Jörg M. Steiner, Dr.med.vet., PhD, DACVIM, DECVIM-CA
Read the German translation: Hilfreiche Laboruntersuchungen zur Diagnose von Erkrankungen des Verdauungstraktes
Diseases of the gastrointestinal tract are a common cause for dogs and cats to be presented to a veterinarian. In order to arrive at the most appropriate diagnosis, veterinarians need to employ diagnostic tests. While no diagnostic test is always positive in a patient with a certain disease and no diagnostic test is always negative in a patient without the disease, the clinician needs to strive to use those tests with the highest clinical accuracy possible. In addition to a high accuracy, diagnostic tests should also be economical and as minimally invasive as possible.
Evaluation of Intestinal Function
Serum Folate Concentration
Folate is a water-soluble B-vitamin (vitamin B9) that is plentiful in most commercial pet foods. However, folate in the diet is mostly supplied as folate polyglutamate, which cannot be readily absorbed. In the proximal small intestine, folate polyglutamate is deconjugated by folate deconjugase and the resulting folate monoglutamate is absorbed by specific folate carriers in the proximal small intestine.
In patients with proximal small intestinal disease, both folate deconjugase and folate carriers can be destroyed. If the disease process is severe enough either folate polyglutamate is no longer deconjugated or folate monoglutamate is no longer absorbed, leading to folate malabsorption. If the condition continues for a significant period of time, folate body stores are depleted and serum folate concentration decreases. The same is true in patients with diffuse small intestinal disease as long as the proximal small intestine is involved in the disease process. Many bacterial species synthesize folate and it is believed that an increased number of bacterial species (i.e., small intestinal bacterial overgrowth) can lead to significant increases in serum folate concentrations.
Serum Cobalamin Concentration
Cobalamin (vitamin B12) is also a water-soluble vitamin that is plentiful in most commercial pet foods. Dietary deficiencies are thus not very common. However, owners who feed their pets exclusively vegetarian diets may inadvertently cause cobalamin deficiency in their pets, unless the vegetarian diet is fortified with cobalamin. Dietary cobalamin is bound to animal-based dietary protein and cannot be absorbed in this form. In the stomach dietary protein is digested by pepsin and HCl and cobalamin is being released. The free cobalamin is immediately bound by R-protein, which is a cobalamin transporter protein that is synthesized in the stomach.1 In the small intestine, R-protein is digested by pancreatic proteases and the free cobalamin is bound by intrinsic factor.1 In dogs and cats the vast majority of intrinsic factor is secreted by the exocrine pancreas.2 This is different from humans where the majority of intrinsic factor is secreted by the stomach.1 Intrinsic factor/cobalamin complexes are absorbed by specific receptors in the ileum.
Distal small intestinal disease, if severe, will lead to destruction of cobalamin receptors in the ileum, leading to cobalamin malabsorption. Cobalamin malabsorption will ultimately lead to depletion of cobalamin body stores and cobalamin deficiency. Diffuse small intestinal disease can also lead to cobalamin malabsorption as long as the ileum is involved in the disease process. Exocrine pancreatic insufficiency also commonly leads to cobalamin deficiency. Finally, an increased number of bacterial organisms in the small intestine will lead to competition for the available cobalamin and may also lead to cobalamin deficiency.
Cobalamin is essential for many biochemical reactions in the body and virtually all tissues need cobalamin for proper function. Clinical signs of cobalamin deficiency can vary. Some patients may just show lethargy, anorexia, and weight loss, while others may show diarrhea, intermittent septic episodes, or even neurological signs. Thus, cobalamin deficiency can lead to further clinical signs and must be addressed therapeutically.3
Fecal α1-Proteinase Inhibitor Concentration
Many gastrointestinal disorders, if severe, can be associated with gastrointestinal protein loss. Traditionally, the gold standard for evaluation of gastrointestinal protein loss is 51Cr-albumin excretion, but this diagnostic test is labor- and time intensive and is also associated with exposure of the patient and personnel to radioactivity. Recently, assays for the measurement of canine and feline α1-proteinase inhibitor concentration in feces have been developed and analytically validated. Alpha1-proteinase inhibitor (α1-PI) is synthesized in the liver and inhibits a variety of different proteinases. Alpha1-proteinase inhibitor has a molecular mass of approximately 60,000 Da, which is similar to that of albumin. Thus, when gastrointestinal disease is severe enough to be associated with gastrointestinal loss of albumin, α1-PI is lost as well. In contrast to albumin, α1-PI is not hydrolyzed by digestive and bacterial proteinases in the gastrointestinal lumen. This is due to the fact that α1-PI is a proteinase inhibitor. Therefore, fecal α1-PI concentration can be used as an estimate for gastrointestinal protein-loss. Currently, assays for measurement of both canine and feline α1-PI are only available through the Gastrointestinal Laboratory at Texas A&M University (http://www.cvm.tamu.edu/gilab).
Clinically, fecal α1-PI concentration should be measured in dogs and cats with hypoalbuminemia that do not have clinical signs of gastrointestinal disease and where an extra-gastrointestinal source of protein loss cannot be identified. Also, dogs belonging to a breed that is associated with a high prevalence of protein-losing enteropathy (e.g., Norwegian Lundehund, Soft Coated Wheaten Terrier, Yorkshire Terrier) that don't have any clinical signs of gastrointestinal disease, but are intended for breeding, should be tested.
Laboratory Evaluation of Hepatobiliary Function
Serum Bile Acids Concentrations
Pre- and postprandial serum bile acids concentrations are used for the diagnosis of severe hepatic dysfunction and portosystemic shunting. Food is withheld from the patient for 12 hours and a serum sample is collected. A small amount of a high-fat food is fed to stimulate gall bladder contraction and another serum sample is collected 2 hours later. When hepatic function is significantly impaired extraction of bile acids from the portal blood becomes less efficient and both pre- and postprandial serum bile acids concentrations may increase. In patients with portosystemic vascular anomalies pre-prandial bile acids concentrations may be only slightly increased, while post-prandial serum bile acids concentrations are often severely increased. This is due to the fact that hepatocytes are functional and are thus able to extract bile acids from the blood, but the blood does not reach the liver. Directly after a fatty meal a large amount of bile acids are being excreted into the duodenal lumen with the bile, are absorbed in the ileum, and reach the vascular space so post-prandial bile acids concentrations tend to be very high. However, over time a small portion of the blood volume circulates through the liver and bile acids are being extracted from that portion of blood so that pre-prandial bile acids concentrations maybe normal or only slightly increased. However, not all patients will show this pattern of serum bile acids concentrations.
Plasma Ammonia Concentration
Ammonia is a metabolite of amino acids. Under physiologic conditions, ammonia is converted to urea in the liver. However, in patients with severe hepatic dysfunction or portosystemic vascular anomaly, ammonia accumulates in the vascular space. Traditionally, measurement of ammonia concentration in plasma was limited to large hospitals or veterinary teaching hospitals as the blood sample needed to be collected, immediately stored on ice, and analyzed within 20 minutes of collection. However, recently several in-clinic analytical devices (e.g., Pocketchem BA), have been introduced that now allow analysis of this parameter inhouse. Unfortunately, plasma ammonia concentration is not very sensitive and hepatic dysfunction has to be severe for plasma ammonia concentrations to be increased. However, an increased plasma ammonia concentration is specific for hepatic dysfunction and/or PSS. The sensitivity of plasma ammonia concentration can be increased by an ammonia challenge. Recently, a Dutch study has suggested that measurement of plasma ammonia concentration is superior to serum bile acids concentrations for the diagnosis of portosystemic shunts in dogs.4 However, further studies are required before routine use of plasma ammonia concentration over serum bile acids concentrations can be recommended.
Laboratory Evaluation of Exocrine Pancreatic Function
Serum Lipase and Amylase Activities
Serum lipase and amylase activities have been used for the diagnosis of human and canine pancreatitis for several decades. However, in both species it has been well recognized that serum lipase and amylase activities are neither very sensitive nor very specific for pancreatitis. Thus, serum amylase and lipase activity are of very limited usefulness for the diagnosis of pancreatitis in dogs and are not useful for the diagnosis of feline pancreatitis.2
Serum Trypsin-like Immunoreactivity Concentration (TLI)
Pancreatic acinar cells synthesize and secrete trypsinogen, an inactive pre-form (zymogen) of trypsin. Almost all trypsinogen is secreted into the duct system and is released into the duodenum. However, a small amount of trypsinogen is also released into the vascular space and can be measured by species-specific immunoassays for measurement of trypsin-like immunoreactivity (TLI). Dogs and cats with exocrine pancreatic insufficiency have a lack of pancreatic acinar cells and thus a severely decreased serum TLI concentration. A serum canine TLI < 2.5 μg/L or a serum feline TLI concentration < 8 μg/L have been shown to be highly specific for EPI.
Serum Pancreatic Lipase Immunoreactivity Concentration (PLI)
Many different cell types in the body synthesize and secrete lipases. In contrast to catalytic assays for the measurement of lipase activity, use of immunoassays does allow for the specific measurement of lipase originated from the exocrine pancreas. Immunoassays for the measurement of serum concentrations of pancreatic lipase (Spec cPL® in dogs and Spec fPL® in cats) have recently been developed and validated. Serum PLI has been shown to be highly specific for exocrine pancreatic function.2 Also, the sensitivity of different minimally-invasive diagnostic tests was compared in dogs with pancreatitis. The sensitivity of serum TLI concentration was below 35%, that of serum lipase activity less than 55%, and ultrasound 68%. In contrast, the sensitivity of serum cPLI concentration for pancreatitis was above 80%.2 Clinical studies in cats have shown similar results. In a study of cats with spontaneous pancreatitis serum fPLI concentration was more sensitive and more specific than serum fTLI concentration or abdominal ultrasonography.5
Thus, in both dogs and cats, serum PLI concentration is the most sensitive and specific diagnostic test for pancreatitis currently available. Also, recently, a patient-side test for the semiquantitative assessment of cPLI concentration, SNAP cPL, has been introduced. A negative SNAP test virtually rules out pancreatitis. A positive test result needs to be followed up by measurement of a Spec cPL in the laboratory to confirm the diagnosis of pancreatitis and to obtain a quantitative result that will allow monitoring the progress of the patient.
1. Dali-Youcef N, Andrès E. An update on cobalamin deficiency in adults. QJM 2009; 102:17-28;
2. Steiner JM. Exocrine pancreas. In: Steiner JM. ed. Small Animal Gastroenterology. Hannover: Schlütersche-Verlagsgesellschaft mbH, 2008;283-306;
3. Ruaux CG, Steiner JM, Williams DA. Early biochemical and clinical responses to cobalamin supplementation in cats with signs of gastrointestinal disease and severe hypocobalaminemia. J Vet Int Med 2005; 19:155-160;
4. Gerritzen-Bruning MJ, van den Ingh TS, Rothuizen J. Diagnostic value of fasting plasma ammonia and bile acid concentrations in the identification of portosystemic shunting in dogs. J Vet Intern Med 2006; 20:13-19;
5. Forman MA, Marks SL, De Cock HEV, et al. Evaluation of serum feline pancreatic lipase immunoreactivity and helical computed tomography versus conventional testing for the diagnosis of feline pancreatitis. J Vet Int Med 2004; 18:807-815.