γ-glutamyl transpeptidase (GGTP)
The enzyme γ-glutamyl transferase (GGT) cleaves C-terminal glutamyl groups from amino acids and transfers them to another peptide or to an amino acid. It is important in glutathione metabolism, amino acid absorption and protection against oxidant injury. Although GGT is found in many tissues, the main source of serum activity is the liver (primarily biliary epithelium), thus GGT is used mainly as a sensitive indicator of cholestasis (interpretation, in general, is similar to ALP), but it also reflects biliary hyperplasia.
It has a half life of 72-96 hours (equine).
GGT is found primarily in the membrane and in microsomes (from smooth endoplasmic reticulum) as aggregates. A small portion (< 5%) is found in the cytoplasm. Disaggregation (solubilization) and increased synthesis result in increased activity in serum. The highest concentrations are found in kidney, pancreas, intestine and mammary glands (the latter in dog, cattle, sheep and goats). In one study in calves and horses, the highest activity in calves was seen in the kidney, followed by the pancreas, liver, lung, small intestine and brain, with very little in skeletal or cardiac muscle. In horses, the highest activity was seen in the kidney followed by the liver, pancreas, brain with low activity in other tissues, including mammary gland, skeletal muscle, small intestine and lung (Barakat and Ford 1988). In the liver, it is found primarily on biliary epithelial cells, but small amounts on canalicular and sinusoidal surfaces of hepatocytes.
- Biliary tract – source of serum activity in health and disease.
- Pancreas, gastrointestinal tract – GGT does not increase in serum in disorders involving these tissues (it is usually shed into the lumen of these organs).
- Kidney – proximal renal tubules. GGT is shed into urine, rather than blood.
- Mammary glands – GGT is excreted into milk, particularly with colostrum.
- Reproductive tract – Epididymis (GGT is secreted into semen).
GGT catalyzes the transfer of the γ- glutamyl group of L-γ- glutamyl-3-carboxy-4-nitroanilide to glycylglycine to generate L-γ-glutamyl-glycylglycine and 5-amino-2-nitrobenzoate. The amount of 5-amino-2-nitrobenzoate produced is measured photometrically and it is proportional to γ-GT activity. The reaction is shown below:
L-γ-glutamyl-3-carboxy-4-nitroanilide + glycylglycine GGT > 5-amino-2-nitrobenzoate + L-γ-glutamyl-glycylglycine
Units of measurement
Enzyme activity is measured in U/L (U = international unit) and µkat/L (SI units), which is defined as the amount of enzyme that catalyzes the conversion of 1 µmol of substrate per minute under specified conditions. Conversion formula is shown below:
U/L x 0.0167 = µkat/L
Serum, plasma, urine
Heparin or EDTA
The stability of GGT activity in human serum and plasma samples are as follows: 7 days at 15 – 25 °C or 2 – 8 °C, and 1 year at -20 °C.
- Lipemia: No significant interference up to 1500 lipemic index.
- Hemolysis: May decrease with moderate to severe hemolysis (≥200 hemolysis index)
- Icterus: Severe icterus may decrease concentrations (>20 mg/dL for unconjugated bilirubin and >50 mg/dL for conjugated bilirubin – the icteric index can be a rough surrogate for these bilirubin concentrations – GGT may be falsely decreased with an icteric index ≥ 20 units).
- Drugs: Increases in GGT occur secondary to therapeutic drugs causing cholestasis. Mild increases may also be seen with anticonvulsant or chronic exogenous corticosteroid therapy, presumably secondary to liver injury causing localized cholestasis or biliary hyperplasia (induction or increased production of GGT cannot be ruled out). Increases in GGT occur as soon as 5 days after corticosteroid administration in dogs but are usually mild.
Increased GGT activity
In adult animals, this is generally thought to reflect two main pathologic processes in the liver: 1) Cholestasis and 2) Biliary hyperplasia. Increases likely occur as a consequence of intratubular pressure within the biliary system. Measurement of GGT activity can be a marker of passive transfer of immunity in some species.
- Artifact: Underfilling of lithium heparin tubes can increase GGT by 10-50% (some above total allowable error of 20% per ASCVP guidelines), particularly at activities <20 U/L (Lippi et al 2012).
- Neonates: Colostrum in most species, except for horses, contains high GGT concentrations. Increases in GGT occur within 24 hours of suckling and are a sensitive indicator of passive transfer. Very high activity of GGT may be observed (up to 1000 x adult levels in 2-3 day old puppies) that gradually decrease with time (over 10 days in dogs, 2-3 weeks in lambs and calves) and is an expected finding in a neonate (i.e. does not indicate hepatobiliary disease) in calves, crias and dogs and cats (Center et al 1991, Weaver et al 2000, Britti et al 2005, Crawford et al 2006). In calves, the increase in GGT has been used to predict efficacy of passive transfer; in one study, GGT < 200 U/L was 80% sensitive for a diagnosis of failure of passive transfer (Perino et al 1993). In another study, calves with GGT activity below 50 U/L had IgG values consistent with failure of passive transfer of immunity (Parish et al 1997), however this testing should be restricted to young calves (<8 days of age) and measurement of IgG concentrations in serum or plasma is the gold standard and superior to GGT activity. The GGT activity gradually decreases after suckling, such that a GGT activity of 54 U/L was equivalent to passive transfer (IgG > 1000 mg/dL by regression analysis) at 17 days of age (Parish et al 1997). In goats, GGT activity does increase immediately after suckling in kids (Braun et al 1984). Although GGT can be high in crias (average of 130 ± 62 U/L in 48 hour old animals), GGT activities were not correlated to serum IgG concentrations, so it is not considered a good marker of passive transfer of immunity in crias (Weaver et al 2000). Even though foals do not derive GGT from the colostrum of mares, GGT can be higher in neonates (Braun et al 1984, Patterson and Brown 1986). In one study, GGT activity increased transiently (up to 169 U/L) in the 7-28 days after birth, before decreasing to within reference intervals by 2 months of age. Note that increases in ALP activity do occur after suckling. This is thought to be derived from intestinal sources in the neonate and is not due to colostrum itself.
- Breed: Some donkeys and burros can have 2-3 x GGT activity of horses, with a published reference interval (per ASVCP guidelines) of 14-69 U/L (Burden et al 2015) versus 8-33 U/L for our reference interval for horses.
- Pathophysiologic: Increases in GGT occur secondary to biliary hyperplasia (increased pressure within the biliary system stimulates hyperplasia) or induction of synthesis.
- Hepatobiliary disease:
- In small animals, GGT is a sensitive indicator of biliary hyperplasia and cholestasis. In the cat, GGT increases may precede increases in ALP and is a more sensitive indicator of liver disease in this species (not in all circumstances; hepatic lipidosis is a notable exception) (Center et al 1986). GGT does not increase in small animals treated with carbon tetrachloride (CCl4) toxicity (which causes centrilobular heptocyte necrosis).
- In large animals, GGT is a sensitive test for biliary hyperplasia (it is a good marker for pyrrolizidine alkaloid toxicity in ruminants) and cholestasis (which is relatively uncommon in large animals). Overall, GGT is considered a better marker of biliary tract disorders in large animals than ALP.
- Horses: GGT activity has been reported to be increased in horses with cholangiohepatitis (e.g. secondary to cholelithiasis or proximal enteritis) or gastrointestinal problems, including proximal enteritis and colonic displacement (Davis et al 2003, Gardner et al 2005) (high GGT activity may be seen with other gastrointestinal disorders in horses, such as small intestinal strangulating obstruction (Johnstone and Morris 1987)) In the Gardner 2005 study, 48% and 2% of 37 and 45 horses with right and left colonic displacement, respectively, had GGT activities above the reference interval (as high as 489 and 92 U/L, respectively). Although the mechanism of the GGT increase is not always clear in an individual horses, some horses with colonic displacement and cholangiohepatitis do have biochemial (increased direct bilirubin in serum) and/or histologic evidence of biliary hyperplasia or cholestasis (Davis et al 2003, Gardner et al 2005). Cholestasis with right colonic displacement could be due to stretching and direct compression of the bile duct by the displaced organ (Gardner et al 2005), whereas cholestasis with proximal enteritis could be secondary to cell swelling from hepatic injury, which may be in response to an ascending bacterial infection from the intestine or absorption of bacterial toxins via the portal circulation. Vacuolar change is seen in hepatocytes and biliary epithelial cells in this disorder and some animals have a mixed lymphoplasmacytic or suppurative cholangiohepatitis on histopathologic evaluation of the liver. A functional cholestasis (inflammatory cytokine downregulation of bile or bilirubin transporters may also be operative in some animals. With cholangiohepatitis, particularly that due to cholelithiasis, a higher fold increase in GGT compared to AST and ALP activity can be a useful diagnostic feature (Peek and Divers 2000). Monitoring of GGT activity is useful in assessing response to antibiotic therapy in this disorder. GGT activity has been shown to increase with 1 day of treatment of 3 horses with chlorof0rm and remained high for 16 days (Barakat and Ford 1988). However, it is not clear if this is truly due to necrosis (of biliary epithelium or hepatocytes) or secondary to localized cholestasis (such as from hepatocyte swelling). The sustained increases in GGT activity in the latter study likely reflect biliary hyperplasia.
- Ruminants: In one study of hepatic injury in ruminants, there were no consistent changes in GGT activity after administration of chloroform in 4 calves, whereas GLDH and SDH activities increased (Barakat and Ford 1988). A mild increase in GGT activity was seen in the latter study in 1 of two sheep 1 day after administration of chloroform with the increases being sustained for 16 days. However, since number of tested animals were low in the latter study and no control was administered, these results should be interpreted with caution.
- Although very low activity of GGT are normally found in hepatocytes from adults (fetal hepatocytes do express GGT), liver disease (viral disease, neoplasia) can induce GGT expression in hepatocytes (often on their sinusoidal surface), so increases in serum GGT activity can reflect biliary (primarily) or hepatocellular disease.
- Renal disease: GGT is expressed on the membranes of proximal renal tubular epithelial cells. Cell injury causes GGT to be shed into the urine and not into blood. The urinary GGT to creatinine ratio has been studied as an early indicator of renal tubular injury, especially due to aminoglycoside toxicity (Garry et al 1990, Grauer et al 1995).
- Hepatobiliary disease:
- In cats with hepatic lipidosis, ALP activity will be increased early in the progression of the disease, followed by an increase in bilirubin concentrations (combination of direct and indirect) and finally in GGT activity(see image to the right). Thus, increases in ALT, AST and ALP activities with hyperbilirubinemia is often indicative (but not 100% specific for) lipidosis.
- Cats can show higher increases in GGT over ALP activity in cases of necroinflammatory disorders involving the biliary structures or pancreas.
- Cats with hyperthyroidism may show mild increases in ALP, AST and ALT activities, whereas GGT activity and total bilirubin concentrations are normal.
- GGT may be increased to a greater degree than ALP activity in diseases associated with biliary hyperplasia in the absence of cholestasis (eg biliary carcinoma, pyrrolizidine toxicosis or Fasciola infections in cattle).
- Sustained increases in ALP and GGT activity have been recognized to occur during the healing phase following hepatic necrosis (Barakat and Ford 1988).