Red blood cell Red blood cell destructiondestruction

Recommended ReadingsRecommended Readings

Wills’ Biochemical basis of Medicine. Wills’ Biochemical basis of Medicine. Chapter 25 Chapter 25

Lippincott’s illustrated reviews biochemistry. Lippincott’s illustrated reviews biochemistry. Chapter 21Chapter 21

Learning objectivesLearning objectives

Know the main location and site of haem breakdownKnow the main location and site of haem breakdown Outline the pathway for haem degradationOutline the pathway for haem degradation Compare/contrast types of Plasma Bilirubin & types of Compare/contrast types of Plasma Bilirubin & types of

HyperbilirubinemiaHyperbilirubinemia Understand the biochemical basis of different types of Understand the biochemical basis of different types of

jaundice.jaundice.

Content Content Main location and site of haem breakdownMain location and site of haem breakdown Pathway for haem degradationPathway for haem degradation

1.1. Formation of bilirubin in reticuloendothelial system Formation of bilirubin in reticuloendothelial system 2.2. Uptake, conjugation & secretion of bilirubin by liverUptake, conjugation & secretion of bilirubin by liver3.3. Catabolism of bilirubin in the gutCatabolism of bilirubin in the gut

Types of Plasma BilirubinTypes of Plasma Bilirubin Definition & types of HyperbilirubinemiaDefinition & types of Hyperbilirubinemia Jaundice Jaundice

Definition &Definition & Types & characteristics of each typeTypes & characteristics of each type Physiological JaundicePhysiological Jaundice Genetic causes of JaundiceGenetic causes of Jaundice

Degradation of hemeDegradation of heme Red blood cells are degraded by the cells of the Red blood cells are degraded by the cells of the

reticuloendothelial system particularly in the liver and reticuloendothelial system particularly in the liver and spleen after 120 days in the circulation (RBC life span)spleen after 120 days in the circulation (RBC life span)

Therefore, The principal sites of heme catabolism are Therefore, The principal sites of heme catabolism are the spleen & the liverthe spleen & the liver

Bilirubin is the end product of heme catabolism:Bilirubin is the end product of heme catabolism: 75% from hemoglobin of old RBCs, 75% from hemoglobin of old RBCs, the rest from hemoglobin of immature RBCs & from the rest from hemoglobin of immature RBCs & from

cytochromes from extra-erythroid tissuescytochromes from extra-erythroid tissues

250-350 mg bilirubin /day is produced in normal adults250-350 mg bilirubin /day is produced in normal adults

Heme catabolism is classified into 3 stages:Heme catabolism is classified into 3 stages:

1.1. Formation of bilirubin in reticuloendothelial Formation of bilirubin in reticuloendothelial

systemsystem (RES)(RES)

2.2. Uptake, conjugation and secretion of Uptake, conjugation and secretion of

bilirubin by the liverbilirubin by the liver

3.3. Catabolism of bilirubin in the gutCatabolism of bilirubin in the gut

Pathway for haem degradationPathway for haem degradation

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1.1. Formation of bilirubin in reticuloendothelial Formation of bilirubin in reticuloendothelial systemsystem (RES):(RES): In reticuloendothelial system (RES), hemoglobin is degraded In reticuloendothelial system (RES), hemoglobin is degraded

into heme and globin.into heme and globin. Globin is either degraded to amino acids or reused for Globin is either degraded to amino acids or reused for

synthesis of hemoglobin.synthesis of hemoglobin. Heme is converted to biliverdin by the microsomal heme Heme is converted to biliverdin by the microsomal heme

oxygenase (a P450 cytochrom) using NADPH and Ooxygenase (a P450 cytochrom) using NADPH and O22 . .

Ferric iron & carbon monoxide are released. Ferric iron & carbon monoxide are released. Biliverdin is water soluble green pigment Biliverdin is water soluble green pigment

Biliverdin is converted to bilirubin by biliverdin reductase. Biliverdin is converted to bilirubin by biliverdin reductase. Bilirubin is water insoluble yellow pigmentBilirubin is water insoluble yellow pigment

Bilirubin is transported in blood to the liver carried by plasma Bilirubin is transported in blood to the liver carried by plasma albumin (it is termed indirect or unconjugated bilirubin).albumin (it is termed indirect or unconjugated bilirubin).

• Certain drugs as salicylates & sulfonamides can displace Certain drugs as salicylates & sulfonamides can displace bilirubin from albumin, allowing bilirubin to enter the CNS bilirubin from albumin, allowing bilirubin to enter the CNS causing neural damage in infantscausing neural damage in infants

2.2. Uptake, conjugation and secretion of bilirubin by Uptake, conjugation and secretion of bilirubin by the liver:the liver:a)a) Uptake of bilirubin:Uptake of bilirubin:

- Bilirubin leaves albumin & is taken by the hepatocytes Bilirubin leaves albumin & is taken by the hepatocytes - Hepatic uptake of bilirubin is mediated by a carrier, & may Hepatic uptake of bilirubin is mediated by a carrier, & may

be competitively inhibited by other organic anionsbe competitively inhibited by other organic anionsb)b) Synthesis of bilirubin-diglucuronide (Conjugation):Synthesis of bilirubin-diglucuronide (Conjugation):

- By UDP glucuronyl transferase enzyme, bilirubin is By UDP glucuronyl transferase enzyme, bilirubin is

conjugated with glucuronic acid, to form bilirubin conjugated with glucuronic acid, to form bilirubin diglucuronide (it is termed direct or conjugated bilirubin diglucuronide (it is termed direct or conjugated bilirubin & it is water soluble) & it is water soluble)

- UDP-glucuronate acts as donor for glucuronate group UDP-glucuronate acts as donor for glucuronate group to form bilirubin monoglucuronide then bilirubin to form bilirubin monoglucuronide then bilirubin diglucuronidediglucuronide

c)c) Conjugated bilirubin is actively secreted by hepatocyte Conjugated bilirubin is actively secreted by hepatocyte into the biliary canaliculiinto the biliary canaliculi (with(with bile) to small intestinebile) to small intestine- Secreation is the rate limiting step in bilirubin metabolismSecreation is the rate limiting step in bilirubin metabolism

3.3.Catabolism of bilirubin in the gut:Catabolism of bilirubin in the gut:

a)a)Conjugated bilirubin is converted by Conjugated bilirubin is converted by bacteria to urobilinogenbacteria to urobilinogen: :

- Conjugated bilirubin is deconjugated and reduced by Conjugated bilirubin is deconjugated and reduced by

intestinal bacteria forming urobilinogen.intestinal bacteria forming urobilinogen.

b)b)Most of urobilinogen is oxidized in colon to a brown pigment, Most of urobilinogen is oxidized in colon to a brown pigment,

urobilin urobilin (stercobilin(stercobilin)), which , which is excreted in fecesis excreted in feces giving feces its giving feces its

color. color.

c)c)Some urobilinogenSome urobilinogen (water soluble) is absorbed from gut into (water soluble) is absorbed from gut into

portal blood and re-excreted by liver in bile (enterohepatic portal blood and re-excreted by liver in bile (enterohepatic

circulation). circulation).

d)d)Traces of this urobilinogen reach the systemic blood & are Traces of this urobilinogen reach the systemic blood & are

excreated by kidneys in urineexcreated by kidneys in urine

Plasma BilirubinPlasma Bilirubin

Normal plasma bilirubin level is <Normal plasma bilirubin level is <1 mg/dL 1 mg/dL (<17 (<17 μμmol/L)mol/L)

Plasma bilirubinPlasma bilirubin is present in 2 forms:is present in 2 forms: Unconjugated bilirubin (Indirect bilirubin)Unconjugated bilirubin (Indirect bilirubin) Conjugated bilirubinConjugated bilirubin (Direct bilirubin)(Direct bilirubin)

Unconjugated bilirubin (Indirect bilirubin)Unconjugated bilirubin (Indirect bilirubin) Main bilirubin present in plasma normallyMain bilirubin present in plasma normally

Water insoluble Water insoluble

Transported in blood as Bilirubin-albumin complexTransported in blood as Bilirubin-albumin complex

Not excreted in urine Not excreted in urine

Increases in blood in cases of:Increases in blood in cases of:

•Hemolysis Hemolysis

•Liver disease: liver fails to uptake or conjugate it.Liver disease: liver fails to uptake or conjugate it. Termed Indirect bilirubin because its estimation by Termed Indirect bilirubin because its estimation by

van den Bergh reaction needs addition of methanol van den Bergh reaction needs addition of methanol to react with the diazo reagent to react with the diazo reagent

Conjugated bilirubin (direct bilirubin)Conjugated bilirubin (direct bilirubin) Bilirubin-diglucuronide which escapes from the liver Bilirubin-diglucuronide which escapes from the liver

to the systemic bloodto the systemic blood

Water-solubleWater-soluble

Excreted in urine: Excreted in urine:

Bilirubinuria is due to conjugated bilirubin and is Bilirubinuria is due to conjugated bilirubin and is

always pathologicalalways pathological

Increases in blood in cases of:Increases in blood in cases of:•Liver disease: liver fails to secrete bilirubin Liver disease: liver fails to secrete bilirubin •Obstruction in the biliary system Obstruction in the biliary system

Termed direct bilirubin because it reacts directly with Termed direct bilirubin because it reacts directly with the diazo reagent of van den Berghthe diazo reagent of van den Bergh

HyperbilirubinemiaHyperbilirubinemia Increased plasma bilirubin level more than Increased plasma bilirubin level more than

1mg/dL 1mg/dL (17 (17 μμmol/L)mol/L)

According to the type of bilirubin increased in According to the type of bilirubin increased in plasma, hyperbilirubinemias are classified into 2 plasma, hyperbilirubinemias are classified into 2 types:types:

Unconjugated hyperbilirubinemia Unconjugated hyperbilirubinemia Conjugated hyperbilirubinemiaConjugated hyperbilirubinemia

According to the underlying defect, According to the underlying defect, hyperbilirubinemias may be: hyperbilirubinemias may be:

Unconjugated , Unconjugated , Conjugated Conjugated BothBoth

Unconjugated hyperbilirubinemiaUnconjugated hyperbilirubinemia High level of unconjugated bilirubin in blood.High level of unconjugated bilirubin in blood. Unconjugated bilirubin can cross the blood-brain Unconjugated bilirubin can cross the blood-brain

barrier & cause severe brain damage (kernicterus)barrier & cause severe brain damage (kernicterus) Unconjugated bilirubin is Unconjugated bilirubin is notnot excreted in urine excreted in urine Causes:Causes:

HemolysisHemolysisImmaturity of the enzyme of bilirubin conjugation in Immaturity of the enzyme of bilirubin conjugation in

neonates (physiological)neonates (physiological)Genetic defect in the enzyme of bilirubin conjugationGenetic defect in the enzyme of bilirubin conjugation

Gilbert's syndromeGilbert's syndrome

Crigler-Najjar syndromeCrigler-Najjar syndromeMost common causes are Hemolysis & Gilbert's Most common causes are Hemolysis & Gilbert's

syndromesyndrome

Conjugated hyperbilirubinemiaConjugated hyperbilirubinemia High level of conjugated bilirubin in blood.High level of conjugated bilirubin in blood.

Due to leakage of conjugated bilirubin from hepatocytes or Due to leakage of conjugated bilirubin from hepatocytes or

biliary system into blood biliary system into blood

Conjugated bilirubin is water soluble, so it is excreted in Conjugated bilirubin is water soluble, so it is excreted in

urine and darkens urine color to urine and darkens urine color to deep orange brown deep orange brown

Causes:Causes:Biliarry obstruction Biliarry obstruction Decreased hepatic secretion of conjugated bilirubin Decreased hepatic secretion of conjugated bilirubin

Dubin-Johnson SyndromeDubin-Johnson SyndromeUnknown causeUnknown cause

Rotor’s Syndrome.Rotor’s Syndrome. Most common cause is biliary obstructionMost common cause is biliary obstruction

JaundiceJaundice

Jaundice is clinically seen as yellow color of skin, Jaundice is clinically seen as yellow color of skin,

nail beds & sclera (due to deposition of bilirubin) nail beds & sclera (due to deposition of bilirubin)

when plasma bilirubin concentration exceeds when plasma bilirubin concentration exceeds

3 mg/dL (50 μmol/L) due to imbalance 3 mg/dL (50 μmol/L) due to imbalance

between bilirubin production and excretion.between bilirubin production and excretion.

Types of jaundice are: Types of jaundice are: Prehepatic (hemolytic) Prehepatic (hemolytic) HepaticHepatic Posthepatic (obstructive) Posthepatic (obstructive)

Prehepatic (hemolytic) Prehepatic (hemolytic)

jaundice:jaundice: Due to excess production of Due to excess production of

unconjugated bilirubin after unconjugated bilirubin after hemolysis which exceeds the hemolysis which exceeds the capacity of liver to conjugate it capacity of liver to conjugate it

Characterized by the following:Characterized by the following: High levels of indirect High levels of indirect

(unconjugated) bilirubin in plasma (unconjugated) bilirubin in plasma i.e., Unconjugated i.e., Unconjugated hyperbilirubinemia hyperbilirubinemia

Dark urine caused by high levels Dark urine caused by high levels of urobilinogen in urine of urobilinogen in urine

Dark stool caused by high levels Dark stool caused by high levels of Fecal urobilinof Fecal urobilin

Posthepatic (obstructive) Posthepatic (obstructive)

jaundice jaundice

Due to biliary obstruction Due to biliary obstruction

Characterized by the following:Characterized by the following:

• High levels of direct (conjugated) High levels of direct (conjugated)

bilirubin in plasma i.e., conjugated bilirubin in plasma i.e., conjugated

hyperbilirubinemia hyperbilirubinemia

• Dark urine due to presence of Dark urine due to presence of

conjugated bilirubin. conjugated bilirubin.

• Urine urobilinogen is absent Urine urobilinogen is absent

• Very pale stool (white, clay) due to Very pale stool (white, clay) due to

absence of Fecal urobilinabsence of Fecal urobilin

Hepatic jaundice: Hepatic jaundice: Due to hepatocyte dysfunction that Due to hepatocyte dysfunction that

cause impaired hepatic bilirubin cause impaired hepatic bilirubin uptake, conjugation, or secretion: uptake, conjugation, or secretion: • impaired uptake → increased unconjugated impaired uptake → increased unconjugated bilirubin in blood), bilirubin in blood),

• impaired conjugation → decreased impaired conjugation → decreased conjugated bilirubin formation in liver→ conjugated bilirubin formation in liver→ decreased urobilinogen formation in gut) decreased urobilinogen formation in gut)

• impaired secretion→increased conjugated impaired secretion→increased conjugated bilirubin in blood & secretion in urine)bilirubin in blood & secretion in urine)

Characterized by the following: Characterized by the following: •High levels of direct (conjugated) & High levels of direct (conjugated) & indirect (unconjugated) bilirubin in indirect (unconjugated) bilirubin in plasma i.e., conjugated & plasma i.e., conjugated & Unconjugated hyperbilirubinemiaUnconjugated hyperbilirubinemia

•Dark urine due to the presence of Dark urine due to the presence of conjugated bilirubin & urobilinogen conjugated bilirubin & urobilinogen

•Pale stool due to decreased Fecal Pale stool due to decreased Fecal urobilinurobilin

increased

Neonatal (physiologic) jaundice:Neonatal (physiologic) jaundice: Transient jaundice, common in neonates (50% of normal Transient jaundice, common in neonates (50% of normal

babies) particularly in premature infants babies) particularly in premature infants Due to immaturity of the enzymes of bilirubin conjugation Due to immaturity of the enzymes of bilirubin conjugation

leading to unconjugated hyperbilirubinemialeading to unconjugated hyperbilirubinemia Unconjugated bilirubin is toxic to the immature brain, it may Unconjugated bilirubin is toxic to the immature brain, it may

cause kernicterus cause kernicterus if it exceeds 20-25 mg/dl, if it exceeds 20-25 mg/dl, kernicterus may result in mental retardation.kernicterus may result in mental retardation. Treatment:Treatment:

to avoid kernicterus, if plasma bilirubin is too to avoid kernicterus, if plasma bilirubin is too high:high:

• Phenobarbital: inducer for UDP-glucuronyl transferase Phenobarbital: inducer for UDP-glucuronyl transferase

• Phototherapy: converts bilirubin to more soluble derivatives Phototherapy: converts bilirubin to more soluble derivatives that are easily excreted in bile (detoxifies bilirubin) that are easily excreted in bile (detoxifies bilirubin)

• Exchange blood transfusion to remove excess bilirubinExchange blood transfusion to remove excess bilirubin

Neonatal jaundice phototherapy

Genetic causes of jaundiceGenetic causes of jaundice::

1.1. Gilbert's syndromeGilbert's syndrome

2.2. Crigler-Najjar syndromeCrigler-Najjar syndrome

3.3. Dubin-Johnson syndromeDubin-Johnson syndrome

4.4. Rotor's syndromeRotor's syndrome

Gilbert's syndromeGilbert's syndrome:: Affect up to 5% of population Affect up to 5% of population

Due to decreased conjugation of bilirubin & decreased Due to decreased conjugation of bilirubin & decreased

uptake in some cases uptake in some cases

Decreased conjugation of bilirubin is due to moderate Decreased conjugation of bilirubin is due to moderate

deficiency in UDP glucuronyl transferase activity deficiency in UDP glucuronyl transferase activity

Mild intermittent Mild intermittent unconjugated hyperbilirubinemiaunconjugated hyperbilirubinemia which is which is

noticed after fasting or infection noticed after fasting or infection

Harmless & asymptomatic → normal lifespanHarmless & asymptomatic → normal lifespan

Crigler-Najjar syndrome:Crigler-Najjar syndrome: Type 1: Type 1:

• Severe Severe unconjugated hyperbilirubinemiaunconjugated hyperbilirubinemia at birth at birth• Due to absence of conjugating enzymesDue to absence of conjugating enzymes

• Fatal due to kernicterusFatal due to kernicterus• Partial response to phototherapy, non to Phenobarbital Partial response to phototherapy, non to Phenobarbital

Type 2:Type 2:• Severe Severe unconjugated hyperbilirubinemiaunconjugated hyperbilirubinemia at birth at birth • Due to partial defect of conjugating enzymes Due to partial defect of conjugating enzymes • Survive to adulthoodSurvive to adulthood• Good response to phototherapy & PhenobarbitalGood response to phototherapy & Phenobarbital

Dubin-Johnson syndrome:Dubin-Johnson syndrome: Due to decreased hepatic secretion of conjugated bilirubin Due to decreased hepatic secretion of conjugated bilirubin

Mild intermittent Mild intermittent conjugated hyperbilirubinemiaconjugated hyperbilirubinemiaBilirubinuria Bilirubinuria Normal lifespanNormal lifespan

Rotor's syndrome:Rotor's syndrome: Cause is unknownCause is unknown

Mild intermittent Mild intermittent conjugated hyperbilirubinemiaconjugated hyperbilirubinemiaBilirubinuriaBilirubinuriaNormal lifespanNormal lifespan