3. PATHOPHYSIOLOGICAL
IMPORTANCE OF BILIRUBIN
METABOLISM
It is the end product of heme degradation.
Serum bilirubin level is an important clinical
marker of hepatobiliary excretory function.
Bilirubin is an endogenous model for plasma
carriage and hepatic throughput of organic
anions.
Hepatic uptake, storage, conjugation and
excretion of bilirubin are finely balanced.
Therefore, enhancement of bilirubin throughput
requires coordinated induction of multiple genes,
which may be mediated by nuclear receptors.
5. Early bilirubin
(15-20%)
0- 3 days
Late bilirubin
(65%)
40- 80 days
Non-Hb
sources (liver)
Increased
erythropoiesis
Erythrocyte
sources
Early and late labeled peaks of radioisotope incorporation into bilirubin
After injection of labeled porphyrin precursor (14
C-glycine)
6. Opening of the heme ring and
Enzyme-catalyzed formation of bilirubin
7. Bilirubin throughput: schema of a hepatocyteBilirubin throughput: schema of a hepatocyte
Sinusoidal
surface
Canalicular
surface
Tight
junction
Liver
sinusoid
Fenestrated
endothelium
8. Bilirubin circulates bound to serum albumin.
BBalb
Albumin-
binding:
Keeps bilirubin
soluble
Prevents
tissue deposi-
tion.
Prevents
renal excretion
Drugs that
displace
bilirubin from
albumin may
precipitate
kernicterus:
Sulfonamides
Coumadin, etc.
9. Bilirubin circulates bound to serum albumin.
At the sinusoidal surface of hepatocytes, it dissociates
from albumin.
BBalb
10. Bilirubin circulates bound to serum albumin.
At the sinusoidal surface of hepatocytes, it dissociates
from albumin.
BBalb
11. Bilirubin circulates bound to serum albumin.
At the sinusoidal surface of hepatocytes, it dissociates
from albumin.
BB
alb
12. Bilirubin circulates bound to serum albumin.
At the sinusoidal surface of hepatocytes, it dissociates
from albumin.
BB
alb
13. Bilirubin enters through the sinusoidal surface, probably by
facilitated diffusion.
Uptake is energy independent and bidirectional.
BB
Bilirubin uptake
is reduced:
In neonates
In cirrhosis
From drug
effect:
novobiocin
In some cases
of Gilbert
syndrome
14. B
Inside the hepatocyte, bilirubin binds to cytosolic proteins
termed ligandins, which are the same as glutathione-S-
transferases (GSTs).
GSTs
B
GST binding
inhibits the
efflux of bilirubin,
thereby increasing
its net uptake
16. B
Conjugation of bilirubin with glucuronic acid is catalyzed
by UGT1A1, which transfers glucuronic acid from
UDP-glucuronic acid to bilirubin
GSTs
UDPUDPGAGA UDPUDP
BB GAGA
UGT1A1
B
Conjugation with
glucuronic acid
makes bilirubin
water-soluble and
non-toxic.
Glucuronidation
is essential for
biliary excretion
of bilirubin.
17. UDP-glucuronosyltransferasesUDP-glucuronosyltransferases
(UGTs)(UGTs)
• UGTs are ER proteins that convert many internal andUGTs are ER proteins that convert many internal and
exogenous toxins to non-toxic metabolites.exogenous toxins to non-toxic metabolites.
• UGT’s are a family of enzymes concentrated in the liver.UGT’s are a family of enzymes concentrated in the liver.
• One UGT isoform, UGT1A1, conjugates bilirubin and isOne UGT isoform, UGT1A1, conjugates bilirubin and is
essential for its excretion.essential for its excretion.
• Inherited UGT1A1 deficiency causes jaundice.Inherited UGT1A1 deficiency causes jaundice.
Substrate
UDPGAUDPGA
•UGTUGT
GlucuronideGlucuronide
UDPUDP
18. Inherited disorders of bilirubin metabolism causingInherited disorders of bilirubin metabolism causing
Unconjugated HyperbilirubinemiaUnconjugated Hyperbilirubinemia
• Crigler-Najjar syndromeCrigler-Najjar syndrome
type 1:type 1:
• Crigler-Najjar syndromeCrigler-Najjar syndrome
type 2:type 2:
• Gilbert syndrome:Gilbert syndrome:
Virtually no UGT1A1 activityVirtually no UGT1A1 activity
UGT1A1 activity below 10%UGT1A1 activity below 10%
UGT1A1 activity ~30%UGT1A1 activity ~30%
19. NEONATAL HYPERBILIRUBINEMIA
• Clinical jaundice appears in newborns when the bilirubin levels is more than
7 mg/dl
• 85% of term newborns and most of the premature infants develop clinical
jaundice
• 6% have bilirubin more than 12.9mg/dl and 35 have bilirubin more than 15
mg/dl.
20. PHYSIOLOGICAL
HYPERBILIRUBINEMIA
• Serum UCB rises to >2mg/dl in first week of life
• Peaks to 6- 8 mg/dl by 3 to 5 days of age
• This normal jaundice is attributed too following conditions:
Increased rbc volume per kg
Increased ineffective erythropoiesis and increased turnover of non heme
proteins
Increased enterohepatic circulation
Decreased uptake of bilirubin from plasma caused by decreased ligandin
Defective conjugation due to decreased UGT activity
Decreased hepatic excretion of bilirubin
21. NON PHYSIOLOGICAL
HYPERBILIRUBINEMIA
• Onset of jaundice is before 24 hours
• Elevation of serum bilirubin that requires phototherapy
• A rise in serum bilirubin levels of >0.2 mg/dl/hour
• Signs of underlying diseases in an infant(vomiting, lethargy, poor feeding,
excessive weight loss
• Jaundice persisting after 8 days in a term infant or after 14 days in a
premature infant.
22. POINTS TO BE NOTED IN HISTORY..
1. A family history of jaundice , anemia, splenectomy or early gallbladder
disease suggests hereditary hemolytic anemia (spherocytosis,
G6PD deficiency)
2. A family history of liver disease may suggest galactosemia, alpha-1 anti
trypsin deficiency, tyrosinosis, hypermethionemia, gilberts syndrome,
criggler najjar syndrome type1 and 2 and cystic fibrosis
3. Sibling with jaundice or anemia may suggest blood group incompatibility
4. Maternal illness during pregnancy may indicate congenital viral infection /
toxoplasmosis
23. • Maternal drugs may interfere with bilirubin metabolism like sulfonamides or
may cause hemolysis in g6pd deficient infants.
• Labour and delivery history may show trauma associated with extravascular
bleeding and hemolysis. Oxytocin use in labour may be associated with
neonatal hyperbilirubinemia
• Infants with HIE , delayed cord clamping.
• Delayed / infrequent stooling
24. BREAST FEEDING JAUNDICE BREAST MILK JAUNDICE
Higher bilirubin levels in infants who are
exclusively breast fed after day 3 of life
Peak level of bilirubin is >12 mg/dl in breast fed
infants
Main factor responsible for breast feeding
jaundice is decreased in take of milk that leads
to slower eliminationof bilirubin and increased
entero hepatic circulation
Late onset of more than 4 days after life
Bilirubin continues to rise and may reach upto
20 to 30 mg/dl by 14 days.
Mechanism thought to be due to unidentified
factors in the milk interfering with the bilirubin
metabolismand due to increased beta
glucorinidase in breast milk causing increased
enterohepatic circulation.
25. POINTS TO BE NOTED IN THE
PHYSICAL EXAMINATION• Prematurity
• SGA ----------- polycythemia and in-utero infection
• Microcephaly ---------- in-utero infection
• Extra vascular blood bruising , cephalhematoma
• Pallor ---------------- hemolytic anemia or extra vascular blood loss
• Petechiae associated withcongenital infection, sepsis or erythroblastosis
• HSM --------- hemolytic anemia, congenital infection/ liver disease
• Omphalitis
• Chorioretinitis ------------- congenital infection
• Evidence of hypothyroidism
26.
27.
28.
29.
30. METHODS OF BILIRUBIN
ESTIMATION
• Visual inspection is not a reliable method of bilirubin estimation
• Screening of total serum bilirubin (TSB) collected predischarge from the baby
and plotted on an hour specific normogram helps to identify the infants at
risk.
• Trans cutaneous bilirubin measurement using multiple wavelength analysis
can reliably estimate serum bilirubin levels
31. TRANSCUTANEOUS BILIRUBIN ESTIMATION
ADVANTAGES DISADVANTAGES
• Reliably estimates irrespective of skin
pigmentation post natal age and weight of
infant
• Reduce the number of invasive blood tests
performed
• Reduce the health care costs
• It’s a screening tool and needs to confirm
the value with total serum bilirubin levels
when
1. TcB is > 70 th percentile for the
phototherapy value
2. When TcB exceeds more than 75th
percentile in bhutani’s normogram
3. At follow up if TcB is more than 13 mg/dl.
• TcB monitoring is unreliable after
phototherapy has begun due to bleaching
of skin.
32. • End tidal corbon monoxide (ETCOc)
o Offers insight into the underlying pathological process contributing to
hyperbilirubinemia
o Only parameter that provide direct information on the rate of bilirubin
production
o Can differentiate between infants with increased production, decreased
elimination or decreased conjugation if the test is used in combination with total
serum bilirubin levels
33. OTHER CLINICAL TESTS TO BE
DONE..
1. Blood type , Rh and antibody screen of the mother
2. Blood test, Rh and antibody screen if the infant
• Routinely recommended if the mother is Rh negative
• Peripheral smear for RBC morphology and reticulocyte count
• Hematocrit
• Identification of antibody on infants RBC if result of direct Coomb’s test is
positive
• Direct bilirubin
• A G6PD screen in male infants
34. • In cases of prolonged jaundice
Measure the direct component
Test for liver disease
Congenital infection
Sepsis, metabolic defects
Hypothyroidism
TPN is an important cause for prolonged hyperbilirubinemia
35. BILIRUBIN TOXICITY
• Bilirubin enters the brain as free unbound bilirubin or bilirubin bind to albumin
in case of disrupted blood-brain barrier.
• FFAs and some drugs interfere with the binding of bilirubin to albumin
• The blood brain barrier is disrupted in cases of hyperosmolarity, asphyxia and
hypercarbia.
• More permeable in premature infants.
36. KERNICTERUS
• Pathological diagnosis
• Yellow staining of brain with bilirubin and evidence of neuronal injury
• Most commonly in basal ganglia, various cranial nuclei, brainstem nuclei,
cerebellar nuclei, hippocampus, anterior horn cells of spinal cord.
• Microscopically there is necrosis, neuronal loss and gliois.
• Clinically characterized by athetosis, auditory neuropathy, upward gaze
limitation, dental dysplasia and intellectual defects.
37. ACUTE BILIRUBIN
ENCEPHALOPATHY
3 phases :
1.Early phase : hypotonia, lethargy, high-pithced cry and poor suck
2.Intermediate phase : hypertonia of extensor muscles, irritability, fever and
seizures. Mostly fatal. If infants survive this phase then chronic bilirubin
encephalopathy (clinical diagnosis of kernicterus is made).
3.Advanced phase : pronounced ophisthotonus, shrill cry, apnoea, seozures,
coma and death.
38. MANAGEMENT OF UNCONJUGATED
HYPERBILIRUBINEMIA
• GENERAL PRINCIPLES
• PREVENTIVE MEASURES AND EARLY IDENTIFICATION OF CAUSE IMPORTANT
• ANY MEDICATION THAT INTERFERE WITH BILIRUBIN METABOLISM/ INTEGRITY OF
BLOOD-BRAIN BARRIER SHOULD BE DISCONTINUED
• CORRECT INADEQUATE FEEDING OF INFANTS AND EVACUATION OF MECONIUM
• ASPIRATION OF CEPHALHEMATOMA
• TREATMENT OF SEPSIS AND HEPATITIS
• CORREECT HYPOTHYROIDISM IF PRESENT
39. • ROLE OF PHENOBARBITONE:
• Barbiturates shown to improve the maturation of microsomal enzymes,
ligandin and gludoronyl transferace thus improving the uptake conjugation
and maturation of bilirubin
• Lag period of 48 to 72 hours exists before enzyme activity is induced by
phenobarbitone so is administered within 72 hours of life even administered
prenatally
• Indicated in infants with
i. Cord bili >2.5 mg/dl
ii. Early onset jaundice due to any cause
iii. Difficult or instrumental / oxytocin induced delivery
iv. G6PD deficiency and type ii crigler najjar syndrome
• Added benefit of phenobarbitone is that it protects fetal brain against
hypoxic damage and may reduce the incidence of intraventricular
hemorrhage.
40. • Clofibrate is another drug which is a potent enhancer of glucoronyl
transferace than phenobarbital .
• 100% increase in hepatic metabiolism in one study in one week
• But it is slow in action and takes several days for its action .
42. PHOTOTHERAPY
• Relatively safe and effective method for treatment of neonatal
hyperbilirubinemia
• Bilirubin absorbs light maximally at 425- 475nm
• 3 photochemical reactions occur when bilirubin is exposed to light:
A. Structural isomerization
B. Photoisomerization
C. Photo- oxidation
43. • STRUCTURAL ISOMERIZATION
• Intramolecular cyclization of bilirubin to lumirubin.
• Lumirubin is rapidly excreted in bile and urine without conjugation
• The conversion is irreversible
• Most important pathway in the lowering of serum bilirubin
• Strongly related to the dose of phototherapy used in the range of 6 to 12 micro
watts/cm^2/ nm
44. N
H
M
V
O
N
H
M CH2
OH
CH2
N
H
N
H
CH2
CH2
C
OOH M M
V
O
CH2
C
O
Internal hydrogen bonds are disrupted
transiently upon exposure of bilirubin to light.
C
C
The dipyrrole carbon bridges switch direction.
49. Phototherapy changes the
configuration of bilirubin making
it transiently water soluble
• Photo isomerization
• Occurs in extravascular spaces of skin
• The natural isomer of UCB is converted to less toxic polar isomer and it diffuses
into blood and is excreted in to bile without conjugation
• But it’s a reversible reaction and the UCB thus formed again will be reabsorbed
from the gut
• Not the major pathway of bilirubin is excretion after starting phototherapy
• Photo – oxidation
• Converts bilirubin to small polar compounds that are excreted in urine
• Least important reaction for lowering of bilirubin
50. INDICATIONS OF PHOTOTHERAPY
• Levels of bilirubin are hazardous to the infant although it has not reached
the levels requiring exchange transfusion
51. • Prophylactic phototherapy is indicated in
• ELBW infant and Severely bruised infants where bilirubin is anticipated to
increase rapidly
• In hemolytic disease of newborn when rise in serum bilirubin is plotted
• phototherapy is contraindicated in
• Infants with direct hyperbilirubinemia caused by liver disease or obstructive
jaundice, and phototherapy may lead to bronze baby syndrome
52. TECHNIQUE OF PHOTOTHERAPY
• Effective phototherapy depends on :
• Light spectrum
• Irradiance
• Distance from the infant
• Extent of skin exposure
• Conventional phototherapy delivers spectral irradiance at infants level of 8
to 10 micro watts/cm^2/nm, 430 to 490 nm when positioned at 20cm above
the infant
• Intensive phototherapy delivers atleast 30micro watt/cm^2/nm
53. • Methods to increase the efficacy of phototherapy:
• Reducing the efficacy of phototherapy unit and infant to 15 to 20 cm
• Using compact fluorescent tubes
• Providing double surface phototherapy
• Using slings/ cloth of white colout to reflect light on to the baby
• Using high intensity gallium nitride light emitting diodes
54. PRECAUTIONS TO BE TAKEN IN
PHOTOTHERAPY
• Eye patches and genital shield to be ensured
• Infants are turned every 2 hours
• Eye patches should not occlude the nares as asphyxia and apnoea can
result
• Infant temperature should be properly maintained and servo controlled
• Infants should be weighed daily
• Skin colour is NOT a guide to bilirubin level so bilirubin has to repeated every
10 to 12 hours
55. SIDE EFFECTS OF PHOTOTHERAPY
1. Insensible water loss – especially those under radiant warmers. Term infants
40% and pre term infants 80% - 190%, servo controlled warmers decrease
this water loss
2. Redistributuin of blood flow -
• in term infants the cerebral and pulmonary velocity increases and the left
ventricular and renal velocity decreases. Returns to normal after discontinuation
of phototherapy
• In preterm infants the cerebral blood flow velocity increases and the renal bood
flow decreases. Also the usual post prandial increase in superior mesenteric
artery blood flow is blunted
56. 4. Watery diarrhea and increased fecal loss. The diarrhea is due to
increased bile salts and UCB in the bowel.
5. Low calcium levels seen in preterm infants receiving phototherapy
6. Retinal damage
7. Tanning of skin
8. Bronze baby syndrome
9. Mutations, sister chromatid exchange and DNA strand breakage
10. Tryptophans are reduced. Also methionine and histidine.
57. EXCHANGE TRANSFUSION
• Removes the partially hemolysed and antibody coated RBCs as well as
unattached antibodies and replaces it with donor RBCs lacking the
sensitizing antigen
• Within in half an hour of exchange bilirubin levels decrease to 60% of its
value
• Furthur increase in post exchange bilirubin is due to the antibody coated
RBCs sequestrated in bone marrow and spleen.
58. INDICATIONS OF EXCHANGE
TRANSFUSION
• When phototherapy fails to prevent a rise in bilirubin to toxic levels
• To correct anemia and heart failure in hydropic infants with hemolytic
disease
• To stop hemolysis and bilirubin production by removing antibody and
sensitized RBCs
• In hemolytic disease immediate exchange is usually indicated if:
• Cord bilirubin level is more than 4.5 mg/dl and cord Hb level is under 11
mg/dl.
• Bilirubin level is rising more than 1mg/dl/hour despite phototherapy or more
than 0.5 mg/dl/hour if Hb is between 11 and 13 mg/dl
• Progression of anemia despite adequate control of bilirubin levels
59.
60. BLOOD FOR EXCHANGE
TRANSFUSION• We use fresh (< 7 days old), irradiated and reconstituted whole blood(PCV
45%-50%) made from PRBCs and FFP collected in CPD.
• In Rh hemolytic disease blood should be type Rh negative cross
mathched against the mother
• In ABO incompatibility blood should be type O Rh negative or Rh
compatible with the mother and infant, be cross mathched against the
mother and the infant and have low titre of naturally occurring anti –a or
anti- b antibodies. Usually type O cells with AB plasma used
• In other isoimmune hemolytic diseases, blood should not contain
sensitizing antigen cross matched against the mother.
• Double the volume of infants blood is used for exchange
transfusion(160ml/kg)
61. TECHNIQUE OF EXCHANGE
TRANSFUSION
Infant under servo controlled radiant warmer and cardiac, blood pressure,
and oxygen saturation monitor in place
An assistant should be assigned to the infant to record volume of blood,
observe the infant and check vital signs
Measurement of potassium and pH of the blood may be indicated if the
blood is > 7 days old or if metabolic abnormalities are noted following
exchange transfusion
62. • Blood should be warmed to 37 c⁰
• Sterile technique to be used. Old,dried umbilical cord can be softened with
saline soaked gauze to facilitate loading the vein and inserting the catheter.
• Exchange is done by push-pull technique through the umbilical vein inserted
only as far as to permit free flow of bood. Catheter in heart can lead to
arrhythmia.
63. • Isovolumetric exchange transfusion involves simultaneously pulling blood out
of umbilical artery and pushing new blood into the umbilical vein. Better
tolerated in small sick or hydropic infants
• If not possible to insert catheter in umbilical vein, exchange transfusion can
accomplished through a central venous catheter placed in the anterior
cubital fossa or into the femoral vein through the saphenous vein
64. • In push pull technique blood is removed in aliquots that are tolerated by
infants:
• 5ml for <1500 g
• 10 ml for 1500-2500g
• 15 ml for 2500-3500g
• 20 ml for >3500g
• Blood in the pint should be gently mixed after every deci litre of clood
transfused to prevent settling of RBCs.
• After exchange transfusion bilirubin levels are measured every 4 hourly**
65. COMPLICATIONS OF EXCHANGE
TRANSFISION1. Hypocalcemia and hypomagnesemia
• Due to citrate in the blood
1. Hypoglycemia
• Glucose conc of CPD is about 300mg/dl and may stimulate insulin secretion and
hypoglycemia
1. Acid – base balance disturbance
• Citrate is metabolized to alkali and if not metabolized as the baby is ill will cause
acidosis
1. Hyperkalemia
• Potassium levels are greatly elevated in stored PRBCS, but washing before before
reconstitution removes the excess potassium
1. Cardiovascular complications
• Perforation of vessels, embolization, vasospasm, thrombosis, infarction
66. 5. Bleeding
• Due to thrombocytopenia and deficient factors
6. Infections
• Bacteremia, hepatitis, cytomegalovirus, HIV etc
7. Hemolysis
• Hemoglobinemia, hemoglobinuria and hyperkalemia caused by overheating
of the blood
8. Graft- versus host diseases
• Transient maculopapular rash, eosinophilia, lymphopenia and
thrombocytopenia
• Can be prevented by using irradiated blood.
9. Miscellaneous ( hypothermia, hyperthermia)
67. IV IMMUNOGLOBULINS
• Adjunctive treatment for hyperbilirubinemia due to hemolytic disease
• Recommended when serum bilirubin is approaching exchange levels
despite maximal interventions including phototherapy
• IVIg (0.5-1 g/kg/dose; repeated in 12 hours) has been shown to reduce the
need for exchange transfusion in both ABO and Rh incompatibility
• Presumably by reducing hemolysis
68. METALLOPORPHYRINS
• Proposed mechanism is competitive enzymatic inhibition of rate-limiting
conversion of heme-protein to biliverdin by heme- oxygenase
• A single intramuscular dose of metalloporphyrins reduce the need for
subsequent phototherapy.
• Particularly useful when jaundice is anticipated like ABO incompatibility/
G6PD deficiency
• Complications include transient erythema if the infant is receiving
phototherapy
• Data on efficacy, toxicity and long term benefits are being evaluated.
69. CONJUGATED
HYPERBILIRUBINEMIA
• Failure to excrete conjugated bilirubin from the hepatocyte in the
duodenum
• Manifested by a CB level of >2mg/dl or CB level >15% of total bilirubin
• a/w hepatomegaly, splenomegaly and pale stools
70. DIFFERENTIAL DIAGNOSIS
1. Liver cell injury(normal bile ducts)
a) Prologed use of parentral nutrition . Sepsis and ischemic necrosis also cause
cholestasis
b) Infection : viral , bacterial, parasitic
c) Metabolic : alpha 1 antitrypsin deficiency, cyctic fibrosis, galactosemia,
tyrosinemia, hypermethionemia,storage disorders, etc.
2. Excessive bilirubin load(insipissated bile duct syndrome) :
• seen in any hemolytic condition especially in infants with erythroblastosis fetalis
• Also in infants supported on ECMO
71. 3. Bile flow obstruction
• Extra hepatic : isolated, a/w choledochal cyst, trisomy 13 or 18 or polysplenia
• Intrahepatic : allagille syndrome , choledochal cyst, bile duct stenosis, rupture of
bile duct, lymph node enlargement
• In the NICU the most common cause of unconjugated hyperbilirubinemia
in decreasing order are :
• Parentral nutrition
• Idiopathic hepatitis
• Biliary atresia
• Alpha 1 anti trypsin deficiency
• Intra uterine infection > choledochal cyst > galactosemia > hemolytic disorders
72. DIAGNOSTIC TESTS AND
MANAGEMENT
• Evaluate for hepatomegaly, splenomegaly, petechiae, chorioretiitis and
microcephaly
• Evaluate liver damage by assessing liver function by SGOT, SGPT, PT, APTT,
and serum albumin levels
• Establish enteral feeds so that PN can be stopped
• Test for bacterial, viral and intra uterine infections
• Serum analysis of alpha 1 antitrypsin deficiency
• Serum and urine amino acid determination
• Urinalysis for glucose and reducing substances
73. • If known causes are ruled out we have to differentiate idiopathic neonatal
hepatitis from bile duct abnormalities
• Investigations done for the same are:
o abdominal usg for choledochal cyst or mass
o Hepatobiliary scan with Tc
o Nasoduodenal tube to be passed for duodenal fluid collection
74. • Most cholestasis in NICU is due to prolonged parenteral nutrition. After ruling
out other causes(sepsis, metabolic disorders, choledochal cyst and
presence of gallbladder) the following management is done :
Enteral feedings even @volumes of 10ml/kg/day is initiated safely
Once enteral feedings are restarted , infants should receive fat soluble vitamin
supplements of A,D,E and K
patients on parenteral nutrition should have there LFTs regularly checked.
decrease the mineral content ( like copper and manganese)
Phenobarbital should not be used to treat cholestasis in this population