Tests of Hepatic Function

Many biochemical tests are available to evaluate the liver's anabolic and/or catabolic function and hepatic circulation. These include measurement of serum bile acid, plasma ammonia and bile pigment (bilirubin) concentrations and the ability to excrete organic dyes. Other tests of hepatic function include measurement of serum albumin, glucose and urea nitrogen, and clotting factor analysis. Hepatic function can be markedly abnormal despite maintenance of the hepatocellular membrane (and therefore normal serum activities of hepatic enzymes). Examples include portosystemic shunts, terminal cirrhosis and metastatic hepatic neoplasia. Likewise, the liver can continue normal anabolic or catabolic function despite severe hepatocyte leakage of intracellular enzymes because of its marked reserve capacity. This can occur, for example, in certain cases of hepatocellular necrosis or primary hepatic neoplasia.

Bile Acids

Bile acids are synthesised in the liver as a result of cholesterol metabolism and are secreted into bile. Following feeding, bile acids enter the intestine and undergo an efficient enterohepatic circulation following active absorption from the ileum. Once absorbed, they are removed from the portal circulation by the liver and re-excreted into bile. Only small amounts of bile acids are lost in the faeces. Several studies have shown serum bile acid measurements to be a sensitive and specific indicator of hepatic function in the dog and cat.

Several artefacts can affect bile acid measurement. Moderate to marked lipaemia artefactually increases the serum bile acid measurement determined by the enzymatic method, but artefactually decreases the measurement when determined by radioimmunoassay (RIA). Moderate to marked haemolysis artefactually decreases the serum bile acid value determined by the enzymatic method but probably does not affect the measurement determined by RIA.

In dogs, fasting serum bile acid concentrations are significantly increased with congenital portosystemic shunts, glucocorticoid-induced hepatopathy, hepatic neoplasia, hepatitis, cholestasis, hepatic necrosis and cirrhosis. Of these diseases, glucocorticoid-induced hepatopathy results in the lowest concentration of serum bile acids. Therefore, marked elevations (i.e., >75-100 µmol/l) are not likely to be caused by glucocorticoid-induced hepatopathy. Although serum bile acid concentrations are a sensitive indicator of hepatic function, they do not distinguish the cause of the disease process. The determination of 2-hour postprandial concentrations further increases the sensitivity of this test for most diseases. If an animal is not eating, force-feeding a small amount of food is usually sufficient. Occasionally the level in the preprandial sample is higher than that in the postprandial sample. This does not have any diagnostic significance, and the higher of the two levels is always used as the diagnostic measurement. For our laboratory, when either sample exceeds 30 µmol/l in the dog and 20 µmol/l in the cat, further diagnostic efforts, such as hepatic biopsy, are warranted.

Serum and Urine Bilirubin

Hepatobiliary excretion of bilirubin requires adequate uptake, conjugation and secretion by the hepatocyte. There must be considerable hepato-cellular disease or increased bilirubin load (haemolysis) to result in hyperbilirubinaemia. Because the dog has a very low renal threshold for bilirubin excretion, the finding of +1 to +3 bilirubinuria in a concentrated sample is normal. The concentration of bilirubin in the urine increases before that in the serum; serum bilirubin concentration is an insensitive indicator of hepatocellular disease, and serum concentrations are not increased until there is marked decrease in hepatic function. Therefore, slight elevations in serum bilirubin are significant, suggesting hepatobiliary disease. The exception to this is with artefactual increases in serum bilirubin concentration, as would occur with lipaemia or haemolysis. If there is not significant bilirubinuria associated with a serum bilirubin concentration greater than normal, artefact should be considered. The cat has a high renal threshold for bilirubin excretion, and any amount of bilirubin in the urine is abnormal. When serum bilirubin is in the normal range, other tests of hepatic function are needed for detection, such as serum bile acid measurements. However, when the serum bilirubin concentration is elevated, there is no need to run additional function tests if haemolysis can be excluded. In this setting bilirubin represents an accurate and specific indicator of hepatic function.

Increased total serum bilirubin concentration can result from prehepatic (haemolysis), intrahepatic (primarily hepatocellular disease) or posthepatic (biliary obstruction) causes. The relative amounts of conjugated or unconjugated bilirubin are variable in all three general categories of hyperbilirubinaemia because secondary events can change the relative concentrations of the two forms. Therefore, their measurement does not aid the clinician in localising the nature of the lesion. Other methods of localising the cause of hyperbilirubinaemia include measuring the haematocrit (to rule out haemolysis) and using ultrasonography, laparoscopy and laparotomy (to distinguish intrahepatic from posthepatic causes). Notably patients with portosystemic shunts and steroid hepatopathy are rarely hyperbilirubinaemic.

Serum Hepatic Enzyme Activities

Serum hepatic enzyme activities do not reflect hepatic function. Enzyme activities reflect either the integrity of the hepatocyte membrane or the patency of the biliary system. Severe hepatic dysfunction can occur in the face of normal enzyme activities, whereas hepatic function may be near normal despite marked increase in serum enzyme activities. Enzymes that leak into plasma following increased hepatocellular membrane permeability include alanine aminotransferase (ALT) and aspartate aminotransferase (AST). Serum enzyme activities that increase with biliary obstruction include alkaline phosphatase (ALP) and gamma-glutamyl transpeptidase (GGT).

Serum Alanine Aminotransferase

ALT is the most liver-specific enzyme in the dog and cat. It is used to detect hepatocyte membrane damage and necrosis. Serum activity increases when there is increased permeability of the hepatocyte membrane, resulting in leakage from the hepatocyte. The extent to which enzyme leakage occurs depends on both the severity and the number of cells damaged (i.e., how diffuse the lesion is) but does not indicate the reversibility of the injury or the functional status of the liver. The activity of serum ALT is highest in cases of chronic active hepatitis, primary hepatic neoplasia and hepatic necrosis. There is often normal or only mildly increased ALT activity in cases of portosystemic shunts, cirrhosis and metastatic neoplasia. In rare cases, increased serum ALT activity can occur with severe muscle disease.

Serum Aspartate Aminotransferase

AST, like ALT, has high activity in the liver. However, there is also high AST activity in skeletal muscle and red blood cells, and serum AST activity is therefore less specific than ALT activity. Increased activity of serum AST occurs with both hepatocyte and muscle damage, as well as with haemolysis (in vitro or in vivo). In hepatic disease, serum AST activities usually parallel those of ALT. Usually the magnitude of increase in serum ALT activity exceeds that of AST activity. When the activity of serum AST exceeds that of ALT, isoenzymes other than hepatic should be considered as the source (i.e., muscle and red blood cells (RBC)).

Serum Alkaline Phosphatase

The serum activity of ALP usually increases with biliary stasis, steroid hepatopathy and bone lesions. The activity of ALP increases in hepatic disease primarily when there is biliary obstruction (intrahepatic or extrahepatic). ALP activity is also often elevated in primary hepatocellular disease of many causes as a result of swelling of hepatocytes, resulting in intrahepatic cholestasis. Diseases that are periportal in location tend to cause more marked increases in ALP activity than centrilobular disorders because they tend to affect bile flow though canaliculi more.

In addition to the isoenzyme induced by biliary obstruction, there is a steroid-induced isoenzyme of ALP. Although this isoenzyme also is produced in the liver, it is a separate entity from that induced by biliary obstruction. The dog is uniquely very sensitive to the effects of glucocorticoids in this regard, in contrast to the cat. The magnitude of increase depends on the dose administered, duration, route and individual sensitivity. Measurement of the steroid-induced isoenzyme is not a reliable test for distinguishing animals with steroid hepatopathy from those with other hepatopathies because activity of the steroid-induced isoenzyme of ALP is variably increased with many types of hepatic diseases and non-hepatic illness.

The activity of ALP is much lower in feline than in canine serum. This is because the half-life of ALP is much shorter in the cat (6 hours versus 3 days), and less feline ALP is produced secondary to biliary obstruction than in the dog because the feline liver contains only one third the concentration of ALP per gram of liver compared to the canine liver. Therefore, even mild elevations of ALP activity in the cat are indicative of marked hepatobiliary disease. The magnitude of increase in ALP activity is most marked with feline hepatic lipidosis, almost always exceeding the magnitude of increase in serum GGT activity in this syndrome.

Gamma-Glutamyl Transpeptidase

In most cases the activity of serum GGT parallels that of ALP, and its measurement is only of occasional value in the dog and cat. Only in feline hepatic lipidosis does the magnitude of increase in serum ALP activity exceed that of serum GGT activity. In general, GGT activity is less influenced by non-hepatic diseases or enzyme-inducing drugs.