Total Serum Bile Acids in Hepatic Disease in Small Animals
Total serum bile acid (TSBA) concentrations can sensitively detect cholestatic disorders and conditions associated with portosystemic shunting, including microvascular dysplasia in small dog breeds. TSBA concentration should be measured before and 2 hours after meal ingestion; fasting is not required. Insufficient hepatic mass or portal circulation deviated to the systemic circulation via extrahepatic portosystemic shunts (congenital or acquired) or microscopic shunting within the liver (congenital microvascular dysplasia) causes high TSBA concentrations.
TSBA concentrations are usually lower before a meal than 2 hours after a meal. However, ~15%–20% of dogs and 5%–10% of cats have higher TSBA concentrations before meal ingestion than after, likely reflecting physiologic variables influencing the enterohepatic circulation of bile acids (ie, the rate of gallbladder contraction, gastric emptying, and intestinal transit of bile acids to the ileum, where they are actively resorbed). TSBA concentrations in dogs > 25 mcmol/L or in cats > 20 mcmol/L are abnormal (either before or after a meal; fasting ranges should not be applied because of variations influencing the TSBA enterohepatic circulation). Collecting a single sample for TSBA measurement (random fasting or a single postprandial sample) can miss detection of abnormal values.
Because TSBA concentrations are a more sensitive indicator of cholestasis than total bilirubin, measuring TSBA concentration is redundant in animals with nonhemolytic jaundice. Use of TSBAs as a liver function test can indicate need for a liver biopsy.
To best use TSBA concentrations as a means of assessing liver function, they should be routinely measured in all young (6 months), small, terrier-type breeds to detect microvascular dysplasia (MVD) during normal health screenings. Finding increased TSBA concentrations in apparently healthy young terrier-like breeds and other small-breed dogs allows detection of dogs in which TSBA concentration will be misleading if discovered in later life during evaluation of illness. Breeds to assess include but are not restricted to the following:
Blood samples that are markedly hemolyzed are unsuitable for analysis. Blood samples that are hyperlipidemic should be ultracentrifuged prior to assay to clarify serum/plasma.
Treatment with ursodeoxycholic acid (UDCA) can increase measured TSBA concentrations and should be suspended at least 1 day prior to testing (UDCA is measured by the bile acid assay).
Ammonia in Hepatic Disease in Small Animals
Measurement of blood ammonia can detect hepatic disorders associated with hepatic encephalopathy (HE); in some cases with acute increases in transaminase activity, documentation of hyperammonemia heralds onset of fulminant hepatic failure.
Ammonia is derived predominantly from protein degradation, with most generated in the intestines from consumed food and enteric bacterial ureases that catabolize urea into ammonia and carbon dioxide. In health, portal transport of ammonia from the colon to the liver results in its direct detoxification (~85%) to urea.
Ammonia intolerance (impaired clearance) occurs in disorders associated with portosystemic shunting and in acute fulminant hepatic failure. Ammonia is not influenced by cholestasis or liver disorders that do not impair the normal portal-to-liver perfusion (ie, portosystemic shunting) or that do not cause extensive loss of functional hepatic parenchyma.
Although ammonia is regarded as a pivotal cause of HE, animals with overt HE may have normal blood ammonia concentrations because there are numerous substances and complicated pathological mechanisms driving HE. A single normal ammonia value cannot discount HE in an animal with suspected chronic liver disease. Furthermore, serial ammonia measurements may not correlate with an evolving clinical scenario of suspected HE. Thus, ammonia measurements cannot reliably diagnose HE unless associated with other features of hepatic insufficiency or portosystemic shunting or in the presence of concurrent ammonium biurate crystalluria.
Measurement of blood ammonia is complicated. Spurious hyperammonemia can reflect a variety of circumstances:
slow blood collection
tight tourniquet technique
conditions promoting ammonia liberation from muscle (seizures, crush injuries as observed in dogs with automobile injury)
sample contamination (human sweat, cigarette smoke, aerosolized urea from open urine vials)
spontaneous generation of ammonia in samples not immediately cooled after collection or not promptly analyzed
Being highly volatile, samples for ammonia quantification cannot be mailed for analyses. Blood samples should be collected into precooled tubes and transported on melting ice to the analysis site within 20 minutes. Enzymatic-based methodologies are difficult to standardize. A dipstick test using an automated analyzer that has been used in Europe is not validated for sale in the US.
If a random or 30-minute postprandial blood ammonia concentration is within the reference range but hepatic insufficiency and portosystemic shunting is suspected, an ammonia tolerance test can be considered. Ammonium chloride is given at 100 mg/kg as a 5% solution orally (may provoke vomiting) or at 2 mL/kg of a 5% solution administered rectally (instilled 30 cm deep) after a cleansing enema. Blood ammonia is measured at baseline and then 20, 30, 40, and 60 minutes later. Unfortunately, ammonia tolerance testing may provoke iatrogenic HE in some animals (with portosystemic shunting).
Finding hyperammonemia always warrants consideration of occult nonhepatic causes, especially if clinical details implicating primary hepatic disease are less than convincing. The most common nonhepatic disorder provoking nonhepatic hyperammonemia is bacterial infection of the urinary tract with urease-producing organisms; usually this involves uroabdomen or an obstructive uropathy rather than just a simple infection. Extreme skeletal muscle activity or massive muscle injury also can provoke hyperammonemia; in this scenario, high transaminase activity accompanies extreme increase in creatine kinase.
While muscle normally functions as a net consumer of ammonia (transforming glutamate to glutamine via glutamine synthetase [GS]), muscle also may convert to local ammonia production when GS is overwhelmed by glutamate availability. Increased muscle ammonia production becomes proportionate to exercise intensity, muscle utilization of branched-chain amino acids for energy (AA catabolism), and deamination of adenosine monophosphate (AMP) for energy. Treatment with valproic acid, an antiepileptic drug, can inhibit carbamoyl phosphate synthetase 1 (CPS1), limiting carbamyl phosphate for its obligatory combination with ammonia as it enters the urea cycle.
Polypharmacy regimens of valproic acid with P450 inducers (ie, phenytoin, phenobarbital, carbamazepine) or topiramate heighten risk for hyperammonemia. Topiramate, an anticonvulsant also used in animals, is thought to aggravate hyperammonemia by blocking carbonic anhydrase. This effect favors development of a metabolic acidosis and shifts ammonium ion equilibrium towards ammonia. This drug is also suspected of inhibiting cerebral GS, thwarting ammonia detoxification in the brain. L-asparaginase, a chemotherapeutic drug (also used in animals) hydrolyzes asparagine (highly used by certain neoplastic cells) into aspartic acid and ammonia, thereby contributing to ammoniagenesis.
Any cause of GI bleeding has potential to provoke hyperammonemia, especially in patients with hepatic insufficiency or portosystemic shunting (congenital or acquired). Simulation of upper GI bleeding in cirrhotic humans confirms inducible hyperammonemia with kidneys as the source of exaggerated ammoniagenesis. The branched-chain amino acid alanine is implicated as a major substrate. Mechanistically, degradation of blood by enteric flora and mucosal oxidation directly increases ammonia generation. Because alanine is the predominant amino acid in hemoglobin, degradation of blood increases its local release.
Increased enteric absorption of ammonia, glutamine, and alanine collectively promotes renal ammoniagenesis. In the context of liver insufficiency, diminished hepatic glycogen reserves instigate glucagon release, which drives peripheral gluconeogenesis (including within the kidney). A cascade of biochemical interactions involving alanine, alpha-ketoglutarate, pyruvate, glutamine, and ammonia is operational in this process.
Alanine deamination increases glutamine, which promotes ammonia generation; pyruvate escalates gluconeogenesis. The collective impact leads to enhanced renal tubular ammonia generation with some eliminated in urine and the remainder absorbed into the systemic circulation. The impact of this complex process has been demonstrated in a cirrhotic human treated with IV glucose infusions (12-hour treatment with 20% glucose solution [rate 28 mcmol/kg/min]) before simulating enteric hemorrhage. Substantial attenuation of renal ammoniagenesis and systemic hyperammonemia was demonstrated.