JAUNDICE

Question # 1

Discuss the
catabolism of heme.

Hemoglobin Catabolism:




Hemoglobin
Globin + Heme
Iron + Porphyrin
Bilirubin
REC, of liver, spleen, BM cells
1. Formation of bilirubin:

STEPS:
Catalyzed by microsomal heme oxygenase system of
reticuloendothelial cells.
In the presence of NADPH and oxygen, the enzyme adds a
hydroxyl group to he methenyl bridge between the two
pyrrole rings with a concomitant oxidation f ferrous iron to
ferric state.
A second oxidation of the same enzyme cleaves he
porphyrin ring.
Ferric iron and CO
2
are released.
Biliverdin is produced.
Bilirubin reductase reduces biliverdin to bilirubin.

2. Uptake of bilirubin by the liver:

Bilirubin is non-covalently bound to
albumin while being transported to
the liver.

Bilirubin dissociates from albumin
and enters a hepatocyte where it
binds to ligandin (cytoplasmic anion
binding protein).


3. Conjugation of bilirubin

diglucuronide / conjugation
Enzyme: bilirubin glucuronyl
transferase

Glucuronate donor: UDP-
glucuronic acid.


4. Excretion of bilirubin into bile:

Conjugated bilirubin is actively transported against a
concentration gradient into the bile canaliculi and then into
the bile.

5. Formation of urobilins in the intestines:

Bilirubin diglucuronide is hydrolyzed and reduced by
bacteria in the gut to urobilinogen.

Urobilinogen can be reabsorbed in the gut and transported
to the kidneys to form urobilin (yellow color of urine).

Most of the urobilinogens are reabsorbed to the
enterohepatic pathway.
Question # 2

Differentiate conjugated
bilirubin from
unconjugated bilirubin.
Unconjugated bilirubin Conjugated Bilirubin
Present normally in plasma Present normally in bile
Attached non covalently to
albumin
Conjugated to glucuronic acid
Has high molecular weight and
cannot be filtered through the
kidney
Has small molecular weight and
if present in plasma can be
filtered through the kidney.
Non-polar, insoluble in plasma
and can cross blood brain barrier
in neonates
Polar, soluble in plasma and
cannot cross blood brain barrier
Gives indirect Van den Bergh
reaction
Gives direct Van den Bergh
reaction
Van den Bergh Reaction:

This is a reaction between bilirubin and
Ehrlich diazo reagent giving a reddish
purple compound.

Conjugated bilirubin reacts directly to the
reagent. Thus, it is called direct bilirubin.

Unconjugated bilirubin does not directly
react with the compound except after the
addition of methyl alcohol. It may be called
indirect bilirubin.


Question #3:

Discuss the different phases of
bilirubin metabolism. What are the
roles of proteins (albumins, ligandins,
etc.) in bilirubin metabolism?

TWO PHASES OF BILIRUBIN METABOLISM:
A. HEPATIC PHASE

1. Hepatic Uptake of Bilirubin
2. Bilirubin Conjugation
3. Biliary Excretion

B. INTESTINAL PHASE
A. HEPATIC PHASE
Liver has a central role

Divided into 3 distincts processes:
1. Hepatic Uptake
2. Bilirubin Conjugaction
3. Biliary Excretion
A. HEPATIC PHASE: Hepatic Uptake
Bilirubin is only sparingly soluble in water,
but its solubility in plasma is increased by
noncovalent binding to albumin.
Each molecule of albumin appears to have
one ____ for bilirubin:
High-affinity site bilirubin, drugs (antibiotics)
Low-affinity site (loose) - bilirubin
In 100 mL of plasma, approximately 25 mg of
bilirubin can be tightly bound to albumin at its
high-affinity site.

A. HEPATIC PHASE: Hepatic Uptake
Things involved in this process:

a. OATP2 a family of organic anion-transporting
polypeptidase (OATPs)

b. Proteins (OATPs)
Ligandin (glutathione S-transferase B)
Protein Y and Protein Z

A. HEPATIC PHASE: Hepatic Uptake
Steps:
1. B1-Albumin complex dissociates
2. B1 (nonpolar) enters the hepatocyte by diffusion via
OATP2; and albumin goes back to circulation for
more binding of B1
3. In the cytosol, B1 binds to proteins like Ligandin and
Protein Y& Z for transport to ER for conjugation.


A. HEPATIC PHASE: Hepatic Uptake
A. HEPATIC PHASE: Bilirubin Conjugation

Occurs in smooth endoplasmic reticulum

Things involved in this process:
o 2 molecules of Uridine Diphosphate - Glucuronyl
Transferase (UDP-GT)
o UDP- Glucuronic acid (bilirubin-UGT) glucuronate
donor

A. HEPATIC PHASE: Bilirubin Conjugation


A. HEPATIC PHASE: Biliary Excretion

Things involved in this process:
Energy (ATP)
MRP2 (multidrug resistance-like protein 2) or
cMOAT (canalicular multispecific organic anion
transporter) located in plasma membrane of the
biliary canaliculi; a member of the family of ATP-
binding cassette (ABC) transporters; for biliary
excretion of B1


A. HEPATIC PHASE: Biliary Excretion
A. HEPATIC PHASE: Summary
Determination of Bilirubin Concentration
A. Van den Bergh Reaction / Diazo Reaction

Reaction: Diazotized sulfanilic acid ------ red
+ Bilirubin azodipyrroles

Coupling Accelerator:
Methanol provides a solution in which both B1 and diazo
reagent are soluble


Determination of Bilirubin Concentration
B. High Performance Liquid Chromatography

Most accurate

Most sensitive

Quantitative method

B. INTESTINAL PHASE
Conjugated Bilirubin (B2) in small amount
Biliary duct
Duodenum
and
Colon
Enzymatic Deconjugation
(ex. bacterial B-glucuronidase)
Nonenzymatic Deconjugation
(ex. alkaline hydrolysis)
Unconjugated Bilirubin
B. INTESTINAL PHASE
UROBILINOGEN
(Soluble)
Kidney
Liver
Intestine
STERCOBILINOGEN STERCOBILIN
bacteria
O
2
4 mg
Question #4:

What are the causes of jaundice?

JAUNDICE
Yellowish discoloration of the skin and sclera
Newborn: there is a decreased level of hepatic
bilirubin glucuronyl tranferase.
It is also known as icterus
a.skin
b.conjunctival membrane over the sclera
(scleral icterus but more properly called
conjunctival icterus)
c.mucous membranes
Jaundice itself is not a disease


JAUNDICE
Types of Jaundice:
1. Hemolytic Jaundice
2. Obstructive or Cholestatic Jaundice
3. Hepatocellular Jaundice

Causes of JAUNDICE
HEMOLYSIS
LIVER DAMAGE
BILE DUCT
OBSTRUCTION
JAUNDICE
Increased Production of Bilirubin
Decreased Excretion of Bilirubin
JAUNDICE: 3 Categories
PRE-HEPATIC HEPATIC POST-HEPATIC
Pathologic
location
Prior to liver Within the liver After conjugation in
the liver
Mechanism Too much
(massive)
destruction of
RBC
Damage to liver cells (
caused by viruses, alcohol,
and parasites); impaired
cellular uptake, defective
conjugation, abnormal
secretion of bilirubin by the
hepatocytes
Obstruction of bile
duct (failure of bile to
flow to the intestine /
impaired bilirubin
excretion)
Comparison
a. B1 Increased Increased Normal
b. B2 Normal Increased Increased
c. Urobilinogen Normal Decreased Decreased
d. Urine Bilirubin Negative Positive Positive
e. Stool Color Normal Normal Pale
f. ALP / ALT / AST Normal Increase Increased
PRE-HEPATIC
HEPATIC
POST-HEPATIC
Location ?
Mechanism?
B1 ?
B2 ?
Urobilinogen ?
Urine Bilirubin ?
Stool Color ?
ALP ?
AST ?
ALT ?
Question # 5
Differentiate between the different
types of jaundice (conjugated vs
unconjugated hyperbilirubinemia);
(choluric vs acholuric jaundice)
Unconjugated
Hyperbilirubinemia
I. Overproduction
Hemolysis (intra- and extravascular)
Ineffective Errythropoiesis
II. Decreased hepatic uptake
prolonged fasting
sepsis
III. Decreased bilirubin conjugation
(decreased hepatic
glucuronosyl transferase
activity)
A. Hereditary transferase deficiency
Gilberts syndrome (mild transferase
deficiency)
Crigler-Najjar type II (moderate
transferase deficiency)
Crigler- Najjar type I (absence of
transferase)
B. Neonatal jaundice (transient
transferase deficiency; increased
intestinal absorption of unconjugated
bilirubin)
Drug inhibition (e.g., chlorampenicol,
pregnanediol)
Breast milk jaundice (transferase inhibition by
pregnanediol and fatty acids in breast milk)
hepatocellular disease (hepatitis, cirrhosis)
C. Acquired transferase deficiencyof
unconjugated bilirubin)
Conjugated
Hyperbilirubinemia
I. Impaired hepatic excretion
(intrahepatic defects)
A. Familial or hereditary disorders
Dubin-Johnson syndrome
Rotor syndrome
Recurrent (benign)
intrahepatic cholestasis
Cholestatic jaundice of
pregnancy
B. Acquired disorders

Hepatocellular disease (e.g., viral or drug-
induced hepatitis, cirrhosis)
Drug-induced cholestasis (e.g., oral
contraceptives)
Alcoholic liver disease
Sepsis
Post operative state
Parenteral nutrition
Biliary cirrhosis (primary or secondary)
II. Extrahepatic biliary
obstruction
A. Intraductal obstruction
Gallstones
Biliary malformation (e.g., stricture,
atresia, choledochol cyst)
Infection (e.g., Clonorchis, Ascaris)
Malignancy (cholangiocarcinoma,
ampullary carcinoma)
Hemobilia (trauma, tumor)
Sclerosing cholangitis
B. Compression of biliary ducts
Malignancy (e.g., pancreatic carcinoma,
lymphoma, metastases to portal lymph
nodes
Inflammation (e.g., pancreatitis)
Choluric Jaundice
Conjugated bilirubin is water soluble
therefore can be excreted in urine
To which imparts a yellow-brown
coloration
This is called choluric jaundice
Acholuric jaundice
Hemolytic jaundice
Jaundice without bilirubinemia
Associated with elevated unconjugated
bilirubin that is nnot excreted by the kidney

Question # 6 Explain the
occurrence of jaundice in the
following conditions:
Physiologic jaundice of the newborn

Decreased hepatic bilirubin glucuronyl
transferase at birth
Immature liver
Increase in unconjugated bilirubin
Toxic encephalopathy/kernicterus
Treated with blue flourescent light
Hemolytic jaundice

Important cause of unconjugated
hyperbilirubinemia
High levels of hemoglobin released from
erythrocytes due to hemolysis
Rate of bilirubin production exceeds the rate of the
liver clearance





Defects in conjugation

Toxic hyperbilirubinemia
Toxin induced liver dysfunction
(chloroform, arsphenamines, carbon
tetrachloride, acetaminophen, hepatitis virus,
cirrhosis and Ammantia mushroom poisoning)
Due to hepatic parenchymal cell damage (which impairs
conjugation)
Dubin Johnson syndrome
Rotor syndrome

Breast milk jaundice

Begins 1
st
week after birth
Peaks within two weeks after birth
Infants immature liver and intestines
Milk of some mothers has non-esterified long chain
fatty acids or other substances which inhibit
glucuronyl transferases conjugation activity
Nursing may be discontinued until bilirubin levels fall
rapidly, then nursing may be resumed.

Obstruction of the biliary tract

most common cause of conjugated hyperbilirubinemia
blockage of the hepatic or common bile ducts
gallstone or cancer of the head of the pancreas
bilirubin diglucuronide cannot be excreted
regurgitates into the hepatic veins and lymphatics
conjugated bilirubin appears I the blood and urine
Phototherapy works through a
process of isomerization that
changes trans-bilirubin into the
water-soluble cis-bilirubin isomer.
TREATMENT
Phenobarbital was used to treat neonatal
jaundice by increasing liver metabolism
and thus lowering bilirubin levels.