appearance correlating with the clinical and biochemical sta(cid:173)
`tus. Gross neuropathologic changes included brain swelling,
`flattening of cortical gyri, and herniation of cerebellar tonsils.
`Light and electron microscopy revealed astrocyte swelling
`with abnormal mitochondria. However, neurons, axons, den(cid:173)
`drites, and synapses all appeared unaffected. Thus, the physi(cid:173)
`ologic response to hyperammonemia appears to be swelling of
`the astrocyte leading to alterations of cerebral blood flow,
`brain swelling, and increased intracranial pressure in the
`absence of neuronal pathology.
`Several lines of evidence suggest that these pathologic
`changes are mediated by excessive glutamine (Fig. 35.3) (85).
`It has been shown that plasma glutamine levels increase prior
`to hyperammonemia and that there is a strong correlation
`between plasma glutamine and ammonium levels (86,87).
`During hyperammonemic coma, the cerebral spinal gluta(cid:173)
`mine concentrations in patients with OTC deficiency and
`argininosuccinase deficiency are extraordinarily high: 6,300
`and 8,660 ~, respectively (normal limit 373-855) (88,89).
`Magnetic resonance spectroscopy has been used to show that
`hyperarnmonemic encephalopathy in OTC deficiency is
`related to increased glutamine levels in the brain (90). Finally,
`it has been well documented that ammonium is rapidly incor(cid:173)
`porated into brain glutamine and that the astrocyte is the site
`of this incorporation (91,92). Astrocytes are rich in glutamine
`synthetase, and it has been suggested that some of the edema
`associated with hyperammonemia ·is a consequence of the
`osmotic affects of an increased glutamine content of the astro(cid:173)
`cyte, leading to astrocyte swelling. Indeed, animal studies have
`shown that the cerebral edema associated with hyperarn(cid:173)
`monemia can be prevented by inhibiting glutamine accumu(cid:173)
`lation. This suggests that hyperammonemia is necessary but
`not sufficient to produce cerebral edema (93). Similar studies
`show that the prevention of glutamine accumulation in
`hyperammonemic rats averted the alterations in brain energy
`metabolism observed in hyperammonemic rats, in which glu(cid:173)
`tamine content increased (86).
`
`
`Page 8 of 18
`
`Owner Ex. 2015
`Par Pharm. v. Horizon
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`

`
`Hyperammonemia
`
`~
`Astrocyte Glutamine Accumulation
`I
`
`Regulatory
`
`err
`Cerebral Edema___.. ENCEPHALOPATHY J
`~
`t
`
`Astrocyte Swelling __ ___ --~~~o- Astrocyte Dt.ftmction
`
`Brain Stem Compression
`
`Disorders of the Urea Cycle
`
`833
`
`the
`representation of
`FIGURE 35.3. Diagrammatic
`for hyperammonemic
`pathophysiology
`responsible
`encephalopathy. The primary event is the activation by
`ammonia of glutamine synthetase within the astrocyte,
`leading to the accumulation of glutamine within the
`astrocyte. This has a dual effect: (a) glutamine serves as
`an intracellular osmolyte, causing entry of water and
`astrocyte swelling and cerebral edema, and (b) the
`swollen astrocyte and/or the high glutamine concentra(cid:173)
`tion causes astrocyte dysfunction. Regulatory osmolytes
`taurine) may minimize swelling.
`(myoinositol and
`[Reprinted from Brusilow SW, Horwich AL. Urea cycle
`enzymes. In: Scriver C, Beaudet A, Sly W, et al., eds. The
`metabolic and molecular bases of inherited disease, 8th
`ed. New York: McGraw-Hill (in press); with permission.]
`
`Substrate accumulation also m