Urea Cycle & OTC Deficiency Dolores, Jimenez, Kinomes, Legitimas, Manapat

Where does excess Nitrogen go? Amino portion of amino acids undergoes transamination reactions in breakdown as well as biosynthesis Excreted in 1 of 3 forms: Ammonia, Urea , Uric acid Fishes Terrestrial animals Birds

Amino acid catabolism If there are more than enough amino acids for protein synthesis, amino acids can be used as metabolic fuel (cell lining in small intestine uses a.a. as a main source of energy) Glucogenic AA-yields alpha-ketoglutarate, oxaloacetate, pyruvate Ketogenic AA-yields acetoacetyl-CoA, acetyl CoA

Ammonia (NH3) is a waste product from amino acid catabolism But excreting ammonia is impractical in humans: -High concentration can cause alkalosis or the increase of alkalinity in the blood -Ammonia is highly toxic! (can enter the brain, combine with alpha KG-->glutamate) Story of Urea Cycle: Toxic Ammonia must become a nontoxic substance that can be excreted

enzyme: Glutamate dehydrogenase (mitochondria) Glutamate is the main supplier of nitrogen to the urea cycle enzyme: Glutamate dehydrogenase (mitochondria)

Urea Cycle

Steps

Step 1 Rate defining step Investment of 2 ATP Reactants: NH4+ and CO2 Products: Carbamoyl Phosphate Catalyzed by: Carbamoyl Phosphate Synthetase I (CPS1) Located: Liver Cell Mitochondria

Step 2 Reactants: Carbamoyl phosphate and ornithine Products: citrulline catalyzed by: Ornithine transcarbamoylase (OTC)

Step 3 Citrulline leaves the mitochondria via citrulline transporter Reactants: ATP, Citrulline, Aspartate Products: Argininosuccinate Catalyzed by: Argininosuccinate synthetase (ASS)

Step 4 Argininosuccinate gets broken down Reactants: Argininosuccinate Products: arginine and fumarate* Catalyzed by: argininosuccinate lyase (ASL)

Step 5 Reactants: Arginine and water Products: Urea and ornithine Catalyzed by Arginase 1 (ARG1)

Regulation and Control N-acetylglutamate Arginine

Connection with the TCA Cycle

Ornithine Transcarbamylase (OTC) Deficiency PIC: OTC enzyme. it’s a trimer. seen in the last reaction of the proximal phase of the urea cycle http://www.ncbi.nlm.nih.gov/Structure/mmdb/mmdbimage.fcgi?uid=13331&type=1&unm=1

INTRODUCTION Most common urea cycle disorder X-linked disorder Deletion in OTC gene (Xp21.1) Lack of the enzyme ornithine transcarbamylase Ornithine transcarbamylase (OTC) deficiency is a rare X-linked defect that is the most common of the urea cycle disorders. It is inherited in an X-linked recessive manner, meaning males are more commonly affected than females The gene coding for ornithine transcarbamylase is located on the short arm of the X chromosome at position p21.1 There are no common mutations that cause disease, however 10 - 15% of disease causing mutations are deletions Ornithine transcarbamylase, found primarily in liver mitochondria, is an early enzyme in the urea cycle. When ornithine transcarbamylase is missing, carbamyl phosphate cannot combine with ornithine. As carbamyl phosphate concentration rises in the liver, ammonia is released. When ammonia reaches the CNS via the blood, nervous system degradation leads to the symptoms of the defect

PREVALENCE Approximately 1 in 80,000 live births The estimated incidence of early-onset OTC deficiency is 1 in 80,000 live births. When male late-onset OTC deficiency and variable female expression are included, estimates run as high as 1 in 20,000 live births

SYMPTOMS Hyperammonemia The primary symptom of OTC deficiency is hyperammonemia. Metabolic condition characterized by elevated levels of ammonia in the blood

Early-Onset OTC Deficiency Lethargy Anorexia Failure to thrive Hypotonia Vomiting Disorientation Seizures Somnolence In early-onset OTC deficiency, as the excess ammonia reaches the central nervous system, lethargy (abnormal drowsiness/state of being lazy,sluggish), anorexia (loss of appetite), and a general failure to thrive (subsequent growth delay and cognitive deficiencies) are often the first readily apparent symptoms. These are followed by hypotonia (decrease muscle tone), vomiting, disorientation, seizures, somnolence (sleeping more than often), coma, and finally death. In those that survive, mental retardation is common. Many of these same symptoms, sometimes less severe, are seen in late-onset OTC deficiency

What exactly happens? This is the step where OTC is so crucial. PIC: https://amit1b.files.wordpress.com/2012/09/9.png

What exactly happens? LACK OF OTC = Ornithine and carbamoyl phosphate do not get turned into citrulline But doesn’t it just lead to carbamoyl phosphate and ornithine accumulation? PIC: https://amit1b.files.wordpress.com/2012/09/9.png

What exactly happens? How does this lead to ammonia build-up, lethargy, coma etc? PIC: http://intranet.tdmu.edu.ua/data/kafedra/internal/chemistry/classes_stud/en/nurse/BSN/ptn/2/16.%20Investigation%20of%20detoxification%20processes%20and%20biosynthesis%20of%20urea.files/image012.gif

Hyperammonemia Substrate build-up due to lack of OTC Build-up goes all the way back to NH4⁺ Hyperammonemia is common in most urea cycle disorders. Basically, the deficiency in enzymes leads to increased respective substrates, continuing all the way back to NH4+. The rate of the rxn is decreased + build-up causes problems When the capacity of the liver to metabolize ammonia is overcome, either because ammonia production exceeds the metabolic capacity of the liver or because the liver is unable to metabolize ammonia, elimination is dependent on the kidney, muscle, and brain. In the setting of hyperammonemia, the kidney decreases ammonia production and increases urinary excretion of ammonia.4,10 The muscle4–6,11–12 and brain metabolize excess ammonia to glutamine.1,11–13 The process of metabolizing ammonia to glutamine is physiologically costly, particularly in the brain where the symbiotic relationship between neurons and astrocytes is disrupted by excess glutamine production. PIC: http://upload.wikimedia.org/wikipedia/commons/7/76/Symptoms_of_hyperammonemia.svg

Lethargy, Confusion, Vomiting Elevated NH4⁺ leads to brain complications affecting movement, behavior, appetite short-circuits potassium transport in glial cells leading to swelling drives glutamine synthase rxn (glutamate → glutamine) hence depleting glu (precursor of GABA) high ammonia + low glu reverses glutamate dehydrogenase rxn (glutamate → a-ketoglutarate). low a-ketoglutarate inhibits Krebs cycle thus less energy early symptoms. Ammonia is especially toxic to CNS. How, no one is sure. Some theories above. addtional: Ammonia crosses the blood–brain barrier through passive diffusion and cation channels. It affects multiple neurotransmitters, specifically the excitatory glutaminergic NMDA receptors and modulation of GABA receptors. Ultimately, neuronal apoptosis occurs.10Hyperammonemia alone appears sufficient to cause brain edema in vivo and astrocytic swelling in vitro, increasing intracranial pressure via an increase in cerebral blood volume.11

EARLY ONSET LATE ONSET Rapid decline (~24 hours) in health if OTC deficiency is not treated in time Intellectual disabilities, Coma, Brain death, Death Hyperammonemia induced by stress or large protein intake Developmental delays, Protein aversion Later symptoms include these

Without efficient urea cycle, ammonia disposal is left to muscle, kidney, brain. Again, especially taxing on brain. PIC: http://journal.publications.chestnet.org/data/Journals/CHEST/22062/zcb0100709840001.jpeg

Diagnosis & Treatment PIC: http://www.devilbisshc.com/images/standard-page-images/1051/cf-diagnosis__large.jpg

Diagnosis Demonstration of hyperammonemia Very low blood urea nitrogen (BUN) level Normal liver and kidney function in most cases, unless hypoxia or shock supervenes Elevated ornithine, glutamine, and alanine levels; low citrulline level Elevated urinary orotic acid level

TREATMENT & MEDICATION Hemodialysis Dietary Management - regulated protein, carbohydrates, and fat intake - supplements (Cyclinex, EAA) - calorie modules (Prophree, Polycose) - antioxidants hemodialysis - fastest method for lowering ammonia level too much dietary protein - excessive ammonia production too restricted proteins/insufficient calories - body will catabolize lean muscle mass (catabolism creates excessive ammonia) supplementation wtih special amino acid formulas such as Cyclinex and EAA. can provide ~50% daily dietary protein allowance UCD people often lack appetite prescription of calorie modules such as Prophpree, Polycose to be used in combination with AA formulas antioxidants - suggested to be helpful in minimizing free radical damage to cells in tissue and brain

TREATMENT & MEDICATION Drug Treatment Buphenyl Sodium benzoate L-citrulline Antacids Buphenyl aka sodium phenylbutyrate primary medication being used to treat urea cycle disorder sodium benzoate is almost same

TREATMENT & MEDICATION Last Resort Liver Transplant

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