Hyperammonemia

FOR SEMINAR

1. vvydehindraneel 1 st YEAR PG BIOCHEMISTRY OSMANIA MEDICAL COLLEGE HYDERABAD INDIA

2. Hyperammonemia a metabolic disturbance characterised by an excess of ammonia in the blood

3. Ammonia is a normal constituent of all body fluids . At physiologic pH, it exists as ammonium ion . adults 15 - 45g/ dL or 11 - 32 mol/L Children 40 - 80 g/ dL or 28 - 57 mol/L Newborns 90 - 150 g/ dL or 64 -107 mol/L

4. Sources of ammonia bacterial hydrolysis of urea and nitrogenous compounds in the intestine the purine -nucleotide cycle amino acid transamination in skeletal muscle, metabolic processes in the kidneys and liver

5. Ammonia is produced from dietary AA and by catabolism of amino acids, amines, nucleic acids, glutamine glutamate (nitrogenous wastes) ( skeletalmuscle ).

6. coliforms ,anaerobes (colon , cecum ) convert dietary AA and urea into ammonia The ammonia is absorbed into the portal circulation, taken up by the liver and converted in the liver, via the urea cycle, into urea.

7. Urea is then excreted into the gastrointestinal system (producing a futile cycle) and into the urine Of total ammonia , 80-90% into the urea cycle, 10-20% metabolised by kidney, heart, and brain. 

8. It is a product of the catabolism of protein. It is converted to the less toxic substance urea prior to excretion in urine by the kidneys. The metabolic pathways that synthesize urea are located first in the mitochondria and then into the cytosol . The process is known as the urea cycle,.

9. Types Primary vs. secondary Primary hyperammonemia is caused by several inborn errors of metabolism that are characterised by reduced activity of any of the enzymes in the urea cycle.

10. Secondary hyperammonemia is caused by inborn errors of intermediary metabolism characterised by reduced activity in enzymes that are not part of the urea cycle e.g. . Propionic acidemia , Methylmalonic acidemia or dysfunction of cells that make major contributions to metabolism (e.g . hepatic failure ).

11. N- acetylglutamate synthetase deficiency carbamoyl phosphate synthetase I ornithine transcarbamylase Specific types Type I Hyperammonemia Type II Hyperammonemia Hyperammonemia , type III

12. hyperinsulinism hyperammonemia syndrome ( glutamatedehydrogenase 1) hyperornithinemia-hyperammonemia-homocitrullinuria syndrome ( ornithine translocase hyperlysinuria with hyperammonemia

13. Methylmalonic acidemia Isovalemic acidemia Propionic acidemia Carnitine palmitoyl transferase II deficiency Transient hyperammonemia of the newborn, specifically in the preterm.

14. Causes Enzyme defects in urea cycle N - Acetylglutamate Synthetase (Nags) Deficiency: ↓Nags ↓Nag Nag is an activator of CPS I autosomal recessive . if ↓ acetyl- CoA ↓ NAG

15. Carbamoyl phosphate synthetase I (CPS I) deficiency : nag HCO 3 + NH 4 ----------------- CARBAMOYL PHOSPHATE CPS I 2atp mg 2adp + pi

16. Autosomal recessive short arm of chromosome 2 hepatic mitochondria. first day of life. infants die in the neonatal period

17. Ornithine transcarbamoylase (OTC) deficiency: ORNITHINEMIA mitochondria. carbamoyl po 4 + ornithine ------- ---- citrulline OTC Citrulline is then transported out of the mitochondria.

18. absence of enzyme,carbamoyl phosphate enters cytosol participates in pyrimidine synthesis in presence of CPS II . most common urea cycle defect incidence of 1 case in 14,000 persons. X-linked trait

19. . Neonatal onset is seen in males who have null mutations , thus no residual enzyme activity. Males who and females who are heterozygous for OTC deficiency have significant residual enzyme activity present later with quite variable clinical pictures.

20. Thus, as many as 60% of OTC deficiency diagnoses are made in non-neonates. The oldest reported patient was aged 61 years Mother of affected child also ehibits hyperammonemia and aversion to protein foods Blood ,urine , csf :↑glutamine, ↑ ammonia, ↑ ornithi ne

21. Argininosuccinic acid synthetase (AS) deficiency: CITRULINEMIA Citrulline combines with aspartate to form argininosuccinic acid. AS deficiency results in citrullinemia . Onset is usually between hours 24 and 72 of life, autosomal recessive. chromosome 9.

22. TWO TYPES OF DEFICIENCY: ONE TYPE :mutation in regulatory gene : enzyme is absent in liver . normal for citulline Other type : mutation in structural gene . Amount of enzyme normal in liver .affects catalytic site and has abnormally high km value for citrulline

23. Clinically : mental retardation , ammonia toxicity Biochemically : blood and csf : increased ammonia , ↑ citrulline Urine : large quantities (1—2gm/day ) of citrulline excreted Feeding arginine in these patients enhances citulline excretion

24. Argininosuccinic lyase (AL) deficiency: This enzyme cleaves argininosuccinic acid to yield fumarate and arginine . The lack of this enzyme leads to argininosuccinic aciduria . It is the second most common urea cycle disorder . chromosome 7. autosomal recessive

25. Enzyme deficiency seen in liver ,kidney brain , RBC. Early diagnosis can be made by demonstrating by demonstrating enzyme defeciency in RBC from cord blood and in amniotic fluid by amniocentecis

26. Terminates fatally in early life. Symptoms appear in the neonatal period or later in life. (2 yrs) Abnormally fragile hair ( trichorrhexis nodosa ) observed in these infants of age 2 weeks.

27. Arginase deficiency : the final step arginine ---------urea and ornithine . argininemia , neurotoxicity chromosome 6q23 . least frequent

28. Hyperammonemia is not severe uneventful. progressive spastic diplegia quadriplegia, intellectual impairment, recurrent vomiting, delayed growth, seizures.

29. hyperinsulinism hyperammonemia syndrome ( glutamatedehydrogenase 1) PATIENT : Since the neonatal period, a white girl had been treated for hyperammonemia and postprandial hypoglycemia with intermittent hyperinsulinism . ammonia 100 to 300 micromol /L and was independent of the protein intake.

30. METHODS: Enzymes of the urea cycle glutamine synthetase , glutamate dehydrogenase (GDH) were assayed in liver and/or lymphocytes.

31. RESULTS: The activity of hepatic GDH ↑ ratio glutamine/blood ammonia low. Oral -N- carbamylglutamate resulted in ↓ ammonia.

32. CONCLUSION : glutamate ---------- GDH- ---------------------2-oxoglutarate ↓ glutamate needed for the synthesis of N- acetylglutamate , the catalyst of the urea synthesis ↓ glutamate ----------------------↓ glutamine by glutamine synthetase . GDH stimulates the release of insulin,.

33. Organic acidemias ketosis , acidosis , hyperammonemia accumulation of CoA derivatives of organic acids, which inhibit the formation of NAG the activator of CPSI in liver .

34. Disorders in this group include the following: Isovaleric acidemia Propionic acidemia Methylmalonic acidemia Glutaric acidemia type II Multiple carboxylase deficiency beta- ketothiolase deficiency

35. Congenital lactic acidosis ↑lactate (10-20 mmol /L), ↑ lactate/ pyruvate ratio, metabolic acidosis, ketosis. Hyperammonemia and citrullinemia in some cases. Pyruvate dehydrogenase deficiency Pyruvate carboxylase deficiency Mitochondrial disorders

36. ↓PYRUVATE DEHYDROGENASE ↓ acetyl CoA ↓ NAG

37. Fatty acid oxidation defects Deficiency of medium- or long-chain acyl CoA dehydrogenase hyperammonemia secondary to hepatic dysfunction. .

38. Systemic carnitine deficiency: Carnitine :transport of long-chain fatty acids into mitochondria. ↑liver transaminases , hepatomegaly ↑ammonium , liver dysfunction.

39. Dibasic amino acid transport defects Lysinuric protein intolerance hyperlysinuria with hyperammonemia ↓ membrane transport ↓ lysine, ↓ ornithine ↓ arginine . Citrulline ( orally)------------ ↓ammonia it is transported by a different mechanism in intestine. normal neurologic development when treated.

40. Hyperammonemia-hyperornithinemia-homocitrullinuria (HHH) first few weeks of life seizures, feeding difficulty, altered level of consciousness .

41. A defect in transport of ornithine cytosol ----│--- → mitochondria ornithinemia ↑ disruption of the urea cycle causes ↑ ammonemia . In absence of ornithine , mitochondrial→ carbamoyl po 4 + lysine------→ homocitrulline

42. Transient hyperammonemia of the newborn premature day 1,day2 before introduction of protein Hyperammonemia ↑ ↑ ↑ hemodialysis . 30% die 35-45% ↓neurologic development. slow maturation of the urea cycle function .

43. seizures ↓consciousness, fixed pupils loss of oculocephalic reflex. HIE, ICH

44. first 24 hours of life. ↑ ↑ ↑ ammonia elevated SGOT Asphyxia

45. Reye syndrome acquired influenza A or B or varicella aspirin ingestion. cerebral and hepatic dysfunction — .

46. vomiting, altered level of consciousness, seizures, cerebral edema , and hepatomegaly without jaundice. ↑ liver transaminases , hyperammonemia , lactic acidosis

47. Renal Urinary tract infection with a urease -producing organism, such as Proteus mirabilis, Corynebacterium species, or Staphylococcus species , can produce a hyperammonemic state. high urinary residuals and an alkaline pH .

48. Other causes neonatal herpes simplex pneumonitis , The increase in ammonia level resulted from protein catabolism caused by prolonged hypoxia. Parenteral hyperalimentation : Increased nitrogen load in patients receiving parenteral alimentation can cause hyperammonemia . thyroid disease and Hashimoto encephalopathy Hyperammonemia is a rare but severe complication of multiple myeloma and is with high mortality. [17]

49. Drugs Valproate topiramate Carbamazepine Salicylate

50. Pathophysiology ↑ ammonia ↑ECF glutamate in the brain ↑ N -methyl D- aspartate (NMDA) receptor. ↓ ATP .

51. ↓ phosphorylation by protein kinase C. ↑ Na + /K + - ATPase ↓ATP depletion Activation NMDA receptor → seizures

52. . ↓ gap–junction channel connexin 43, water channel aquaporin 4 K channel, ∆ K and water transport brain edema.

53. ∆ astrocyte morphology

54. p53, a tumor suppressor protein transcriptional factor, Activation of p53 astrocyte swelling glutamate uptake ↓ brain edema

55. ↑ lactate, pyruvate , glutamine, glucose, ↓ glycogen, ketone bodies, glutamate.

56. ↑inhibitory neurotransmission as a consequence of 2 factors. ↑ glutamate ↓ glutamate receptors

57. ∆ glutamate-nitric oxide- cGMP pathway ∆ signal transduction associated with NMDA receptors cognition and learning

58. ↑ GABAergic tone from BZDreceptor overstimulation Activation of GABA(A) receptors reduces the function of the pathway. ↓ intellectual function, ↓consciousness, coma.

59. ↑ the transport of aromatic AA ( eg , tryptophan) across BBB ↑ serotonin, anorexia

60. Astrocyte edema ↑ ROSand ↑NO species, RNA oxidation ↑ intracellular zinc. RNA oxidation ↓postsynaptic proteins involved in learning and memory consolidation

61. Epidemiology 1 in 25,000 live births

62. Presentation Family history unexplained neonatal deaths or undiagnosed chronic illness. males affected Suggestive of OTC deficiency, X-linked trait . Consanguinity ↑risk of inheriting disorder . .

63. Early-onset : neonatal period. The baby is well day 1,2 lethargy, irritability, poor feeding, vomiting. hyperventilation ,grunting respiration, seizures ammonia level of 100-150 µmol/L, 2-3 times the reference range.

64. Late-onset hyperammonemia typically is due to urea cycle disorders, present later in life. Adults with partial enzyme deficiency become symptomatic postpartum stress, heart-lung transplant, short bowel and kidney disease, parenteral nutrition with high N intake, gastrointestinal bleeding.

65. Intermittent ataxia: unstable gait , dysmetria . periodic ↑ ammonia Intellectual impairment : Episodic hyperammonemia may produce subtle intellectual deficits even in clinically asymptomatic individuals. Failure to thrive: ( poor feeding and frequent vomiting )

66. Gait abnormality: In arginase deficiency, spastic diplegia , ( toe walking) Behavior disturbances : sleep disturbances, irritability, hyperactivity, manic episodes, psychosis. Epilepsy Intractable seizures in a few patients secondary to urea cycle defect

67. Recurrent Reye syndrome Episodic headaches cyclic vomiting Protein avoidance: Females with OTC deficiency

68. Physical Dehydration ← vomiting Tachypnea : stimulation of the medullary center of respiration by the ammonium ion Hypotonia ← acute stress

69. Bulging fontanelle :↑ICP odor of "sweaty feet" in isovaleric acidemia abnormally fragile hair in argininosuccinic aciduria . Infants with argininosuccinic lyase ↓: hepatomegaly

70. Other diagnostic considerations the neonatal period : nonspecific c/f c/f indicate distress sepsis, intracranial hemorrhage , cardiac disease, gastrointestinal obstruction should be ruled out Plasma ammonium level should be determined in all such scenarios. :

71. NH3 >200 µmol/L), Plasma and urinary amino acids Urinary organic acids Serum glucose Arterial blood gases Bicarbonate Lactate Citrulline , acylcarnitine Urinary ketones Urinary orotate

72. Arterial blood gas analysis: acid-base status respiratory alkalosis → a urea cycle defect stimulation of the central respiratory drive ↓ hyperventilation

73. Serum amino acid tests ↑ Glutamine , alanine ↑ in all urea cycle defects except for arginase deficiency . ↓ Citrulline mildly in CPS/NAGS and OTC deficiencies but ↑markedly in AS deficiency and moderately in AL deficiency.

74. Arginine level ↑ arginase deficiency ↓mildly in all the other enzyme deficiencies of the urea cycle. Argininosuccinic acid level ↑ in AL deficiency

75. Urinary orotic acid tests : ↑markedly in OTCdeficiency ↑mildly in other enzyme deficiencies for CPS/NAGS deficiency, in which it is ↓ mildly

76. Urinary ketone tests: ketosis indicates an organic acidemia Plasma and urinary organic acid tests: These levels screen for an organic acidemia

77. Enzyme assays: tissue specimens obtained by percutaneous liver biopsy CPS, NAGS, and OTC deficiency red blood cells (for arginase deficiency),

78. fibroblast from skin biopsy (ASS, ASL, and HHH), intestinal mucosa (CPS, OTC). replaced by genetic analysis. It is still indicated in selected cases with negative genetic testing

79. DNA mutation analysis is the method of choice in confirming the diagnosis of UCD as it is clinically available for all genes of the urea cycle. Heterozygote identification in OTC-deficient pedigrees

80. Allopurinol loading test : establishes the carrier status of women at risk for OTC deficiency. After a loading dose of allopurinol , urinary orotidine excretion is ↑ greatly in carriers. DNA analysis: determine the presence of a mutation at the OTC locus.

81. Antenatal diagnosis: DNA analysis on chorionic villus or amniotic fluid cells, measurements of amniotic fluid metabolites or enzyme activities in the amniotic cells, chorionic villi , fetal liver, fetal RBC

82. Imaging Studies Neuroimaging : CT or MRI of the brain cerebral edema chronic liver disorders is hyperintense signal in the globus pallidum due ↑manganese . 21]

83. Diffuse Brain Edema Findings: Absence of sulcal markings and poor differentiation of white and grey matter.

84. MR spectroscopy: ↑glutamine/glutamate ↓ myoinositol and choline signals. diffusion tensor imaging damage to corticospinal tracts : arginase deficiency. [ Multiple strokelike lesions MRI finding in a patient with HHH syndrome

85. Histologic Findings prominent Alzheimer type II astrogliosis

86. Assay Measurement of ammonia is problematic as it is very unstable . Arteria l blood samples are preferable to venous blood samples as results are more consistent. Heparinized plasma samples are preferable to serum

87. Due to the instability of ammonia and leakage of ammonia from RBC , samples need to be assayed for ammonia as soon as possible after sample collection and maintained at 4 C (or on ice) until assay (stable for a maximum of 3 hours under these conditions)

88. The sample must be separated from cells as soon as possible as leakage of ammonia from erythrocytes occurs within 30 minutes , resulting in artifactually high values. This is not easy to accomplish under most situations, therefore ammonia assays are not routinely performed.

89. . Furthermore, any ammonia in the environment (air, water supply ) can contribute to the ammonia in the patient sample. Control samples (from a clinically healthy animal) should always be run in conjunction with patient samples, to ensure that sample collection and handling are not responsible for elevations in ammonia.

90. MEASUREMENT OF AMMONIA IN BLOOD fasted : 6 hours Plasma ammonia levels exercise, smoking, GI bleeding, blood transfusions, high protein intake medications. ↑

91. . Heparin is the preferred anticoagulant, because it has been shown to reduce red blood cell ammonia production. The patient’s arm relaxed ,because muscle exertion leads to↑ venous ammonia levels Prolonged application of a tourniquet or fist-clenching while obtaining the blood sample avoided

92. The blood sample should be drawn into a chilled, sodium heparinized vacuum tube that is immediately placed on ice. centrifuged and the plasma removed within 15 minutes of draw. .

93. It is crucial to keep blood samples cold after collection, because the ammonia conc of standing blood and plasma↑ spontaneously

94. this increase is bcoz of generation,release of ammonia from rbc deamination of amino acids,( glutamine .) capillary blood avoided, platelet aggregation, clotting → ↑ ammonia levels .

95. Measurements should be taken at the same time of day ( a diurnal variation ) ammonia levels in whole blood samples maintained at 4oC are stable for <1 hour. , plasma ammonia levels are stable at 4oC for 4 hours. DO NOT FREEZE Samples must arrive at laboratory within 3 hours after collection.

96. Do not use block ice or dry ice, as this will freeze the specimen. Hemolyzed specimens and Specimens received at ambient temperatures, will not be analyzed, as falsely increased ammonia concentrations may result.

97. The decrease in absorbance at 340 nm, due to the oxidation of NADPH, is proportional to the ammonia concentration. 0.2–15 mg/ml. Spectrophotometer

98. NADH is converted to NAD+ in the presence of NH3, ketoglutarate and glutamate dehydrogenase . The decrease in optical density at 340 nm or fluorescence intensity Quantitative Colorimetric/ Fluorimetric Determination of Ammonia

99. bunchman Flow Diagram to Evaluate Hyperammonemia Increased ammonia acidosis No acidosis Urine for organic acids Plasma amino acids Lactate/ pyruvate

100. HYPERCITRULLINURIA HYPERORNITHINEMA HYPERAMMONEMIA

101. ARGINASE DEFICIENCY

102. ↑ removal of nitrogen waste. convert nitrogen into products other than urea, which are then excreted; the load on urea cycle ↓ .

103. Protein intake : stopped. Calories : hypertonic 10% glucose. Hemodialysis : all comatose neonates with plasma ammonium levels greater than 10 times reference range. the total dialysis time is shorter with hemodialysis than with peritoneal dialysis. Treatment should be started if the plasma ammonium level is 3 times the reference level

104. The first sodium benzoate and arginine . Later, phenylacetate now replaced by phenylbutyrate (orally) . Sodium phenylbutyrate is a prodrug and is metabolized to phenylacetate

105. arteriovenous or venovenous hemofil tration as an alternative method IV sodiumbenzoate phenylacetate : ammonium level falls to 3-4 times the upper limit IV arginine

106. Na benzoate with arginine for (CPS), (OTC), (ASS), (ASL) deficiencies

107. Antiemetic control nausea and vomiting associated with IV administration of sodium benzoate and phenylacetate Ondansetron Granisetron Palonosetron Dolasetron

108. Arginine supplementation: is an essential AA in patients with urea cycle defects . In neonates and in OTC and CPSI deficiencies, citrulline can be given as a source of arginine as it gives one less nitrogen atom; in late-onset cases, arginine is acceptable because of increased nitrogen tolerance .

109. Citrulline levels are ↑in ASS and ASL deficiencies and citrulline should not be administered in patients with unknown enzyme deficiency. Provide enough calories to meet energy requirements

110. Carglumic acid ( Carbaglu ) N - carbamoyl -L-glutamate, Structural analogue of NAG enables activation of CPS I (first enzyme of urea cycle) ammonia i nto urea. More resistant to enzymatic degradation by hydrolysis compared with N - acetylglutamate .

111. Corticosteroids are not FOR ↑ ICP (induce negative nitrogen balance.) Mannitol is not effective in treating cerebral edema induced by hyperammonemia . Valproic acid should not be used to treat seizures as it decreases urea cycle function and ↑ammonia

112. Osmotic demyelination syndrome serious complication of standard therapy for hyperammonemia in patients with ornithine transcarbamylase deficiency. [23]

113. Diet Low protein intake: 0.7 g/kg/day of protein 0.7 g/kg/day of essential amino acid mixture. first 6 months, an infant may tolerate 1.5-2 g/kg/day of protein .

114. A gastrostomy tube is the most reliable way to administer medications and fluids during illness provide adequate nutritional support to prevent catabolism.

115. Surgical Care Liver transplantation correct the metabolic error. , and requirements for medication and dietary restriction were eliminated. Neurologic outcomes correlated closely with status prior to transplantation. Thus, liver transplantation is a good option for patients with urea cycle defects who have not suffered major brain injury.

116. Liver cell transplantation, as multiple intraportal infusions of cryopreserved hepatocytes

117. Further Outpatient Care monitoring growth and development of the child that would indicate the adequacy of treatment .periodic fasting levels of the Plasma ammonium Plasma glutamine (should be maintained at < 1000 µmol/L) Arginine Total protein

118. Complications Cerebral edema Cortical blindness

119. Prognosis the 5-year survival 22% neonatal-onset 41% late-onset group. neonatal-onset 90% severe neurologicdeficits , late-onset 28%