DM Lecture 23-Ammonia Metabolism Urea Cycle noted_

The non-toxic transport forms of ammonia from peripheral tissues are glutamine and alanine. Glutamine is formed by the incorporation of NH3 to glutamate, while alanine serves as a transport form of NH3 from muscle.

In cases of urea cycle defects, glutamine levels are elevated due to the impaired conversion of ammonia to urea, leading to increased ammonia levels in the blood.

Ammonia is considered neurotoxic, which creates a requirement for its transport in a non-toxic form. This necessitates the use of compounds like glutamine and alanine to safely transport ammonia from peripheral tissues to the liver for detoxification.

Glutamine plays a crucial role in ammonia transport by being formed in most tissues, particularly in the brain. It is synthesized from glutamate through the action of the enzyme glutamine synthetase, which combines glutamate with ATP and NH3, producing glutamine and releasing ADP + Pi as byproducts. In cases of urea cycle defects, glutamine levels are elevated due to impaired ammonia detoxification.

Alanine is synthesized in the muscle through a process called transamination, where pyruvate reacts with glutamate. The enzyme responsible for this reaction is alanine aminotransferase (ALT). Alanine plays a significant role in the ammonia transport cycle by moving back to the liver, where it can be converted to pyruvate, thus linking amino acid metabolism with glucose production.

The cycle involving glutamine and alanine in ammonia transport includes the following steps:

1. Glutamine Formation: In most tissues, glutamine is formed from glutamate by glutamine synthetase.
2. Liver Conversion: In the liver, glutamine is converted back to glutamate by glutaminase, releasing NH3.
3. Glutamate Metabolism: Glutamate is then converted to α-Ketoglutarate via glutamate dehydrogenase.
4. Alanine Synthesis: Pyruvate is converted to alanine in the muscle through transamination with glutamate, facilitated by ALT.
5. Cycle Completion: Alanine returns to the liver, where it can be converted back to pyruvate, completing the cycle. This cycle is significant as it helps in the detoxification of ammonia and links amino acid metabolism with glucose production.

α-KG acts as the amino acceptor in the transamination process, facilitating the transfer of the amino group from amino acids to form glutamate.

The transamination of alanine results in the formation of pyruvate, while the transamination of aspartate leads to the formation of oxaloacetate.

The aminotransferase reaction requires PLP (Pyridoxal phosphate), which is a derivative of Vitamin B6, and it is essential for the transfer of the amino group during the reaction.

Glutamate dehydrogenase catalyzes the conversion of glutamate to alpha-ketoglutarate, releasing free ammonia in the process. This free ammonia then enters the urea cycle for detoxification.

Glutaminase catalyzes the reaction that converts glutamine into free ammonia and glutamate. The ammonia produced can then be converted to NH4+, which is lost in urine, playing a crucial role in acid-base balance and H+ excretion.

The formation of ammonia, particularly through the action of glutaminase in the renal tubule, is significant for maintaining acid-base balance in the body. The ammonia can be converted to NH4+, which helps in the excretion of excess H+, thus regulating pH levels.

Bacterial ureases in the colon convert dietary proteins, bacterial proteins, and cellular proteins into ammonia (NH3) and ammonium (NH4+).

Cirrhosis increases the amount of ammonia entering systemic circulation due to impaired detoxification processes in the liver, leading to potential neurotoxicity.

In liver disease, intestinal ammonia formation becomes significant as the liver is unable to detoxify ammonia effectively, allowing it to enter systemic circulation and potentially cause neurotoxicity.

The two sources of nitrogen in the urea molecule are:

1. Ammonia - provides the first nitrogen atom.
2. Aspartate - provides the second nitrogen atom.

The urea cycle takes place in the liver and is partly mitochondrial (two reactions) and partly cytosolic (three reactions).

N-acetyl-glutamate acts as an allosteric activator and has an absolute requirement for the enzyme involved in the urea cycle.

The rate limiting step in the urea cycle is catalyzed by Carbamoyl Phosphate Synthetase I (CPSI), which requires N-acetylglutamate as an activator.

The main products of the urea cycle are Urea and Fumarate. Urea is excreted from the body, while fumarate enters the TCA cycle to form Oxaloacetate.

The first step of the urea cycle requires HCO3-, NH4+, and 2 ATP to produce Carbamoyl Phosphate.

N-acetylglutamate acts as an allosteric activator in the urea cycle, specifically in the conversion of CO2 and NH3 to carbamoyl phosphate, which is the rate-limiting step of the cycle.

The main products of the urea cycle are urea, which is excreted by the kidneys, and ornithine, which is recycled back into the cycle. Additionally, fumarate is produced as a byproduct during the conversion of argininosuccinate to arginine.

The urea cycle is crucial in amino acid catabolism as it converts excess nitrogen into urea, allowing for its excretion by the kidneys. This process helps to prevent the accumulation of toxic ammonia in the body.

CPS-I (Carbamoyl phosphate synthetase-I) is a mitochondrial enzyme that incorporates free ammonia into the urea cycle. It is activated by N-acetyl glutamate (NAG).

Arginase (ARG) is the final enzyme in the urea cycle that catalyzes the formation of urea from arginine, facilitating the excretion of nitrogenous waste.

The enzyme that uses aspartate in the urea cycle is Argininosuccinate synthetase (ASS).

The key clinical symptoms include:

• Vomiting
• Grunting respiration (hyperventilation)
• Rapid lethargy and unresponsiveness

The laboratory findings include:

• Elevated serum ammonia and glutamine levels
• Respiratory alkalosis (indicated by increased pH, decreased PCO2, and normal or low HCO3)
• Plasma urea at the lower end of normal

Ammonia directly stimulates the respiratory center, leading to hyperventilation, which results in respiratory alkalosis.

Hyperammonemia presents with:

• Flapping tremor (asterixis)
• Slurring of speech
• Somnolence
• Vomiting
• Cerebral edema
• Blurring of vision

Treatment options for hyperammonemia include:

1. Limit protein in diet
2. Lactulose to acidify the GI tract and trap NH4+ for excretion.
3. Antibiotics (e.g., rifaximin) to reduce ammoniagenic bacteria.
4. Benzoate, phenylacetate, or phenylbutyrate which react with glycine or glutamine, forming products that are excreted renally.

Increased ammonia (NH3) causes CNS toxicity by:

• Inhibiting the TCA cycle (decreasing α-ketoglutarate)
• Decreasing glutamate levels
• Increasing GABAergic tone (increasing GABA)
• Increasing glutamine, leading to cerebral edema due to osmotic shifts.

Enzyme deficiency leads to the accumulation of the substrate of the enzyme, resulting in increased blood ammonia (hyperammonemia) and glutamine levels.

Symptoms include:

1. Lethargy
2. Irritability
3. Feeding difficulties
4. Respiratory alkalosis (due to ammonia stimulating the respiratory center, leading to hyperventilation)
5. Neurological manifestations such as seizures, intellectual disability, and developmental delay due to elevated ammonia levels.

CPS-1 (Carbamoyl phosphate synthetase 1) or OTC (Ornithine transcarbamylase) deficiencies are the most severe among urea cycle disorders, leading to significant decreases in urea formation and severe hyperammonemia.

CPSI (Carbamoyl Phosphate Synthetase I) is the rate-limiting enzyme in the urea cycle, catalyzing the conversion of bicarbonate and ammonia into carbamoyl phosphate. Dysfunction of CPSI leads to Hyperammonemia I, characterized by elevated ammonia levels in the blood.

Hyperammonemia II is associated with a deficiency in the enzyme Ornithine Transcarbamylase (OTC). Clinical manifestations include elevated ammonia levels and the presence of orotic acid in urine due to disrupted urea cycle function.

Citrullinemia is characterized by elevated levels of citrulline in the blood and urine, resulting from a deficiency in the enzyme Argininosuccinate Synthetase (ASS). Clinically, this condition can lead to severe hyperammonemia, which can cause neurological damage if not managed promptly.

N-acetylglutamate acts as an allosteric activator of carbamoyl phosphate synthetase 1, which is crucial for initiating the urea cycle by facilitating the conversion of ammonia and bicarbonate into carbamoyl phosphate.

The main products of the urea cycle include urea, which is excreted by the kidneys, and fumarate, which can enter the citric acid cycle. The cycle also involves the conversion of ammonia and aspartate into urea.

The rate-limiting step in the urea cycle is catalyzed by carbamoyl phosphate synthetase 1. This step is significant because it regulates the overall rate of the urea cycle, impacting the detoxification of ammonia and the synthesis of urea.

1. Dialysis: Used in acute emergencies.
2. Diet: Low protein and high carbohydrate to minimize nitrogen intake.
3. Stress Prevention: Avoid stress that induces a catabolic state.
4. Medications: Administration of benzoic acid and/or phenylbutyrate/phenylacetate as alternate routes of nitrogen excretion.

The long-term management option for hyperammonemia in Urea Cycle Disorders is liver transplantation.

A diet that is low in protein and high in carbohydrates helps to minimize nitrogen intake, which is crucial in managing hyperammonemia and preventing further accumulation of ammonia in the body.

The combination of Phenylbutyrate and Glutamine forms Phenylacetylglutamine, which is significant as it facilitates nitrogen loss through excretion in urine, helping to manage hyperammonemia.

Benzoic acid combines with Glycine to form Hippuric acid, which is then excreted in urine. This process aids in the loss of nitrogen, thus helping to reduce ammonia levels in the body.

The metabolic pathways involve:

1. Phenylbutyrate → Phenylacetate → Phenylacetylglutamine (excreted in urine)
2. Benzoic acid → Hippuric acid (excreted in urine)

Both pathways result in nitrogen loss, which is crucial for managing hyperammonemia.

Arginine administration allows the urea cycle to progress by increasing substrate concentration. It is converted to ornithine by the arginase enzyme, which facilitates the cycle's function.

Arginine activates NAG synthesis, which is an allosteric activator of CPS-1, enhancing the urea cycle's efficiency.

N-acetyl glutamate (NAG) is crucial as it activates carbamoyl phosphate synthetase I (CPS-1), which is the first step in the urea cycle, thus facilitating the conversion of ammonia to urea.

• Very high blood ammonia and glutamine levels; most severe form of hyperammonemia.
• Low levels of ALL urea cycle intermediates.
• May respond to Arginine supplements which stimulate the formation of N-acetylglutamate (NAG), an allosteric activator of CPS-I.

Arginine supplementation may help by stimulating the formation of N-acetylglutamate (NAG), which can potentially activate the deficient CPS-I enzyme, aiding in the conversion of NH3 and CO2 into Carbamoyl-phosphate.

N-acetylglutamate (NAG) is an allosteric activator of CPS-I, which is crucial for the conversion of NH3 and CO2 into Carbamoyl-phosphate, a key step in the urea cycle.

The most common urea cycle disorder is Hyperammonemia Type II, which is characterized by elevated blood glutamine levels.

OTC deficiency is usually seen in males and is more severe in males due to its X-linked inheritance pattern.

OTC normally combines ornithine and carbamoyl phosphate to form citrulline, which is then transported to the cytoplasm to combine with aspartate to form arginosuccinate.

The key laboratory findings in OTC deficiency include:

• Elevated serum ammonia
• Elevated serum and urine glutamine
• Elevated serum and urine orotic acid

In Ornithine transcarbamoylase deficiency, elevated carbamoyl phosphate drives pyrimidine biosynthesis, leading to increased levels of orotic acid in both serum and urine.

Orotic aciduria is a significant clinical marker in OTC deficiency, indicating disrupted pyrimidine metabolism due to elevated carbamoyl phosphate levels, which can help in diagnosing the condition and monitoring treatment efficacy.

The key diagnostic indicators include hyperammonemia, high glutamine levels, and very high levels of serum and urinary citrulline.

Elevated citrulline levels are significant as they indicate a disruption in the urea cycle, specifically related to argininosuccinate synthetase deficiency, leading to the accumulation of citrulline in serum and urine.

Hyperammonemia can lead to severe neurological complications, as elevated ammonia levels are neurotoxic and can result in cognitive impairment, seizures, and potentially coma if not managed promptly.

Key laboratory findings include:

• Elevated ammonia
• Elevated glutamine
• Elevated argininosuccinate in plasma and urine
• Elevated citrulline due to the metabolic block occurring after its formation.

In argininosuccinate lyase deficiency, hyperammonemia is characterized by:

• Elevated ammonia levels in the blood
• Elevated glutamine levels as a response to ammonia toxicity
• Elevated argininosuccinate in both plasma and urine, indicating a specific metabolic block in the urea cycle.

Elevated citrulline in argininosuccinate lyase deficiency is significant because:

• It indicates that the urea cycle is functioning up to the point of citrulline production.
• The elevation occurs because citrulline is produced before the metabolic block at the argininosuccinate lyase step, leading to its accumulation in the plasma.

• Elevated serum ammonia and elevated arginine levels
• Blood NH3 levels are not as high as in other Urea Cycle Disorders (UCD)
• Treatment includes a diet of essential amino acids excluding arginine
• Significant number of cases are adult onset, often associated with neurological problems

The treatment for Arginase deficiency includes:

1. Dietary management: A diet of essential amino acids that excludes arginine.
2. Other treatments: Additional treatments may be applied as previously listed in the context of Urea Cycle Disorders.

Arginase deficiency is characterized by:

• Elevated serum ammonia and elevated arginine levels, but blood NH3 levels are not as high as in other UCDs.
• A significant number of cases present in adulthood, often leading to neurological problems.

Lab findings for Hyperammonemia Type I include:

• Hyperammonemia (most severe)
• Elevated glutamine
• Low levels of all urea cycle intermediates

Management strategies include:

• Low protein diet to reduce ammonia production
• Phenylacetate and benzoic acid to help remove excess nitrogen
• Arginine supplementation to provide an alternative pathway for nitrogen disposal.

Hyperammonemia Type II is characterized by:

• Hyperammonemia
• Elevated glutamine
• Increased orotic acid excretion in urine, which indicates a blockage in the urea cycle due to the deficiency of ornithine transcarbamylase (OTC).

Management includes a low protein diet, phenylacetate, benzoic acid, and arginine to manage ammonia levels.

Management strategies for Citrullinemia include:

• Low protein diet to limit ammonia production
• Phenylacetate and benzoic acid to facilitate nitrogen removal
• Arginine supplementation, which helps to bypass the metabolic block caused by the deficiency of argininosuccinate synthetase.

Lab findings include:

• Hyperammonemia
• Elevated glutamine
• Increased citrulline levels.

Amino acids are converted into ammonia through transamination and oxidative deamination.

After urea is formed in the liver, it is transported to the kidney and excreted in urine. Additionally, some urea is degraded in the gut, allowing ammonia to reenter circulation, which the liver must then detoxify.

Renal failure leads to elevated Blood Urea Nitrogen (BUN) levels and can result in elevated blood ammonia, which may indicate severe liver disease or inherited urea cycle defects.

Acquired hyperammonemia in liver disease is primarily caused by viral or drug-induced hepatitis and alcoholic cirrhosis. In cirrhosis, there is porto-systemic shunting of blood, allowing portal blood to enter systemic circulation without passing through the liver, leading to increased ammonia levels in the bloodstream.

In cirrhosis, porto-systemic shunting allows ammonia produced in the intestine to enter the systemic circulation directly, bypassing the liver where it would normally be detoxified. This results in elevated ammonia levels in the blood, leading to neurotoxicity.

Bacterial ureases in the colon convert urea into ammonia, contributing to the overall ammonia formation in the gut. This process becomes particularly significant in liver disease, as the ammonia produced is not effectively detoxified by the liver, leading to increased levels in systemic circulation.

The patient exhibits confusion, inability to concentrate, apathetic behavior, disorientation, drowsiness, slurred speech, and flapping tremor (asterixis).

The patient has alcohol-induced liver cirrhosis, which leads to the liver's inability to detoxify ammonia (NH3) in circulation.

The patient has elevated blood ammonia levels and elevated plasma glutamine levels, indicating impaired ammonia detoxification due to liver dysfunction.

A low-protein/high carbohydrate diet is recommended to reduce nitrogen intake, leading to less nitrogen needing disposal.

Lactulose is a disaccharide that is resistant to digestion in the small intestine. It is digested by normal flora, producing lactic acid, which neutralizes NH4+, resulting in more nitrogen being excreted in the feces.

Neomycin, or other antibiotic treatments, help reduce bacterial urease in the gut, which can contribute to ammonia production.

Hyperammonemia leads to an imbalance of neurotransmitters, disrupting the balance between excitatory and inhibitory neurotransmission. This can result in neurological symptoms and cognitive dysfunction.

Hyperammonemia impairs brain ATP metabolism, which is crucial for energy production in brain cells. This impairment can contribute to neuronal dysfunction and cell death.

Brain CT scans in cases of hyperammonemia typically show widespread loss of cortical sulci and gray-white matter differentiation, indicative of cerebral edema.

Hyperammonemia alters the balance between excitatory neurotransmitters (like Glutamate) and inhibitory neurotransmitters (like GABA), leading to potential neurotoxicity.

Glutamate Decarboxylase converts Glutamate (an excitatory neurotransmitter) into GABA (an inhibitory neurotransmitter), thus influencing the excitatory-inhibitory balance in the brain.

The two types of neurotransmitters involved are excitatory neurotransmitters (e.g., Glutamate) and inhibitory neurotransmitters (e.g., GABA).

Elevated blood ammonia leads to the conversion of α-ketoglutarate to glutamate via the enzyme glutamate dehydrogenase, resulting in depletion of TCA cycle intermediates and decreased TCA cycle activity. This ultimately reduces ATP synthesis, which is critical for brain function.

Increased glutamine levels, resulting from the conversion of glutamate to glutamine, can lead to reduced Na+/K+ ATPase activity, causing osmotic pressure changes and edema, which alters neurotransmission.

Decreased TCA cycle activity due to elevated ammonia levels results in reduced ATP synthesis, which is essential for neuronal function, and can lead to neurological impairments due to insufficient energy supply.