Term
| What is the definition of an isoenzyme? |
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Definition
| Proteins that have a different polypeptide structure, but catalyze the same chemical reaction |
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Term
| What is the term for an enzyme without a cofactor vs. an enzyme with a cofactor? |
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Definition
apoenzyme (without a cofactor)
holoenyme (with a cofactor) |
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Term
| What are most enzyme cofactors derived from? |
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Definition
vitamins (e.g. p-5-p from Vit B6, NAD from niacin)
ions (e.g. Ca, Mg) |
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Term
| What are the five mechanisms though which serum enzyme activity may increase? |
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Definition
Increased release from damaged cells
Induction of enzyme synthesis
Cell proliferation (hyperplasia or neoplasia)
Decreased enzyme clearance
Ingestion and absorption |
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Term
| Why does serum levels of enzymes released from damaged muscle raise slowly? |
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Definition
| These enzymes enter the plasma via lymph rather than directly entering the blood (e.g. RBC and hepatocellular enzymes) |
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Term
| What is the primary mechanism for increased serum [enzyme] for membrane associated enzymes? |
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Definition
Induction -there is a stimulus (endogenous substances (e.g. bile acids) and drugs/toxins (e.g. phenobarbital, prednisone) to increase production of enzymes
**to a lesser extent total tissue mass can also increased membrane associated serum [enyzme] (i.e. increased serum [enzyme] with neoplasia or hyperplasia, and decreased serum [enzyme] with hypoplasia or atrophy) |
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Term
| What is a macroenzyme, and what is the effect on circulating lifpan of the affected enzyme? |
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Definition
| Macroenzymes are enzymes that form complexes with immunoglobulin or other proteins --> this prolongs the circulating life span of these enzymes |
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Term
| Describe the theory of enzyme assays |
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Definition
| in the presence of excess substrate, the rate of a reaction catalyzed by a specific enzyme is depended on the quantity of that enzyme |
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Term
| What is the difference between an end point vs. a kinetic enzyme assay? What type of assay is most commonly used to measure serum enzyme activity? |
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Definition
End point- the reaction is stopped after a specified time and only one reading is obtained (either the amount of product produced or the amount of substrate used)
Kinetic- multiple readings are taken during a specified time period and enzyme activity is determined by the rate of produce formation. <-- most current serum enzyme assays are kinetic |
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Term
| What units are typically used to express enzyme activity? |
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Definition
International units
=amount of enzyme that catalyzes conversion of 1 mmol of substrate/min |
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Term
| What reaction does ALP catalyze? And what cofactors are required? |
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Definition
Compound-PO4 --> compound + PO4
e.g. separates PO4 from any bound other compound
*cofactor fMg+2, alkaline pH |
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Term
| What reaction does ALT catalyze? And what cofactor is required? |
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Definition
Alanine + alpha-ketoglutarate --> pyruvate and glutamate
*cofactor P-5-P (pyridoxal-5'-phosphate)
Pyruvate is then used for gluconeogenesis or Kreb's cycle (ATP production) |
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Term
| What reaction does amylase catalyze? And what cofactors are required? |
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Definition
Starch + H20 --> Starch fragments and unhydrolyzed starch
*cofactor: fCa+2, Cl- |
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Term
| What reaction does AST catalyze? And what cofactor is required? |
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Definition
Aspartate + alpha-ketoglutarate --> oxaloacetate glutamate
*cofactor: P-5-P (pyridoxal-5'-phosphate)
Oxaloacetate is then used in the Kreb's cycle (ATP production) |
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Term
| What enzymes use P-5-P (pyridoxal-5'-phosphate) as a cofactor? |
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Definition
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Term
| What reaction does CK catalyze? And what cofactor is required? |
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Definition
creatinine phosphate + ADP ---> creatinine + ATP
*cofactor: fMg+2 |
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Term
| What reaction does GGT catalyze? |
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Definition
| gamma-glutamyl-p-nitroanilide + glycylglycine --> p-nitroaniline + gamma-glutamyl-glyclglycine |
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Term
| What reaction does glutamate dehydrogenase (GMD) catalyze? |
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Definition
| glutamate + NAD --> 2-oxoglutarate + NADH +NH4 |
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Term
| What reaction does Iditol (aka Sorbitol) dehydrogenase (ID) catalyze? And what cofactor is required |
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Definition
Fructose + NAD --> Sorbitol + NADH
*cofactor: Zn+2 |
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Term
| What reaction does lactate dehydrogenase (LD) catalyze? |
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Definition
| lactate + NAD --> pyruvate + NADH |
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Term
| What reaction does lipase (LPS) catalyze? And what cofactors are required? |
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Definition
triglyceride --> diglycerides + monoglycerides + glycerol + fatty acids
*cofactors: colipase, fCa+2, bile salts |
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Term
| What enzymes are measured in mass per unit volume as opposed to activity level in serum? |
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Definition
| PLI and TLI <-- measured using immunologic assays, reported in ug/L |
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Term
| How are enzymes conventionally named? |
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Definition
| substrate first followed by one of six types of reactions the enzyme catalyzes (e.g. transfer amino = transaminase; transfer phosphate = kinase; oxidize or reduce = oxidoreductase and dehydrogenase) |
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Term
| What enzyme is especially susceptible to deterioration following freezing? |
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Definition
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Term
| What enzymes present in erythrocytes can be falsely increased in a serum sample that has not been removed promptly from a clot and/or if the sample is hemolyzed? |
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Definition
AST, LD
**also chemical constituents of erythrocytes may act as substrates of otherwise participate in enzyme assay reactions affecting results. If the sample is hemolyzed free hemoglobin can interfere with light transmission in spectrophotometric assays (bilirubin and lipemia can also affect spectrophotometric results) |
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Term
| T/F the magnitude of serum enzyme elevations does not differentiate reversible from nonreversible damage or local from diffuse damage |
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Definition
true
* typically the magnitude of increased above URI does correlate with the severity of tissue damage however |
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Term
| What common drugs are associated with increased ALT activity? |
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Definition
corticosteroids (secondary to VH, ALT is not induced by steroids), anesthestic agents, tetracycline, carprofen, phenobarbital
**increased enzyme activity in these cases is not thought to be associated with induction |
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Term
| What muscle diseases are associated with increased ALT? |
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Definition
Canine musculodystrophy (typically <7x URL, but sometimes can be very increased)
Dystrophin deficient cats with acute rhabdomyolysis (6-19x URL) |
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Term
| In what species do hepatocytes contain very little ALT precluding it's value as a marked of hepatic injury? |
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Definition
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Term
| How does ALT compare with AST in dogs and cats as a marked of hepatocellular injury? |
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Definition
| ALT is more specific to hepatocytes than AST but AST has a shorter half life and might be a better indicator of active hepatocellular damage (provided hemolysis and/or muscle injury are not also present) |
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Term
| Which enzymes are cytoplasmic? |
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Definition
| ALT, AST (also mitochondrial), LD, ID (sorbitol dehydrogenase), CK, amylase, lipase |
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Term
| Which enzymes are mitochondrial? |
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Definition
| AST (also cytoplasmic), glutamate dehydrogenase (GMD/GLDH) |
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Term
| What enzymes are associate with cell membranes? |
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Definition
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Term
| What is the clinical use for measuring Iditol (sorbitol) dehydrogenase? |
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Definition
| Increased ID is associated with hepatocellular injury, and may be of more use in cattle and horses because ALT is not present in hepatocytes and AST/LD are not specific to hepatocytes, but it's pretty unstable so not routinely run |
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Term
| Even though GLDH/GMD is not specific to hepatocytes, increased serum concentrations indicated hepatocellular damage because of what factors? |
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Definition
| The enzyme either does not enter plasma (e.g. kidney, intestine), or there is too little enzyme in the tissue to significantly affect serum levels (e.g. muscle, salivary gland) |
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Term
| What is the most sensitive serum enzyme to identify hepatocellular damage in dogs? |
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Definition
| Glutamate dehydrogenase (GMD/GLDH) |
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Term
| In what species does colostrum ingestion markedly increase (up to 30x URL) serum ALP activity? |
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Definition
dogs
*ALP concentrations normalize to pre-suckling values within 10days but will still be elevated in comparison with adult dogs due to increased b-ALP activity
**colostrum intake in calves also increased ALP but only 3x URI |
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Term
| T/F increased serum ALP activity in foals is NOT related to colostrum ingestion |
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Definition
TRUE (colostrum ALP does increased serum ALP in dogs, and to a lesser extent cattle)
increased ALP in foals is related to increased b-ALP |
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Term
| How are the different ALP isoenzymes formed? |
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Definition
Post translational modification of tissue nonspecific ALP
**there is a also a gene for I-ALP, but it doesn't contribute to serum ALP activity due to its short half life |
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Term
| How much to c-ALP contribute to total ALP activity (as measured on routine chemistry assay) in healthy dogs? How does this vary with age? |
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Definition
| c-ALP is 10-30% of total ALP in healthy dogs and increases in contribution as dogs age |
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Term
| What are two methods of estimating the contribution of c-ALP to total ALP activity? |
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Definition
| Levamisole selectively inhibits L-ALP and B-ALP activitiy and can be added to the reaction to determine how much C-ALP is contributing to total ALP activity C-ALP is heat stable compared to L- and B-ALP so comparison of serum ALP activity before and after a hot water bath can help determine the contribution of C-ALP **gold standard method is affinity electrophoresis but this is time consuming and requires special equipment and is not routinely performed |
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Term
| How does wheat germ lectin affect ALP assay? |
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Definition
| B-ALP and C-ALP are selectively precipitated by wheat germ lectin, this can be used to estimate B-ALP activity when combined with levamisole inhibition (inhibits L- and B-ALP) |
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Term
| How does cholestasis induce increased serum ALP activity? |
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Definition
Bile acids stimulate production of L-ALP on sinusoidal hepatocyte membranes (normally limited to canalicular membranes)
*bile acids may also promote release of L-ALP from hepatocyte membranes by promoting the activity of glyosylphosphatidylinositol phospholipase D |
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Term
| what common drugs can induce increased L-ALP production? |
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Definition
corticosteroids, phenobarbitol, primidone
*increased ALP can persist for 2-4 weeks after stoping the drug |
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Term
| What hormone is associated with increased B- and L-ALP? |
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Definition
Thyroxine
**hyperthyroid cats have increased ALP |
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Term
| What is a hypothesized source of ALP in canine mammary neoplasms? |
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Definition
myoepithelial cells
*increased ALP (up to 8x URI) has been reported in both benign and malignant canine mammary neoplasia without liver or bone metastasis |
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Term
| What breed has a familiar hyperphosphatasemia (increased ALP)? |
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Definition
| Siberian Husky (6x URI, B-ALP) |
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Term
| In what species might placental ALP contribute to increased serum ALP in the mother? |
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Definition
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Term
| T/F Increased ALP activity may be noted prior to icterus in dogs with cholestasis |
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Definition
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Term
| What breed of dogs tends to have higher serum ALP activity that other dog breeds? |
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Definition
| Scottish terriers (up to 1350 U/L) |
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Term
| In what species to ALP have poor diagnostic sensitivity for cholestasis? |
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Definition
| Cats and horses * both are typically icteric BEFORE ALP activity is increased |
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Term
| What pattern of ALP and GGT is typical of cats with hepatic lipidosis? |
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Definition
| ALP activity is increased more than GGT |
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Term
| In what species does colostrum have high GGT activity? |
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Definition
| Cattle and dogs, postsuckling calves may have GGT activities 20x URI, 100x URI in pups |
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Term
| What toxins are associated with biliary hyperplasia? |
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Definition
| pyrrolizidine alkaloid-containing plants, sporodesmin, alsike clover |
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Term
| What drugs are associated with GGT induction? |
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Definition
| phenobarbitol, dilantin, primidone, corticosteroids |
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Term
| What enzyme has the best diagnostic sensitivity for diagnosing cholestasis in horses? |
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Definition
| GGT (ALP is also increased but GGT is increased more) |
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Term
| How does age and body size affect serum [CK] in dogs? |
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Definition
higher CK activity in young dogs (< 1yr, highest activity in youngest puppies)
higher mean [CK] in small breed dogs (<10kg) vs. large breed |
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Term
| What creatinine kinase isoenzymes exists and what is their tissue distribution? |
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Definition
CK-1 in brain
CK-2, CK-3 in cardiac and skeletal muscle
CK-MT in mitochondria of many tissues |
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Term
| What is the effect of sample type (serum vs. plasma) on [CK] in dogs? |
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Definition
higher [CK] in serum (~2.5x) compared to plasma, probably from release of CK by platelets during clotting
*difference is unlikely to be clinically relevant |
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Term
| What toxins are most likely to cause muscle injury sufficient to significantly increase serum CK? |
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Definition
| Monensin, castor bean, gossypol |
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Term
| What endocrinopathy in dogs may be associated with increased CK activity in some individuals (and decreased activity in some with the same disease)? |
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Definition
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Term
| What is the proposed mechanism for falsely increased CK activity in samples with in vitro hemolysis? |
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Definition
RBC contain molecules (e.g. glucose-6-phosphate, adenylate kinase, ATP) that interfere with coupled reactions of CK assays
**RBC DO NOT contain CK |
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Term
| In what species can significant smooth muscle injury also result in increased serum [CK]? |
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Definition
Cattle with endometritis
*there was concurrent but relatively minimal increased in AST |
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Term
| How does neurologic disease affect serum CK? |
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Definition
seizures or recumbency can be associated with increased serum [CK] from skeletal muscle injury
Necrosis or demyelination of neural tissues can result in increased CSF [CK], but it doesn't cross the blood brain barrier into serum |
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Term
| Which species have salivary alpha-amylase? |
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Definition
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Term
| What are the major tissue sources that contribute to serum [AMS]? |
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Definition
| pancreas, intestine, and macroamylase (e.g. protein bound) +/- liver |
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Term
| What are the three major types of assays to measure serum amylase activity? |
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Definition
saccharogenic assays: amylase degrades starch into glucose and maltose which are coupled to other reactions
amyloclastic assays: amyalse degrades starch in a reaction chamber with a dye that binds to starches, less dye binding = greater amylase activity (these are preferred in dogs because they have serum glucoamylase and maltase that can falsely increase AMS activity via saccharogenic assays, but this assay is harder to automate)
chromogenic assays: amylase catalyzes the cleavage of a dye bound to a synthetic substrate (some are acceptable for measuring dog AMS activity) |
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Term
| Increases in serum amylase and lipase is most specific for pancreatitis in what species? |
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Definition
Dogs (although some dogs with acute pancreatitis will have serum [amylase and/or lipase] WRI)
**amylase and/or lipase may be increased in cats with acute pancreatitis, but magnitude of increase is less than in dogs. Serum amylase and lipase are totally useless for assessing pancreatic disease in horses and cattle |
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Term
| T/F intestinal amylase has not been shown to increase total serum amylase activity |
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Definition
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Term
| What are six different tissue sources of lipase? |
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Definition
gastric lipase (hydrolyzes ingested TG)
pancreatic lipase (hydrolyzes ingested TG)
Lipoprotein lipase (many cells including adipocytes and myocytes; lipoprotein lipase migrates to luminal surface of endothelial cells to hydrolyze TG in serum lipoproteins)
hepatic lipase (present on endothelial cells of hepatic sinusoids, hydrolyzes TG into LDLs)
hormones sensitive lipase (adipocytes, catalyzes hydrolysis of TG stored in adipocytes and release of free fatty acids)
lysosomal acid lipase (intracellular many tissues, hydrolysis of cholesterol esters) |
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Term
| What hormones stimulate production of hormone sensitive lipase? What is its function? |
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Definition
| epinephrine and glucagon stimulate production of hormone sensitive lipase --> hydrolysis of TG stored in adipocytes and liberation of free fatty acids |
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Term
| What medication has been associated with increased lipase activity in dogs? |
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Definition
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Term
| What is the most sensitive test for diagnosing pancreatitis in cats? |
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Definition
| feline pancreatic lipase immunoreactivity (fPLI) |
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