Liver Enzymes

Front 1

define hepatitis

Back 1

any type of hepatocellular inflammation and necrosis due to various agents (viruses, parasites, bacteria, drug toxicity, alcohol, circulatory failure, autoimmune process, and metabolic disorders)

Front 2

What is fulminant liver failure?

Back 2

Severe liver damage leads to secondary problems:

• encephalopathy within 8 weeks
• coagulopathy
• electrolyte imbalance
• death

Front 3

What are the clinical findings of hepatitis?

Back 3

anorexia, weakness and jaundice (but not diagnostic)

Front 4

What is the treatment for hepatitis?

Back 4

Supportive treatment: low fat, high carbohydrate diet and follow up care is needed to monitor progress

Front 5

What are the 3 phases of hepatitis?

Back 5

acute

subacute

chronic

Front 6

Acute hepatitis is usually caused by what?

Back 6

viruses (hepatitis A, B, C, D, E, G, CMV, EBV and HSV)

Front 7

What is the most common cause of acute hepatitis which is usually short-lived and not chronic?

Back 7

hepatitis A (foodborne virus)

Front 8

What is chronic hepatitis?

Back 8

destruction of the liver due to immune response to the virus

Front 9

What is hepatitis B?

Back 9

HBV is the most common form of chronic hepatitis worldwide

Front 10

Which hepatitis is the most common form of chronic in North America?

Back 10

Hepatitis C

Front 11

What is hepatitis C?

Back 11

• #1 reason for liver transplants
• kills 15,000 per year
• most people don't know they have the virus until there is liver damage
• about 50% of the people who are infected go on to develop chronic infections

Front 12

What are the risk factors associated with hepatitis C?

Back 12

• people who share needles
• healthcare workers exposed to infected blood
  

Front 13

What are the symptoms of hepatitis C?

Back 13

many patients are asymptomatic

• pain in upper right quadrant (liver damage)
• nausea and vomiting
• loss of appetite
• jaudnice
• fatigue
• itching
• can result in cirrhosis
  

Front 14

What is cirrhosis?

Back 14

• It is the end-stage of a number of destructive process affecting the liver due to the chronic "recurring injury" to the liver
• results in destruction of hepatic architecture and loss of the normal structural relationships between portal tracts, hepatocytes and central veins
• morphologically, bands of fibrous tissue encircle parnenchymal liver nodules (tissue scarring)
  

Front 15

What are the most common causes of cirrhosis?

Back 15

hepatitis and alcoholism

• develops in 15% of individuals who drink heavily for more than a decade
• hepatitis B is most common cause world-wide
  

Front 16

What are the clinical manifestations of cirrhosis?

Back 16

• hepatocyte dysfunction: increase in bilirubin, decrease albumin, decrease in clotting factors (can't synthesize it), and increased ammonia and decreased urea (can't convert ammonia to urea)
  
• hemodynamic alterations: fall in platelet count and increase PT (prothrombin time)
  
• increase in alpha fetoprotein
  

Front 17

What are the two MOST consistent features of advanced liver cirrhosis are what?

Back 17

• decreased serum albumin
• increased serum alkaline phosphatase
  

Front 18

What should be known about the enzyme activity levels to interpret conditions?

Back 18

• rate of enzyme release
• rate of enzyme clearance from the blood
  

Front 19

T/F: enzyme activity in the plasma is not helpful in the indicator of cell damage

Back 19

False (enzyme activity is an extremely sensitive indicator of even minor cell damage)

Front 20

Why do enzymes leak from cells?

Back 20

• decreased oxygen leads to deterioration of the plasma membrane so the cells become leaky
• if there is severe damage, cell death occurs so all the cell's contents are released
  
• hypoxia can be caused by hypoxemia (low oxygen content caused by carbon monoxide poisoning or drowning), heart failure and shock
• direct attack on cell membrane by agents (viruses or chemicals such as organic pesticides)
• cell damage and/or trauma
  

Front 21

Why would enzyme production be increased?

Back 21

• increased synthesis due to ethanol and GGT
• increase in cell proliferation such as cancer of prostate and ACP

Front 22

Why would enzyme production be decreased?

Back 22

• genetic deficiency (genetic mutation)
  
• depressed production due to disease (such as decreased production of clotting factors)
  

Front 23

How are enzymes cleared from the plasma?

Back 23

• most enzymes can't be cleared because it's too big to be removed by the kidneys
• only amylase is significantly cleared by the kidneys
• other inactivated enzymes are cleared primarily by receptor-mediated endocytosis
• half-life of enzyme varies between 6-48 hours depending on the enzyme
  

Front 24

What is the function of aspartate aminotransferase (AST)?

Back 24

catalyzes the interconversions of amino acids and 2-oxacids by transfer of amino group

transfers the amine group from L-aspartic acid to 2-oxoglutarate to produce L-glutamic acid and oxaloacetate (reversible to make aspartate or glutamate)

Front 25

Where is AST found?

Back 25

• cytosol and mitochondria
• found in relatively equal amounts in heart, liver and skeletal muscle
• also found in RBCs
• good general indicator of disease
• must be used with other enzyme measurements in order to establish diagnosis
  

Front 26

What is the preferred specimen for aspartate aminotransferase measurement?

Back 26

serum (avoid hemolysis) which the aspartate aminotransferase for about a week at 2-6 degrees (refrigerated)

Front 27

What is the principal for aspartate aminotransferase measurement?

Back 27

clinical assays couple oxaloacetate formed by enzyme activity to a second indicator reaction that results in a colored product UV absorbance change

Front 28

What is the function of alanine aminotransferase?

Back 28

catalyzes the interconversions of amino acids and 2-oxacids by transfer of amino group

transfers of the amine group from L-alanine to 2-oxoglutarate to produce L-glutamic acid and pyruvate (reversible to make alanine or glutamate)

Front 29

Where is ALT found?

Back 29

• found in the cytosol of a variety of tissues but highest concentration in the liver
• ALT is more specific for hepatocellular damage than AST
  
• ALT elevations last longer than AST
  

Front 30

What does measuring the ALT tell you?

Back 30

• increase in ALT is apparent before jaundice
• ALT decreases with bilirubin as liver disease resolves
• persistent elevation of ALT after acute hepatitis is used to diagnose chronic hepatitis
• in obstruction, bilirubin can increase, but if ALT can be normal which indicates no hepatocyte damage

Front 31

What is the principle of alanine aminotransferase measurement?

Back 31

• chemical equilibrium favors formation of alanine so clinical assys force reaction to favor pyruvate formation because pyruvate is easier to measure
• pyruvate produced is coupled to an indicator reaction, either at 340 nm measuring NADH to NAD+ at 505 nm with a chromophore formed (dinitrophenylhydrazine to hydrozone which is blue)
  

Front 32

What is the preferred specimen for alanine aminotransferase measurement?

Back 32

serum (avoid hemolysis) which the aspartate aminotransferase for about a week at 2-6 degrees (refrigerated)

Front 33

What is alkaline phosphatase (ALP)?

Back 33

total ALP varies with age and gender (men more than women, and women increase levels during trimester in pregnancy

Front 34

Where is alkaline phosphatase found in the body?

Back 34

• in the serum, ALP isoenzymes primarily originates from liver and bone
• activity present in liver, bone (osteoblasts), intestinal epithelium (in sera of B & O secretors), kidney tubules and the placenta
• localized in cell membranes
• in the liver, it is found in sinusoidal surface of hepatocytes and in microvilli of bile canaliculi and ducts
  

Front 35

What is alkaline phosphatase useful for?

Back 35

differentiating biliary obstruction

Front 36

Why would alkaline phosphatase serum levels increase?

Back 36

due to cell membrane localized synthesis induced by biliary stasis/duct obstruction, not from cell injury

Front 37

Increased levels of alkaline phosphatase in bone isoenzyme occur in what?

Back 37

rapid bone growth (puberty), pregnancy and bone diseases:

• Paget's disease (osteitis deformans)
• osteomalacia
• Rickets
• cancers
  

Front 38

Alkaline phosphatase's activity is optimal at what pH in vivo?

Back 38

pH 10

Front 39

Alkaline phosphatase's activity is optimal at what pH in vitro?

Back 39

depends on the substrate and magnesium concentration

Front 40

What are the activators of alkaline phosphatase?

Back 40

divalent ions: Mg, Co, Mn and An

Front 41

The correct ratio of what two divalent ions are required for optimal activity of alkaline phosphatase?

Back 41

Mg:Zn

Front 42

What are the inhibitors of alkaline phosphatase?

Back 42

phophate, borate, oxalate and cyanide ions

Front 43

What is the most widely used substrate for ALP?

Back 43

4-nitrophenyl phosphate (4-NPP)

Front 44

What is the preferred specimen for alkaline phosphatase?

Back 44

fasting samples of the serum or heparinized plasma and no hemolysis

Front 45

What should be avoided in specimens collected for alkaline phosphatase?

Back 45

the specimen should NOT have citrate, oxalate (inhibits the enzyme) or EDTA anticoagulants (binds and inactivates divalent ions)

Front 46

What happens to alkaline phosphatase in the specimen over time in storage?

Back 46

its activity increases

Front 47

What are some techniques to differentiate the ALP isoenzymes?

Back 47

• electrophoretic separation at alkaline pH
• heat inactivation
• chemical inhibition
• immunochemical techniques
  

Front 48

Describe electrophoretic separation at alkaline pH for ALP differentiation.

Back 48

• Liver is most anionic, then bone, intestine then kidney
• placental isoenzyme migrates similar to liver and bone
• may require pretreatment of serum such as neuraminidase may be necessary to separate liver and bone fractions
  

Front 49

Describe heat inactivaton for ALP differentiation.

Back 49

• requires tight control of heating conditions with immediate cooling
• after 30 mins at 65 degrees, the placental isoenzyme is the only fraction still active
• after 10 mins at 56 degrees, liver is active, but the bone is not

example: if 80% of enzyme activity is present after 19 mins, then 80% is liver enzyme and 20% was bone

Front 50

Describe chemical inhibition for ALP differentiation

Back 50

varying resistance to inactivation by urea

placental (most resistant) > liver > bone (least resistant)

Front 51

Describe immunochemical techniques

Back 51

• monoclonal and polyclonal antibodies are available
• polyclonal will cross-react
  

Front 52

What is gamma glutamyl transferase?

Back 52

• catalyzes the transfer of aa from 1 peptide to another peptide
• localized in the microsomes and cell membranes
  

Front 53

What is the role of GGT?

Back 53

role in glutathione metabolism in the liver and aa absorption in nephrons and in the intestine

Front 54

If the GGT is elevated, what does it indicate?

Back 54

almost always indicates liver damage: obstruction, necrosis and neoplasms

Front 55

Concentration of GGT vary with what?

Back 55

gender but not age

Front 56

GGT synthesis is induced by what?

Back 56

chronic alcohol and/or drug ingestion

Front 57

Which screening test has been advocated for alcohol abuse?

Back 57

GGT (although alcohol related GGT levels will return to normal 14-16 days after cessation of alcohol consumption)

Front 58

What is gamma glutamyl transferase levels used for?

Back 58

• used to differentiate bone vs. hepatic ALP elevations
• good indicator of cirrhosis, particularly alcoholic cirrhosis, but levels must be interpreted with caution because of induction by exogenous agents
  

Front 59

What is the principle for measuring GGT?

Back 59

the assay utilize gamma-glutamyl-p-nitroanilide as a substrate or donor and glycylglycine as the amino acid receptor which is a coupled reaction and observed spectrophotometrically

Front 60

What is the specimen preferred for GGT analysis?

Back 60

serum preferred, EDTA plasma acceptable, avoid hemolysis

Front 61

What is 5'-nucleotidase?

Back 61

localized in microsomes and cell membranes

Front 62

What is 5'-nucleotidase useful for?

Back 62

• indicator of obstruction of biliary
• good indicator of graft vs. host disease
  
• used to differentiate increased ALP as bone or hepatic
• parallel rise in ALP in liver disease
  
• activity does not vary with gender or age
• not elevated in bone disorders, bone growth or pregnancy
  

Front 63

What is the principle of the assay used to measure 5'-nucleotidase?

Back 63

The principle of the assay used to measure 5'-nucleotidase will vary depending on the technique instrumentation and substrate being used. The enzyme will react only with the nucleoside 5'-phosphate. Adenylate monophosphate or PMP is used as a substrate. The 5'-nucleotidase removes the phosphate to produce adenosine. This reaction is coupled to two additional enzymatic reactions in order to obtain a measurable product. The adenosine is the adenosine deaminase seen which results in the formation of ammonia. The ammonia then reacts with two oxoglutarate and NADH with the help of the enzyme glutamate dehydrogenase to produce glutamate and NAD. The conversion of the NADH to NAD can be measured spectrophotometrically

Front 64

What are the specimen preferred for 5'-nucleotidase analysis?

Back 64

serum preferred, hemolysis avoided