Module 1: Liver Chemistries (PPT&BISHOP)

Liver

Gross Anatomy

 located below the diaphragm

 abdominal upper right quadrant
 largest internal organ
 1.2 to 1.5 kg in a healthy adult
 extremely vascular organ that receives its blood supply from two major sources:
o hepatic artery 
 branch of the aorta
 supplies oxygen-rich blood from the heart to the liver
 responsible for providing approximately 25% of the total blood supply to the liver
o portal vein
 supplies nutrient-rich blood (collected as food is digested) from the digestive tract
 responsible for providing approximately 75% of the total blood supply to the liver
 two blood supplies eventually merge into the hepatic sinusoid
o lined with hepatocytes capable of removing potentially toxic substances from the blood
 From the sinusoid
o blood flows to the central canal (central vein) of each lobule
 through the central canal that blood leaves the liver
 right lobe is approximately six times larger into two lobes than the left lobe
 Approximately 1,500 mL of blood passes through the liver per minute
 drained by a collecting system of veins that empties into the hepatic veins and ultimately into the
inferior vena cava
 Excretory system
o begins at the bile canaliculi
 canaliculi are small spaces between the hepatocytes that form intrahepatic ducts
 where excretory products of the cells can drain
 2 major cells of liver
o hepatocytes
 making up approximately 80% of the volume of the organ
 large cells that radiate outward in plates from the central vein to the periphery of the
lobule
 perform the major functions associated with the liver
 responsible for the regenerative properties of the liver
o Kupffer cells
 macrophages that line the sinusoids of the liver
 act as active phagocytes capable of engulfing bacteria, debris, toxins, and other
substances flowing through the sinusoids

4 Major biochemical functions

o Synthetic Function
 makes:
 plasma proteins (albumin & globulin)
 cholesterol
 triglycerides
  lipoproteins
 produces bile
o Detoxification and Excretion
 converts blood ammonia to  urea ( for kidney excretion)
 excretion of heme waste products
 breaks own acetaminophen, alcohol and other drugs
o Strorage
 Storage of Vitamins A,D,E,K and B12
o Transformation
 Metabolizes carbohydrates
 Breakdown and synthesis of cholesterol

Question: Imagine if the liver has difficulty performing its functions due to injury , what do you think will
happen?

Answer: Your abdomen might swell accompanied by painful episodes. Or Jaundice can appear.

Explanation:

 Jaundice occurs when the liver is not able to remove the heme waste product called bilirubin (yellow
pigment) from the blood causing the yellowing of the sclera and the skin and imparting dark color to the
urine.
 Clinically, jaundice appears when the serum bilirubin is greater than 2.5 mg/dL, or more commonly
when it reaches 3.0 mg/dL.
 “Approximately, 200-300 mg of bilirubin is produced per day and it takes a normally functioning
liver to process the bilirubin and eliminate it from the body” (Bishop et, al) .
 Largely, after processing, this waste is excreted in the feces and in the urine in small quantity.
 Jaundice can be classified based on the site of disorder as pre-hepatic, hepatic and post-hepatic.
Knowing this will allow doctors how to manage patients with this condition.

Question:

 When does pre-hepatic, hepatic and post-hepatic jaundice occur? And what happens in these instances?
1 (question #1) Cite examples of condition under these categories of Jaundice.
 Also, what else can you think of the probable consequences of having damaged liver cells? 2 (question
#2)
 Like, what if it cannot make adequate proteins or metabolize the carbohydrates that we eat or cannot
process the ammonia in our blood?

Question: But what if I do not have those signs or symptoms, how can I be sure that my liver is functioning
well?

Answer:

 There are tests called “Liver Chemistry Tests” that can be done to evaluate if you have a healthy liver.
 Hepatic panel consists of blood tests that will measure the levels of proteins, enzymes or bilirubin in
your blood to diagnose or monitor liver disease or liver damage.
  As the liver performs its various functions, it produces metabolites that are released into the
bloodstream.
 In the presence of liver disorders, the levels of these metabolites may be altered that can provide us a
clue on the type of abnormality. Hence we can do liver tests.
 From our definition, Liver Chemistries may be classified as:
o Bilirubin tests
 can be serum bilirubin tests or urobilinogen tests.
o Liver enzyme tests
 can be measurement of albumin, globulin or prothrombin time
o Liver protein tests
 can consist of measurement of ALT, AST, GGT etc.

Biochemical Functions Bishop's

Excretory and secretory

 important functions of the liver :

o processing and excretion of endogenous and exogenous substances into the bile or urine such
as the major heme waste product, bilirubin.
 liver
o only organ that has the capacity to rid the body of heme waste products
 Bile is made up of:
o bile acids or salts,
o bile pigments
o cholesterol
o other substances extracted from the blood
 body produces :
o approximately 3 L of bile per day
 body excretes :
o 1 L of what is produced
 Bilirubin
o principal pigment in bile
o derived from the breakdown of red blood cells
o Approximately 126 days after the emergence from the reticuloendothelial tissue
 red blood cells are phagocytized and hemoglobin is released
o Hemoglobin is degraded to :
 heme
 converted to bilirubin in 2 to 3 hours.
 globin
 degraded to its constituent amino acids
 reused by the body
 iron
 bound by transferrin
 returned to iron stores in the liver or bone marrow for reuse
o Bilirubin is bound by albumin and transported to the liver
 referred to as unconjugated or indirect bilirubin
o Unconjugated bilirubin
 insoluble in water
 cannot be removed from the body until it has been conjugated by the liver
 Once at the liver cell:
  unconjugated bilirubin flows into the sinusoidal spaces
 released from albumin so it can be picked up by a carrier protein called ligandin
 Ligandin
 which is located in the hepatocyte
 responsible for transporting unconjugated bilirubin to the
endoplasmic reticulum, where it may be rapidly conjugated
o Conjugation (esterification) /  Conjugated bilirubin
 occurs in the presence of the enzyme uridine diphosphate glucuronosyltransferase
(UDPGT)
 which transfers a glucuronic acid molecule to each of the two propionic acid side chains
of bilirubin to form bilirubin diglucuronide, also known as conjugated bilirubin
 water soluble
 able to be secreted from the hepatocyte into the bile canaliculi
 Once in the hepatic duct
 it combines with secretions from the gallbladder through the cystic duct
 expelled through the common bile duct to the intestines
 Intestinal bacteria
 especially the bacteria in the lower portion of the intestinal tract
 work on conjugated bilirubin to produce mesobilirubin,
 which is reduced to form mesobilirubinogen and then urobilinogen (a colorless
product)
 urobilinogen formed (roughly 80%)
 oxidized to an orange-colored product called urobilin (stercobilin)
 excreted in the feces.
 gives stool its brown color
 2 things that can happen to the remaining 20% of urobilinogen formed
 majority will be absorbed by extrahepatic circulation to be recycled
through the liver and re-excreted
 other very small quantity left will enter systemic circulation and will
subsequently be filtered by the kidney and excreted in the urine
Detoxification and Drug Metabolism

 liver
o serves as a gatekeeper between substances absorbed by the gastrointestinal tract and those
released into systemic circulation
 first pass
o Every substance that is absorbed in the gastrointestinal tract must first pass through the liver
o can allow important substances to reach the systemic circulation
o can serve as a barrier to prevent toxic or harmful substances from reaching systemic circulation
 body has two mechanisms for detoxification of foreign materials (drugs and poisons) and metabolic
products (bilirubin and ammonia)
o Either it may bind the material reversibly to inactivate the compound
o or it may chemically modify the compound so it can be excreted in its chemically modified form
 drug metabolizing system of the liver
o responsible for the detoxification of many drugs through:
 oxidation,
 reduction
 hydrolysis
 hydroxylation
 carboxylation
 demethylation.
 Many of these take place in the liver microsomes via the cytochrome P-450 isoenzymes.

Liver Function Alterations during disease

Jaundice

 comes from the French word jaune, which means “yellow"

 one of the oldest known pathologic conditions reported (Hippocratic physicians)
 also known as icterus
o most commonly used in the clinical laboratory to refer to a serum or plasma sample with a
yellow discoloration due to an elevated bilirubin level
 used to describe the yellow discoloration of the skin, eyes, and mucous membranes most often resulting
from the retention of bilirubin
 may also occur due to the retention of other substances.
 Although the upper limit of normal for total bilirubin is 1.0 to 1.5 mg/dL:
o jaundice is usually not noticeable to the naked eye (known as overt jaundice) until bilirubin
levels reach 3.0 to 5.0 mg/dL
 most commonly classified based on the site of the disorder:
o prehepatic jaundice
o hepatic jaundice
o posthepatic jaundice
 Prehepatic and posthepatic jaundice
 caused by abnormalities outside the liver, either before, as in “prehepatic,”
or after, as in “posthepatic.”
 In these conditions, liver function is normal or it may be functioning at a
maximum to compensate for abnormalities occurring elsewhere.
 hepatic jaundice
   due to a problem with the liver itself
  intrinsic liver defect or disease.

Prehepatic jaundice

 occurs when the problem causing the jaundice occurs prior to liver metabolism
 most commonly caused by an increased amount of bilirubin being presented to the liver
 seen in acute and chronic hemolytic anemias
 Hemolytic anemia
 causes an increased amount of red blood cell destruction and the
subsequent release of increased amounts of bilirubin presented to the liver
for processing
 The liver responds by functioning at maximum capacity;
 therefore, people with prehepatic jaundice rarely have bilirubin levels that exceed
5.0 mg/dL because the liver is capable of handling the overload
 also known as unconjugated  hyperbilirubinemia
 because the fraction of bilirubin increased in people with prehepatic jaundice is
the unconjugated fraction
 This fraction of bilirubin (unconjugated bilirubin) is :
 not water soluble
 bound to albumin
 not filtered by the kidneys
 not seen in the urine

Hepatic jaundice

 occurs when the primary problem causing the jaundice  resides in the liver (intrinsic liver
defect or disease)
 can be due to disorders of bilirubin metabolism and transport defects
 Crigler-Najjar syndrome
 Dubin-Johnson syndrome
 Gilbert's disease
 neonatal physiologic jaundice of the newborn)
 or due to diseases resulting in hepatocellular injury or destruction.
 Gilbert's disease, Crigler-Najjar syndrome, and physiologic jaundice of the newborn are:
 hepatic causes of jaundice that result in elevations in unconjugated bilirubin
 Conditions such as Dubin-Johnson and Rotor's syndrome are :
 hepatic causes of jaundice that result in elevations in conjugated bilirubin

Gilbert Syndrome

 first described in the early twentieth century

 a benign hereditary disorder that affects approximately 5% of the U.S. population.
 Of the many causes of jaundice, Gilbert syndrome is the most common cause
 interestingly, it carries no morbidity or mortality in the majority of those affected and carries generally
no clinical consequences
 It is characterized by intermittent unconjugated hyperbilirubinemia in the absence of hemolysis and
underlying liver disease due to a defective conjugation system
 The hyperbilirubinemia usually manifests during adolescence or early adulthood.
  Total serum bilirubin usually fluctuates:
o between 20–50 umol/L, and it rarely exceeds 85 umol/L.
 The molecular basis of Gilbert syndrome (in whites) is related to  the UGT (uridine diphosphoglucose
glucuronyltransferase) superfamily
o which is responsible for encoding enzymes that catalyze the conjugation of bilirubin.
o The UGT1A1 (the hepatic 1A1 isoform of UGT)
 contributes substantially to the process of conjugating bilirubin
o The UGT1A1 promoter contains the sequence (TA)6TAA.
o The insertion of an extra TA in the sequence, as seen in Gilbert syndrome, reduces the
expression of the UGT1A1 gene to 20%–30% of normal values.
 That is, the liver’s conjugation system in Gilbert syndrome is working at approximately 30% of normal

Crigler-Najjar syndrome

 was first described by Crigler and Najjar in 1952

 syndrome of chronic nonhemolytic unconjugated hyperbilirubinemia
 an inherited disorder of bilirubin metabolism resulting from a molecular defect within the gene involved
with bilirubin conjugation
 may be divided into two types:
o type 1 -  where there is a complete absence of enzymatic bilirubin conjugation
o type II - where there is a mutation causing a severe deficiency of the enzyme responsible for
bilirubin conjugation.
 rare and is a more serious disorder

Note:

 Gilbert disease and Crigler-Najjar syndrome

o unconjugated hyperbilirubinemias
 Dubin-Johnson syndrome and Rotor syndrome
o conjugated hyperbilirubinemias

Dubin-Johnson syndrome

 a rare inherited disorder caused by a deficiency of the canalicular multidrug resistance/multispecific

organic anionic transporter protein (MDR2/cMOAT)
 the liver’s ability to uptake and conjugate bilirubin is functional
o however, the removal of conjugated bilirubin from the liver cell and the excretion into the bile
are defective.
o This results in accumulation of conjugated and, to some extent, unconjugated bilirubin in the
blood, leading to hyperbilirubinemia and bilirubinuria
 obstructive in nature
 so much of the conjugated bilirubin circulates bound to albumin.
 This type of  bilirubin (conjugated bilirubin bound to albumin)
o referred to as delta bilirubin
o An increase in delta bilirubin poses a problem in laboratory evaluation
o delta bilirubin fraction reacts as conjugated bilirubin in the laboratory method to measure
conjugated or direct bilirubin
  distinguishing feature :
o appearance of dark-stained granules (thought to be pigmented lysosomes) on a liver biopsy
sample
 Usually the total bilirubin concentration remains :
o between 2–5 mg/dL with more than 50% due to the conjugated fraction
 relatively mild in nature with an excellent prognosis.
 People with Dubin- Johnson have a normal life expectancy, so no treatment necessary

Rotor syndrome

 clinically similar to Dubin-Johnson syndrome but the defect causing Rotor syndrome is not known.
 It is hypothesized to be due to a reduction in the concentration or activity of intracellular binding
proteins such as ligandin.
 Unlike in Dubin-Johnson syndrome,
o a liver biopsy does not show dark pigmented granules
 seen less commonly
 a relatively benign condition
 carries an excellent prognosis
 therefore treatment is not warranted.
 However, an accurate diagnosis is required to aid in distinguishing it from more serious liver diseases
that require treatment

Physiologic jaundice of the newborn

 a result of a deficiency in the enzyme glucuronyl transferase

o one of the last liver functions to be activated in prenatal life
o since bilirubin processing is handled by the mother of the fetus
 In premature births, infants may be born without  glucuronyl transferase
o the enzyme responsible for bilirubin conjugation
 This deficiency results in the rapid buildup of unconjugated bilirubin
o which can be life threatening.
 When this type of bilirubin builds up in the neonate
o it cannot be processed
o it is deposited in the nuclei of brain and nerve cells, causing kernicterus

Kernicterus

 often results in cell damage and death in the newborn

 this condition will continue until glucuronyl transferase is produced
 Infants with this type of jaundice are usually treated with ultraviolet
o to destroy the bilirubin as it passes through the capillaries of the skin
 In extreme cases, some infants require an exchange transfusion
 bilirubin levels are carefully and frequently monitored so the dangerously high levels of unconjugated
bilirubin (approximately 20 mg/dL) can be detected and treated

Posthepatic jaundice
  results from biliary obstructive disease, usually from physical obstructions (gallstones or
tumors), that prevent the flow of conjugated bilirubin into the bile canaliculi.
 Since the liver cell itself is functioning
 bilirubin is effectively conjugated;
 however, it is  unable to be properly excreted from the liver.
 Since bile is not being brought to the intestines,
 stool loses its source of normal pigmentation and becomes clay-colored

Cirrhosis

 a clinical condition in which scar tissue replaces normal, healthy liver tissue
 As the scar tissue replaces the normal liver tissue
o it blocks the flow of blood through the organ and prevents the liver from functioning properly
 rarely causes signs and symptoms in its early stages
 but as liver function deteriorates the signs and symptoms appear, including :
o fatigue
o nausea
o unintended weight loss
o jaundice
o bleeding from the gastrointestinal tract
o intense itching
o swelling in the legs and abdomen
 some patients may have prolonged survival but they have poor prognosis
 Causes:
o chronic alcoholism
o chronic  hepatitis C virus infection
o chronic hepatitis B and D virus infection
o autoimmune hepatitis
o inherited disorders
 alpha1-antitrypsin deficiency,
 Wilson disease
 hemachromatosis
 galactosemia
o nonalcoholic steatohepatitis
o blocked bile ducts
o drugs
o toxins
o infections
 cannot easily be reversed
o but treatment can stop or delay further progression of the disorder
 Treatment depends on the cause of cirrhosis and any complications a person is experiencing
o For example
 cirrhosis caused by alcohol abuse is treated by abstaining from alcohol
 Treatment for hepatitis-related cirrhosis:
 involves medications used to treat the different types of hepatitis
 interferon for viral hepatitis
 corticosteroids for autoimmune hepatitis
Tumor

 Cancers of the liver are classified as primary or metastatic.

 Primary liver cancer
o is cancer that begins in the liver cells.
 Metastatic cancer
o occurs when tumors from other parts of the body spread (metastasize) to the liver
o much more common than primary liver cancer
o 90%–95% of all hepatic malignancies are classified as metastatic.
 Cancers that commonly spread to the liver include:
o colon
o lung
o breast cancer.
 Cancers of the liver may also be classified as benign or malignant
o The common benign cancers of the liver include:
 hepatocellular adenoma
 rare condition occurring almost exclusively in females of child-bearing age
 hemangiomas
 masses of atypical blood vessels usually mesenchymal in origin with no
known etiology
o Malignant tumors of the liver include :
 hepatocellular carcinoma (HCC)
 most common
 develops in those with liver cell damage that eventually progresses to cirrhosis,
which is a predisposing condition for the development
 of HCC
 hepatocarcinoma
 hepatoma

Reye syndrome

 is a term used to describe a group of disorders caused by:

o infectious
o metabolic
o toxic
o druginduced disease
 found almost exclusively in children although adult cases of Reye syndrome have been reported.
 precise cause of Reye syndrome is unknown
 it is often preceded by a viral syndrome such:
o varicella
o gastroenteritis
o an upper respiratory tract infection
 influenza
 strong epidemiologic association between the ingestion of aspirin during a viral syndrome and the
subsequent development of Reye syndrome
 an acute illness characterized by:
o noninflammatory encephalopathy
o fatty degeneration of the liver
o with a clinical presentation of profuse vomiting accompanied
 o with varying degrees of neurologic impairment such as :
 fluctuating personality changes and deterioration in consciousness.
 encephalopathy
o characterized by a progression from mild confusion (stage 1) through progressive loss of
neurologic function to loss of brainstem reflexes (stage 5)
 degeneration of the liver
o characterized by a mild hyperbilirubinemia
o and threefold increases in ammonia and the aminotranferases (AST and ALT)
o Without treatment, rapid clinical deterioration leading to death may occur

Drug- and Alcohol-Related Disorders

 liver
o is a primary target organ for adverse drug reactions
o because it plays a central role in drug metabolism
 Many drugs are known to cause liver damage:
o ranging from very mild transient forms to fulminant liver failure
 mechanism of toxicity
o via an immune-mediated injury to the hepatocytes
o the drug induces an adverse immune response directed against the liver itself
o and results in hepatic and/or cholestatic disease
 Of all the drugs associated with hepatic toxicity:
o the most important is ethanol
o In very small amounts, ethanol causes very mild, transient, and unnoticed injury to the liver
 however, with heavier and prolonged consumption, it can lead to alcoholic cirrhosis.
 Approximately 90% of the alcohol absorbed from the stomach and small intestines
o transported to the liver for metabolism
o Within the liver, the elimination of alcohol requires the enzymes:
 alcohol dehydrogenase
 acetaldehyde dehydrogenase
o to convert alcohol to acetaldehyde and subsequently to acetate
o acetate can then be oxidized to water and carbon dioxide,
o or it may enter the citric acid cycle.
 Long-term excessive consumption
o liver abnormalities
o alcoholic fatty liver with inflammation (steatohepatitis)
o scar tissue formation, as in hepatic fibrosis,
 destruction of normal liver structure seen in hepatic cirrhosis.
 liver injury category
o alcoholic fatty liver
 mildest category
 very few changes in liver function are measurable
 slight elevations in AST, ALT, and GGT, on biopsy,
 fatty infiltrates are noted in the vacuoles of the liver.
 tends to affect young to middle-aged people with a history of moderate alcohol
consumption
 complete recovery within 1 month is seen when the drug is removed
o alcoholic hepatitis
  more evidence of liver damage
 moderately elevated AST, ALT, GGT, and ALP
 and elevations in total bilirubin up to 30 mg/dL.
 Serum proteins, especially albumin
 decreased
 prothrombin time is prolonged.
o alcoholic cirrhosis
 dependent on the nature and severity of associated conditions such as a gastrointestinal
bleeding or ascitis;
   5-year survival rate ;
 60% in those who abstain from alcohol
 30% in those who continue to drink.
 more common in males than in females,
 symptoms tend to be nonspecific and include:
 weight loss
 weakness
 hepatomegaly
 splenomegalY
 jaundice
 ascites
 fever
 malnutrition
 edema
 Laboratory abnormalities
 increased liver function tests (AST,ALT, GGT, ALP, total bilirubin)
 decreased albumin
 a prolonged prothrombin time.
 drugs causes liver injury
 tranquilizers
 some antibiotics
 antineoplastic agents
 lipid-lowering medication
 antiinflammatory drugs,

ASSESSMENT OF LIVER FUNCTION/LIVER FUNCTION TESTS

Liver function test  (LFTs) from the video

 hepatic panel
 blood tests used to monitor liver function and damage
 provide insights into several aspects of liver health
o Synthetic function - liver's ability to synthesize enzymes and proteins
 can be assessed based on its ability to produce plasma
 albumin
 albumin (normal : 40 to 6g/L)& coagulation factors
 serum albumin levels FALL with all liver diseases
 low albumin = liver disease
 liver disease
 Non-hepatic causes:
 low protein intake
  Malabsorption
 protein loss in urine
 prothrombrin time (PT)
 pro time test
 normal PT :  10.9 to 12.5 secs
 measures the time the blood takes to clot
 a decline in the liver function leads to less coagulation factors produced and
delays coagulation time
 High PT -  Liver disease
 Non hepatic causes:
 Bleeding disorder
 Vitamin K deficiency
 use of blood thinning medications (warfarin)
o Bilirubin processing - liver's ability to process bilirubin and secrete bile
 bilirubin
 Alkaline phosphatase (ALP)
 gamma-glutamyltransferase (GGT)
 Bilirubin
 unconjugated bilirubin (water insoluble) ---> conjugated bilirubin (water
soluble)--> bile drains into small intestine
 a water insoluble product of normal heme breakdown
 transported to the liver loosely bound to albumin
 liver converts into a water-soluble form to be secreted into bile
 accumulation of bilirubin in the blood indicates problems with biliary function
 high levels of bilirubin give the skin and the whites of the eyes  a yellowish color
known as jaundice
 normal bilirubin: 2 to 17umol/L
 Other enzymes measured for biliary function
 Alkaline phosphatase (ALP) - normal: 30 to 120IU/L
 found in: bone, intestines, placenta
 ALP elevations may also be due to a number of non-hepatic causes
 bone diseases
 chronic real failure
 lymphoma, infection
 Normal ALP are higher in children/teens, preganancy, aging
women
 gamma-glutamyltransferase (GGT) - normal: 0 to 30IU/L
 found in: kidney, pancreas, intestines
 Elevated GTT is more specific for biliary disease compared to
ALP (not present in bones)
 GTT are higher in infants
 both enzymes are found in the tiny bile ducts (canaliculi of the liver)
 present in several other tissues
 damage to the biliary tract releases these enzymes into the bloodstream
o Liver damage
 when liver cells are injured, their content including liver enzymes, are leaked into the
blood streams
 The levels of these enzymes can be measured to assess the extent of the liver damage
 Increased level enzymes = Liver damage
 2 enzymes are usually included in a hepatic panel
 aspartate transaminase (AST) = normal: 0 to 35 IU/L
 alanine transaminase (ALT) = normal: 0 to 45IU/L
  more sensitive and specific for liver damage than AST
 lower level in other tissues
 both are involved in protein metabolism in the liver
 are present at high concentrations in the liver
 but also found in a number of other tissues
 heart
 skeletal muscles
 kidneys
 brain
 pancreas
 lungs
 Normal AST and ALT values are higher in men than women; higher with obesity
 may also result from non-hepatic causes
 heart attacks
 muscle diseases
 lungs
 abnormal liver function test do not always indicate liver disease
o some abnormalities are transient (impermanent)
o or they may result form different, non-hepatic causes

Liver Chemistries

Detoxification &Excretion Function Test

 Serum Bilirubin
 Plasma Amonia

Liver Cell damage Test

 ALT&AST
 ALP
 GGT

Synthetic Function Test

 Serum Albumin & Globulin

 Prothrombin Time

Serum Bilirubin Test

Bilirubin

 is the breakdown product of heme metabolism from red blood cell’s hemoglobin.

Liver cells

 take this unconjugated bilirubin to render it water soluble by attaching to it glucuronic acid
molecule.

Once conjugated
  this bilirubin can now be released from the hepatocytes and mix up with other gallbladder
secretions and then be released in the common bile duct in the intestine

An elevated blood level of conjugated bilirubin

 occurs in various liver and bile duct conditions.

 It is particularly high if the flow of bile is obstructed
 For example:
o by a gallstone stuck in the common bile duct.
o It can also be increased with hepatitis, liver injury, or long-term alcohol abuse.

An increased level of unconjugated bilirubin

 occurs when there is excessive destruction of erythrocytes like in the case of hemolytic anemia.

Methods of Bilirubin Determination

 Jendrassik & Grof

o bilirubin reacts with a diazo reagent which results in the production of purple product called
azobilirubin
 diazo reagent:  two solutions, one of sodium nitrite, the other of
acidified sulfanilic acid, used in bringing about diazotization. (Ehrlich
diazo reagent)
o Caffeine-benzoate
 accelerates the coupling of bilirubin with the diazo reagent
o Ascorbic acid
 stops the reaction
o Alkaline tartrate
 converts the purple azobilirubin to a blue azobilirubin
o This product is measured spectrophotometrically at 600 nm.
o Advantages
 not affected by pH changes
 maintains optical sensitivity at low bilirubin concentrations
 Insensitive to high protein concentrations

 Evelyn &Malloy Method

o This method of bilirubin is based on Van Den Bergh reaction
o Bilirubin reacts with diazotized  sulfanilic acid to produce purple colored azobilirubin.
o Methanol is added to solubilize unconjugated bilirubin
o susceptible to hemoglobin interference

Plasma Ammonia

 Ammonia
o waste product of protein metabolism
o liver converts the ammonia into urea to facilitate its removal from the body
 o measuring ammonia level
 way of determining this particular liver function
o If the liver or kidney for that matter is not efficient in getting rid of this waste,  it accumulates in
the blood and may ultimately cause hepatic coma
 Hepatic coma
 Hepatic encephalopathy
 a nervous system disorder brought on by severe liver disease.
 toxins build up in the blood
 These toxins can travel to the brain and affect brain function
o Method of Ammonia determination
 The basis of ammonia assays come in two approaches
 the diffusion of ammonia from an alkaline medium with trapping in
acid
 use of ion-exchange resin to separate ammonia from the sample
 in enzymatic assays
 ammonia is made to react  with alpha-ketoglutarate and reduced cofactor using
glutamate dehydrogenase to yield L-glutamate and the cofactor.
 The decrease in absorbance is proportional to the concentration of ammonia.
 And in turn its due to the oxidation of reduce cofactor.
 normal range is 15 to 45 µ/dL (11 to 32 µmol/L).

ALT (Alanine Aminotransferase) formerly SGPT 

AST (Aspartate Aminotransferase) formerly SGOT

 these enzymes are responsible for catalyzing the conversion of alanine to pyruvate
 or conversion of aspartate to oxaloacetate by transferring amino groups
 both are found in hepatic cells
 ALT
o mainly found in the liver
o more liver specific than AST
 AST
o more widely distributed in the other tissues
 the levels of these enzymes increased in the serum when hepatic cells are damaged or destroyed.
 the elevations may remain increased from 2-6 weeks
 the degree of elevation of AST and ALT are useful in distinguishing acute chronic liver diseases
 highest levels (500-5000 IU/ml) are found in
o acute stages of viral hepatitis
o drug/toxin related hepatic necrosis and autoimmune hepatitis
 Levels more than 5000 IU/ml can be found in
o acetaminophen associated liver failure
o hepatic ischemia
o herpes simplex hepatitis
 Reference range
o ALT is 7 to 45 U/L
 o AST is 5 to 35 U/L
 AST to ALT ratio can also be very useful such that levels more than 2.0 particularly suggests alcoholic
liver disease
 The deficiency in pyrodixone seen in alcoholic liver disease decreases ALT levels than AST
 ALT/AST  Method of Determination
o determined by measuring the change in absorbance when they transfer their amino groups to
respective ketoacids and become pyruvate and oxaloacetate respectively


 The change in absorbance is proportional to the levels of AST and ALT

Alkaline Phosphatase (ALP)

 alkaline phosphatase being located in the microvilli of the bile canaliculi serves as a good marker of
extra hepatobiliary disorders such as:
 biliary obstruction
 cholestasis
 since it is also found in the placenta and in bones
o physiologically increased in pregnancy and in growing children
 in GIT and kidney disorders
o ALP levels may also increase
 decreased levels =  inherited hyperphosphatasia
 ALP levels vary with age and sex
o due to bone fraction measurements
 Method of ALP Determination
o determined by the Bower's and McComb method based on the hydrolysis of p-nitrophenyl
phosphate to para-nitrophenol

 the corresponding absorbance of p-nitrophenol is directly proportional to the activity of

ALP

Gamma-Glutamyl Transferase (GGT)

 produced in high concentrations in the cell membranes of the liver and gall bladder
 on a lesser degree, it may also be produced in many tissues of the body including:
 intestines
  spleen
 heart
 brain
 seminal vesicles
 clinical utility of GGT is for the evaluation of hepatobiliary and liver diseases
 the first enzyme to increase in bile duct obstruction making, most sensitive liver enzyme in this
condition
 increases are seen in
o chronic alcoholism who take warfarin and phenytoin
 reference range :
o men: 6 to 55 U/L
o women: 5 to 38 U/L
 Method of GGT Determination
o measured when gamma-glutamyl p-nitroanilide transfer its gamma-glutamyl residue to
glycyglycine producing p-nitroaniline

 the absorbance of para-nitroaniline is directly proportional to the GTT

Serum Albumin & Globulin

 together these tests can be used to determine the synthetic function of the liver
 can be useful to assess the extent of liver dysfunction
 when the liver is not functioning well
 there will be decrease in the protein synthesis
 this in turn reflects the decrease in the production of albumin and globulin
 For instance
 decrease in gamma-globulin would suggests alpha 1-antitrypsin deficiency causing the
chronic liver disorder
 however, the corresponding increase in gamma-globulin would implicate chronic active
hepatitis and cirrhosis

Question: From your previous course in CC, what are the methods of albumin and globulin determinations and
what are basis for those tests?

Prothrombin Time PT

 a test used to diagnose bleeding disorders

 in liver diseases:  when it is unable to produce clotting factors
o PT will be increased
 However, it is not routinely used to detect liver problems but rather to assess the risk of bleeding

Summary 
 Liver Function Test
o is a misnomer because some of these test do not actually evaluate the function of the liver but
rather pinpoints the location or source of the problem
 Elevations in
o AST & ALT would implicate hepatocellular damage
o GGT, ALP and Bilirubin would most likely suggests cholestasis
 In essence, the actual liver function would point to its ability to produce clotting factors and plasma
proteins or in its ability to clear the body of toxic waste products as in the case of bilirubin and ammonia
 Elevated liver chemistries :
o do not necessarily mean hepatic pathology
o as some elevations may be transient
o or maybe due to non-hepatic causes