Group

C2
Physiology Laboratory
Small Group Discussion
Output

January 20, 2016

[HEMOLYTIC DISEASE OF THE NEWBORN]

By: ASUBARIO, Olufunmilola Omonike; BALADAD, Alvin Byron; DE JESUS, Chrislou; GURUNG, Man
Bahadur; KALANYEG, Kristie; MAHALEE, Naphitcharak; MONTHATHONG, Thanapol; PANLASIGUI,
Rikkimae Maria; SAMSON, Chino Paolo; SOLONIO, Natalie Keith; VALDEZ, Gregorio
 HEMOLYTIC DISEASE OF THE NEWBORN
Introduction

Hemolytic Disease of the Newborn (HDN), also known as erythroblastosis fetalis,

isoimmunization, or blood group incompatibility, occurs when fetal red blood cells
(RBCs), which possess an antigen that the mother lacks, cross the placenta into the
maternal circulation, where they stimulate antibody production. The antibodies return to
the fetal circulation and result in RBC destruction.

A French midwife was the first to report hemolytic disease of the newborn (HDN)
in a set of twins in 1609. In 1932, Diamond and colleagues described the relationship
among fetal hydrops, jaundice, anemia, and erythroblasts in the circulation, a condition
later called erythroblastosisfetalis. Levine later determined the cause after Landsteiner
and Weiner discovered the Rh blood group system in 1940. In 1953, Chown
subsequently confirmed the pathogenesis of Rh alloimmunization to be the result of
passage of Rh-positive fetal RBCs after transplacental hemorrhage into maternal
circulation that lacked this antigen.

In 1966, 2 groups from the United Kingdom and the United States demonstrated,
in a combined study, that anti-D immunoglobulin G (IgG) prophylaxis soon after delivery
prevented sensitization in Rh-negative women. The World Health Organization (WHO)
technical report in 1971 recommended that a dose of 25 mcg (125 IU) of anti-D
immunoglobulin G (IgG) should be given intramuscularly for every 1 mL of fetomaternal
hemorrhage of Rh-positive packed RBCs or 2 mL of whole blood.

In 1998, this recommendation was reinforced by the American Association of

Blood Banks and the American College of Obstetrics and Gynecologists with inclusion
of prophylaxis at 28 weeks' gestation. Routine use of Rh IgG prophylaxis resulted in a
significant decline in the incidence of RhDalloimmunization, and erythroblastosisfetalis
has become rare. The perinatal effects of maternal Rh alloimmunization are now
referred to as hemolytic disease of the fetus and newborn, and fetal manifestations of
the disease are more appreciated with newer technologies such as cordocentesis and
fetal ultrasonography.

Diagnosis:

The diagnosis of HDN is based on history and laboratory findings:

Blood tests done on the newborn baby

 Biochemistry tests for jaundice

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 Peripheral blood morphology shows increased reticulocytes .Erythroblasts (also
known as nucleated red blood cells) occur in moderate and severe disease.
 Positive direct Coombs test (might be negative after fetal interuterine blood
transfusion)
Blood tests done on the mother

 Positive indirect coombs test

 It is suspected when the mother is Rh-negative and the father is Rh-positive.

Signs and Symptoms:

During the prenatal period, an indirect Coombs test can show previous exposure to
Rh-positive antigens. If there are indications that the mother is Rh-sensitized,
amniocentesis can be done to detect bilirubin levels in the amniotic fluid.

 Anemia
 Jaundice
 Kernicterus
 Enlargement of liver and spleen
 If the infants survive, the condition is marked by:
 Heart failure
 Edema
 Pulmonary congestion
 Lethargy seizures
 Mental retardation

Treatment:

Intrauterine Transfusion (IUT)

• Red Blood Cells (RBCs) are infused into abdominal cavity of fetus and then
absorbed into fetal circulation to avoid hydropsfetalis and fetal death.

. • In intraperitoneal IUT, a needle is passed through the mother’s abdomen and

into the abdomen of the fetus by the help of ultrasound image to determine the position
of the fetus and placenta.

- Selection of Blood for IUT: - Most IUTs are accomplished using group O. - Rh-
negative RBCs that are less than 7 days from collection
- RBCs are usually dry packed to remove residual anti-A and anti-B and
reconstituted with group AB fresh frozen plasma t provide coagulation factors.
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 - The risk of these procedures is now largely dependent on the prior condition of
the fetus and the gestational age at which transfusion is commenced.
- Titer greater than 32 for anti-D and 8 for anti-K OR four fold increase in titer
indicates need for analysis of amniotic fluid.
• Amniocentesis - Perform at 28 weeks if HDN in previous child
- Perform at 22 weeks if previous child severely affected • Perform if maternal
antibody increases before 34th wk.
- High values of bilirubin in amniotic fluid analyses by the Liley method or a
hemoglobin concentration of cord blood below 10.0 g/mL.

Immunomodulators
Class Summary
These agents normalize antibody levels in patients with primary defective
antibody synthesis. They prevent and treat certain bacterial and viral infections and
reduce the immune-mediated hemolysis and phagocytosis.
Intravenous immunoglobulin (Gamimune, Gammagard, Sandoglobulin, Gammar-P)

Several studies have reported success in minimizing the need for exchange
transfusion in severe HDN with IVIG. Effective adjunct to phototherapy. Mechanism of
action appears to be related to blockage of Fc receptors in the neonatal
reticuloendothelial system. Studies have also documented decreased hemolysis after
administration of IVIG using carboxyhemoglobin levels. Administration in doses of 500-
1000 mg/kg in the first few hours of life to a newborn with severe hemolysis should be
considered. However, efficacy depends on timing of administration, duration of
treatment, and severity of hemolysis. Should be prepared by and dispensed from
pharmacy and should not be mixed with normal saline. Dispensed as either 3% or 6%
solution.

Colony-stimulating Factor

Epoetinalfa, recombinant (Epogen, Procrit)

Purified glycoprotein produced from mammalian cells modified with gene coding
for human erythropoietin (EPO). Amino acid sequence is identical to that of endogenous
EPO. Biological activity mimics human urinary EPO, which stimulates division and
differentiation of committed erythroid progenitor cells and induces release of
reticulocytes from bone marrow into the blood stream.

Competitive hemeoxygenase inhibitor

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 Stannsoporfin (SnMP, Stanate)

Also known as tin-mesoporphyrin. Investigational in the United States. Phase III

clinical trials completed. Structural analog of heme that blocks hemeoxygenase (HO-1),
a rate-limiting enzyme in bilirubin production, thereby preventing the conversion of
heme to bilirubin. Heme is excreted unchanged in bile and is not stored in tissue. It is
inert and does not enter the brain or interact with DNA. It does not affect previously
formed bilirubin conjugation or excretion in liver. Several randomized, controlled and,
when possible, blinded studies over the last decade that involved >700 neonates with all
principle forms of neonatal jaundice have shown SnMP to be effective in preventing and
blocking jaundice progression. Phototherapy was eliminated in 97% of treated infants.

Also inhibits nitric oxide synthase and soluble guanylyl cyclase. Repeated doses
lead to inhibition of intestinal hemeoxygenase involved in iron absorption and may lead
to anemia. It also stimulates HO-1 transcription and protein levels. The half-life as
measured in healthy adult volunteers is 3.8 h.

Further Inpatient Care

The following may be indicated in patients with hemolytic disease of newborn (HDN):

 The stabilization of a hydropic newborn requires a high level of intensive coordinated

management by a neonatal team well prepared for the possibly affected infant.

 In general, immediate intubation followed by draining of pleural effusions and ascites

results in immediate improvement in respiratory gas exchange.

 A cautious correction of anemia with packed RBCs or by exchange transfusion is

necessary to prevent circulatory overload.

 These neonates have normal blood volume but elevated central venous pressure.

 A close monitoring of metabolic status (eg, watching for hypoglycemia, hypocalcemia,

hyperkalemia, acidosis, hyponatremia, renal failure) is absolutely essential to achieve
a successful outcome.

 Despite of the first use of phototherapy by Cremer and associates more than 40 years
ago, no standard method for delivering phototherapy is yet available.

 Phototherapy units differ widely with respect to the type and size of lamps used.
The efficacy of phototherapy depends on the spectrum of light delivered, the blue-
green region (425-490 nm) of visible light being the most effective; irradiance
(µW/cm2/nm); and surface area of the infant exposed.

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  High-intensity phototherapy first described by Tan in 1977 uses irradiance greater
than 25 µW/cm2/nm up to 40 µW/cm2/nm when a dose-response relationship to
bilirubin degradation reaches a plateau.

 Nonpolar bilirubin is converted into 2 types of water-soluble photoisomers as a

result of phototherapy. The initial and most rapidly formed configurational isomer 4z,
15e bilirubin accounts for 20% of total serum bilirubin level in newborns undergoing
phototherapy and is produced maximally at conventional levels of irradiance (6-9
µW/cm2/nm).

 The structural isomer lumirubin is slowly formed, and its formation is irreversible and
is directly proportional to the irradiance and surface area of skin exposed to
phototherapy. Lumirubin is the predominant isomer formed during high-intensity
phototherapy.Decrease in bilirubin is mainly the result of excretion of these
photoproducts in bile and removal via stool. In the absence of conjugation, these
photoisomers can be reabsorbed by way of the enterohepatic circulation and
diminish the effectiveness of phototherapy.

 Phototherapy implementation guidelines were recently addressed in recent clinical

practice guidelines published by the American Academy of Pediatrics.The
recommendations are as follows:

 The guidelines are based on total serum bilirubin levels and the direct fraction
should not be subtracted from the total unless it is more than 50% of the total serum
bilirubin level.

 Intensive phototherapy should be started for babies with hemolytic disease. This
implies the use of irradiance in the 430-490 nm band of more than 30 µW/cm2/nm
delivered to as much of the infant's surface area as possible. This can be
accomplished using special blue fluorescent tubes that are labeled F20T12/BB
(General Electric, Westinghouse, Sylvania) or TL52/20W (Phillips, Eindhoven, The
Netherlands) and positioning them 10-15 cm above the infant. When fluorescent
tubes are used, they should be brought as close to the infant as possible to
increase irradiance. However, when halogen spotlights are used, the distance
above the infant should be as per the manufacturer's instructions because
spotlights can cause burns. Phototherapy lights emit minimal UV radiation that does
not cause erythema and is completely absorbed by the acrylic Plexiglas covering of
the tubes.

 Irradiance should be measured using radiometers recommended by the

manufacturers of phototherapy systems at multiple sites on the infant's body
surface illuminated by the phototherapy lamp and the measurements averaged.

 The infant should be in the bassinet, and the sides should be lined with white cloth
or aluminum foil to expose more surface area. The exposed surface area is
increased by the use of 1-2 fiberoptic pads that should be placed under the infant or

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 by the use of BiliBed (MedelaInc; McHenry, Ill) or Bili-Bassinet (Olympic Medical;
Seattle, Wash), which provides phototherapy from above and below. The diaper
should be removed if bilirubin is approaching exchange levels.

 The serum bilirubin declines by 0.5-1 mg/dL in the first 4-8 hours on intensive
phototherapy and should be measured in 2-3 hours to document the effectiveness.

 If the serum bilirubin level continues to rise despite intensive phototherapy or is

within 2-3 mg/dL of exchange level, administer intravenous immunoglobulin (IVIG)
at 0.5-1 g/kg over 2 hours and repeat every 12 hours if needed.

 High-dose IVIG 1 g/kg given early in high-risk neonates with rapid rise of bilirubin
level (>0.5 mg/kg/h) and worsening anemia (Hb< 12 g/dL) despite intensive
phototherapy, is be able to eliminate the need for exchange transfusion and to
reduce duration of phototherapy. The number needed to treat (NNT) is 6.

 Phototherapy is indicated in the term infant with hemolytic disease of the newborn
immediately after birth due to Rh disease and due to ABO incompatibility as follows:

 Unborn (cord blood) - Total serum bilirubin level of more than 3.5 mg/dL
 Age less than 12 hours - Total serum bilirubin level of more than 10 mg/dL
 Age less than 18 hours - Total serum bilirubin level of more than 12 mg/dL
 Age less than 24 hours - Total serum bilirubin level of more than 14 mg/dL
 Age 2-3 days - Total serum bilirubin level of more than 15 mg/dL
 Immediately after birth in all preterms who weigh less than 2500 g

 Exchange transfusion removes circulating bilirubin and antibody-coated RBCs,

replacing them with RBCs compatible with maternal serum and providing albumin with
new bilirubin binding sites. The process is time consuming and labor intensive but
remains the ultimate treatment to prevent kernicterus. The process involves the
placement of a catheter via the umbilical vein into the inferior vena cava and removal
and replacement of 5- to 10-mL aliquots of blood sequentially, until about twice the
volume of the neonate's circulating blood volume is reached (ie, double-volume
exchange).

 This process removes approximately 70-90% of fetal RBCs. The amount of bilirubin
removed directly varies with the pretransfusion bilirubin level and amount of blood
exchanged. Because most of the bilirubin is in the extravascular space, only about
25% of the total bilirubin is removed by an exchange transfusion. A rapid rebound of
serum bilirubin level is common after equilibration and frequently requires additional
exchange transfusions.

 The indications for exchange transfusion are controversial, except for the fact that
severe anemia and the presence of a rapidly worsening jaundice despite optimal
phototherapy in the first 12 hours of life indicate the need for exchange transfusion. In

Physiology Laboratory Small Group Discussion Output |Hemolytic Disease of the Newborn 7
 addition, the presence of conditions that increase the risk of bilirubin encephalopathy
lowers the threshold of safe bilirubin levels.

 Guidelines for exchange transfusion in neonates with hemolytic disease of the

newborn are as follows:
 Total serum bilirubin level of more than 20 mg/dL - Weight more than 2500 g
(healthy)
 Total serum bilirubin level of more than 18 mg/dL - Weight more than 2500 g (septic)
 Total serum bilirubin level of more than 17 mg/dL - Weight 2000-2499 g
 Total serum bilirubin level of more than 15 mg/dL - Weight 1500-1999 g
 Total serum bilirubin level of more than 13 mg/dL - Weight 1250-1499
 Total serum bilirubin level of 9-12 mg/dL - Weight less than 1250

 The following are indications for exchange transfusion:

 Severe anemia (Hb< 10 g/dL)

 Cord bilirubin > 4 mg/dL.
 Rate of bilirubin rises more than 0.5 mg/dL despite intensive phototherapy
 Severe hyperbilirubinemia
 Serum bilirubin-to-albumin ratio exceeding levels that are considered safe

 Exchange transfusion should be considered in newborns born at more than 38 weeks'

gestation with a bilirubin-to-albumin ratio of 7.2 and in newborns born at 35-37 weeks'
gestation with a bilirubin-to-albumin ratio of 6.8. Exchange transfusion is not free of
risk, with the estimated morbidity rate at 5% and the mortality rate as high as 0.5%.
Apnea, bradycardia, cyanosis, vasospasm, and hypothermia with metabolic
abnormalities (eg, hypoglycemia, hypocalcemia) are the most common adverse
effects.

 IVIG has been shown to reduce the need for exchange transfusion in hemolytic
disease of the newborn due to Rh or ABO incompatibility. The number needed to treat
to prevent one exchange transfusion was noted to be 2.7 and was estimated to be 10,
if all the infants with strongly positive direct Coombs test were to receive the
medication.In addition, it also reduced the duration of hospital stay and
phototherapy.Although it was very effective as a single dose, multiple doses were
more effective in stopping the ongoing hemolysis and reducing the incidence of late
anemia.

 A randomized, controlled trial by Smits-Wintjens et al, however, failed to show the

benefit of prophylactic single-dose IVIG at 0.75 g/kg within 4 hours of life in severely
sensitized neonates with prior IUT due to Rh alloimmunization.While IVIG has been
proven to be safe, a retrospective review reported almost 30-times increased risk of
necrotizing enterocolitis (NEC) in late preterm and term infants.

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 Tin-mesoporphyrin in a dose of 4.5 mg/kg (6 µmole/kg) was used in an infant with
persistent hemolysis due to Rh alloimmunization to prevent need for further
phototherapy, without any adverse effects.

Prognosis:
The severity of this condition can vary. Some babies have no symptoms. In other
cases, problems such as hydrops can cause the baby to die before, or shortly after
birth. Severe HDN may be treated before birth by intrauterine blood transfusion.

Prevention:
The most severe form of this disease, which is caused by Rh incompatibility, can
be prevented if the mother is given a shot of a medicine called RhoGAM at certain times
during and after her pregnancy. If you have had a baby with this disease, talk with your
doctor if you plan to have another baby.

References:

1.Bowman JM. Hemolytic disease (erythroblastosisfetalis). Creasy RK,

ResnikR.Maternal-fetal medicine. 4th edition. Philadelphia: WB Saunders; 1999.
736-767.

2.[Guideline] Snyder EL, Lipton KS. Prevention of hemolyic disease of the

newborn due to anti-D Prenatal/perinatal testing and Rh immune globulin
administration. American Association of Blood Banks Association Bulletin. 1998.
98:1-6.

3.Fasano RM. Hemolytic disease of the fetus and newborn in the molecular
era. Semin Fetal Neonatal Med. 2015 Nov 14.

4.van der Schoot CE, Tax GH, Rijnders RJ, de Haas M, Christiaens GC. Prenatal
typing of Rh and Kell blood group system antigens: the edge of a
watershed. Transfus Med Rev. 2003 Jan. 17(1):31-44.

5.Singleton BK, Green CA, Avent ND, et al. The presence of an RHD
pseudogene containing a 37 base pair duplication and a nonsense mutation in
africans with the Rh D-negative blood group phenotype. Blood. 2000 Jan 1.
95(1):12-8.

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