HDFN Notes

Hemolytic Disease of the Fetus and Newborn (HDFN)

General Overview

• Definition: Incompatibility of maternal antibody(ies) towards fetal RBCs.

  • Maternal antibodies destroy fetal RBCs, a condition called Erythroblastosis Fetalis (EF).

  • Occurs both in-utero (prepartum) and ex-utero (postpartum).

• Cause: Fetal RBCs (that leak into the mother's blood) have antigens foreign to the mother, stimulating maternal antibody production.

• Severity: Ranges from mild to severe.

• Antibody Class: IgG.

• Effects of EF: Fetus develops Hydrops Fetalis (HF).

  • HF includes:

    • General anemia.

    • Possible edema.

    • Possible cardiac failure.

    • Potential fetal death.

Classifications of HDFN

• ABO:

  • Caused by mother-baby ABO incompatibility (e.g., Mother type O, Baby type A).

  • Most common.

• Rh:

  • Usually anti-D, potentially with other Rh antibodies (anti-C, -c, -E, -e).

  • Common.

• Other:

  • Unexpected immune antibodies other than Rh (e.g., Jk, K, Fy, S).

  • Rare.

HDFN Caused by Other Antibodies

• Uncommon, occurring in approximately 0.8% of pregnant women.

• Immune alloantibodies usually due to anti-E, -c, -Kell, -Kidd, or -Duffy.

• Anti-K:

  • Disease ranges from mild to severe.

  • Over half of maternal cases with anti-K are caused by previous blood transfusions.

  • Is the second most common form of severe HDFN.

• RBC Alloantibodies in 178 Pregnancies Requiring Intrauterine Transfusions (Ohio State Alloimmunization Committee, 1965-2007):

  • Anti-D: 44%

  • Anti-D and other(s): 42%

  • Anti-c: 6%

  • Anti-Kell: 5%

  • Anti-E: 2%

  • Anti-Jsb, Anti-Fya: 1%

In-Utero (Prepartum) Overview

• Dangers of HDFN in-utero:

  • Fetal severe anemia.

  • Fetal heart failure.

  • Fetal death.

• Maternal antibodies attack fetal RBCs causing fetal anemia; Hydrops Fetalis (HF) can develop.

• Destroyed RBCs release unconjugated/indirect bilirubin into the fetal body.

• Increased unconjugated/indirect bilirubin can be detrimental. While in-utero, the unconjugated bilirubin is sequestered out via the mother's liver.

In-Utero Bilirubin Conjugation

• Fetus: Antibody-coated red cells -> Anemia, Heart Failure, Fetal Death (Hydrops fetalis)

• Indirect bilirubin = unconjugated bilirubin

• Maternal Liver: Maternal conjugation occurs; Indirect bilirubin -> Direct bilirubin = Conjugated bilirubin (Deactivated/harmless); Excreted by mother during pregnancy

Maternal Prenatal Care / Testing

• Prenatal care under Obstetrician guidance is crucial.

• Helps in early detection of potential pregnancy problems.

• Certain lab tests during the initial visit:

  • ABO, Rh (including Du), & ABS tests are performed on first Obstetrical visit

  • If ABS is negative, repeat at 24 weeks.

  • If ABS is positive, perform Ab ID. If Ab is IgG, must perform an Ab titer also.

  • Retain all maternal samples.

  • Periodically, perform Ab titer to check for increase in titration (> 1:32 is significant).

  • When performing the current titer, must run the last previous titer sample in parallel with current titer sample.

  • An increase in Ab titration could indicate a higher risk for HDFN.

• The higher the maternal Ab titer, the higher the risk is for HDFN.

• If maternal serum Ab titer is high, the physician may test amniocentesis fluid for presence of ag (amniocytes).

• Optional: Test father to verify he is ag positive.

Amniocentesis

• Based on maternal Ab titer results & HDFN is suspected:

• Amniocentesis is performed on mother at 18-20 weeks gestation.

  • Amniocentesis is the procedure of obtaining a sample of amniotic fluid from the womb.

• Amniotic fluid bilirubin is a direct indicator of the severity of HDFN.

• Scan fluid in spectrophotometer at steadily increasing wavelengths to determine change in OD when 450 nm is reached (absorbance of bilirubin).

• If \Delta OD increases at a later scan, bilirubin levels are increasing.

Liley's Graph

• Used to assess the severity of HDFN based on amniotic fluid analysis.

• Plots \Delta OD 450 values against gestational age (weeks).

• Zones indicate severity (I, IIA, IIB, IIIA, IIIB) with corresponding risk levels (Mild, Moderate, Severe).

Queenan Curve

• An alternative to Liley's graph for assessing HDFN severity.

• Plots \Delta OD 450 values against gestational age.

• Differentiates between affected (Rh positive) and unaffected (Rh negative) pregnancies.

• Indicates risk of intrauterine death.

Intervention, If Needed

• Intrauterine transfusion criteria:

  1. Fetal hydrops detected on ultrasound.

  2. Fetal Hgb level is <10 g / dL (severe anemia).

  3. Amniotic fluid \Delta OD 450 results on Liley's Graph or Queenan Curve indicate hyperbilirubinemia.

• Risks and benefits must be weighed by the pediatrician.

• Intrauterine blood requirements:

  • RBCs:

    • < 5 days old.

    • CPDA-1 or AS-3 (lacks mannitol, which is a diuretic; avoid diuretics in neonates).

    • Ag negative to corresponding maternal alloantibody(ies).

    • O Neg.

    • CMV neg, if tested OR product is leuko-reduced ("CMV safe").

    • Hgb S negative.

    • Irradiated.

Postpartum Overview

• Dangers of HDFN Postpartum

  • The maternal Ab(s) continue(s) to destroy fetal RBCs postpartum.

  • The following signs & symptoms could be seen:

    • Anemia - Gross anemia can cause fetal cardiac failure

    • Hepatomegaly

    • Splenomegaly

    • Jaundice - Neonate's immature liver cannot conjugate bilirubin efficiently; therefore, an accumulation of neonatal unconjugated / indirect bilirubin (which is toxic) occurs

    • Kernicterus - Untreated accumulation of unconjugated bilirubin can lead to kernicterus that can cause severe retardation; if further left untreated, fetal demise

Neonatal Jaundice

• After Delivery: Antibody-coated cell -> Infant's spleen -> Hemoglobin -> Infant's liver -> Indirect bilirubin

• Indirect bilirubin = unconjugated bilirubin

Effects of Accumulated Unconjugated Bilirubin

• Yellowing of skin-Jaundice

• Yellowing of eyes

• Excess bilirubin in blood

• Kernicterus: Bilirubin moves from bloodstream into brain tissue

HDFN: Neonatal Treatment Options

1. Phototherapy (light wavelength set at 460 to 490 nm):

   • Unconjugated / indirect bilirubin is sensitive to strong light

   • Used to change the unconjugated / indirect bilirubin to isomers which are less lipophilic and less toxic to the brain.

2. Exchange transfusion - indications:

   • Hgb is <10 g / dL

   • Bilirubin level is \geq 20 mg / dL

ABO HDFN

• Most common form of HDFN.

• Can occur at the 1st delivery and subsequent pregnancies; no prior exposure is needed.

• Commonly seen in an "O" mother (with IgG form of Anti-A,B) delivering an "A" neonate; "B" type neonate is less common.

• Baby can have mild to moderate jaundice.

• Usually less severe than Rh HDFN.

• Occurs in only 3%; is severe in only 1%; and <1:1,000 require exchange transfusion.

ABO HDFN: Lab Testing - Maternal Prenatal

• Prenatal testing is usually NOT predictive of ABO HDFN development

Lab Testing - Postpartum AKA, Cord Blood Work-Up

• Mother

  • Repeat ABO / Rh and ABS (usually performed at hospital admission prior to delivery)

• Baby

  • Determine ABO / Rh (including weak D)

  • Perform DAT

    • If baby IgG DAT is positive, may perform elution procedure

      • An elution procedure uses baby RBCs and a supernatant media. The elution is performed using heat or chemical to "dissociate" the Ab off the baby's RBC. The Ab is contained in the supernatant media where Ab ID can be performed. Ab found in eluate is of maternal origin.

  • Perform DAT (con't.)

    • To determine ABO HDFN, test eluate against A1 & B cells

    • Some hospitals may not perform an elution on the baby RBCs (for allo Ab ID) because of the large amount of blood needed to perform the elution. Removing too much blood for lab tests can cause a condition called, iatrogenic anemia*, in the patient.

    • * anemia caused by withdrawing too much blood

ABO HDFN: Prevention & Treatment

1. Not preventable (as in Rh0(D) HDFN)

2. Hyperbilirubinemia / jaundice is commonly treatable by using only phototherapy

3. When serum bilirubin result reaches \geq 20 mg / dL, an exchange transfusion is indicated

Phototherapy

• The treatment of choice in ABO HDFN, dependent on the Lab Test Results, e.g., Hgb & especially the Bilirubin

Rh HDFN

• Most common-causing Ab is anti-D, but Abs to C, c, E, e antigens are known to cause Rh HDFN

• Most severe form of HDFN

• Severe anemia may occur resulting in severe hyperbilirubinemia / jaundice

• Kernicterus could result

• Usually affects only 2nd or subsequent pregnancies* (mother allo-immunized at delivery of 1st pregnancy)

• Has pre-formed Abs during subsequent pregnancies

• * Must consider miscarriages, abortions, & prenatal fetal / maternal bleeds

RH HDFN - Abs

• Anti-D - most common form of Rh HDFN. The disease varies from mild to severe.

• Anti-E - is a mild disease

• Anti-c - range from a mild to severe disease

• Anti-e - rare

• Anti-C - rare

• Ab combinations (i.e., anti-c and anti-E Abs occurring together) - can be severe

RH HDFN: LAB TESTING - PRENATAL

• Mother tested for:

  • ABO & Rh including Du test

  • ABS

    • If Ab screen is positive, determine:

      • Ab ID & Ab Titer

      • Ag status of the father, optional

      • Ig subclass of the maternal Ab, if indicated

• Determine initial Ab titer, then repeat approximately every 4 weeks. Run parallel testing on the last previous sample with current sample.

RH HDFN: LAB TESTING - PRENATAL

• If anti-D (or any other Rh Ab) titer reaches >1:32, consider further testing:

  • Middle Cerebral Artery-Peak Systolic Velocity (MCA-PSV) - color Doppler / Ultrasound Transducer which is a reliable predictor of fetal anemia

  • Amniocentesis

    • Amniocentesis is uncommonly used for the managing of anemia because MCA-PSV is noninvasive and gives the same information

    • The concentration of bilirubin pigment in the amniotic fluid (measured by the \DeltaOD 450 nm procedure) predicts worsening of the fetal hemolytic disease

LAB TESTING - POSTPARTUM AKA, CORD BLOOD WORK-UP

• Mother

  • Repeat ABO / Rh and ABS tests

    • If ABS is positive, must do Ab ID procedure

      • If a weak anti-D is present, it may be due to the administration of an antepartum Rh Immune Globulin shot. Check patient's OB history with RN or physician's office.

      • If the weak anti-D Ab is due to the Rh Immune Globulin shot, the Ab ID report should state presence of a "passive" anti-D…not as an "allo" anti-D.

• Baby

  • Determine ABO / Rh (including weak D)

  • Perform DAT

    • If baby DAT is positive, may perform elution

      • An elution procedure uses baby RBCs and a supernatant media. The elution is performed using heat or chemical to "dissociate" the Ab off the baby's RBC. The Ab is contained in the supernatant media where Ab ID can be performed. Ab found in eluateis of maternal origin.

    • To determine Rh HDFN (or any other allo Ab HDFN), use eluate against Ab ID panel cells

    • Some hospitals may not perform an elution on the baby RBCs for allo Ab ID due to creating iatrogenic anemia condition.

RH HDFN: LAB TESTING - POSTPARTUM

• Enumerate # of fetal cells present in mother's blood

• Fetal Hgb Screen Test / Rosette Test (qualitative test) - demonstrates small number of D-positive cells in mother's D- negative cell suspension.

RH HDFN: LAB TESTING - POSTPARTUM

• Kleihauer-Betke (KB) test (quantitative test)

  • Blood sample from mother treated with acid, then stained

  • Fetal cells are resistant to acid; mother's RBCs become “ghost cells”

  • Determine # of fetal cells in first 2,000 maternal cells counted

  • Volume of fetal cells = # of fetal cells x maternal blood volume divided by 2,000

HDFN Comparison: ABO vs Rh(D)

EXCHANGE TRANSFUSION

• Can occur in either ABO or Rh HDFN, dependent on the Lab Test Results, e.g., Hgb & Bilirubin

EXCHANGE TRANSFUSION - NEONATE TREATMENT

• Occurs after birth

• When phototherapy is unsuccessful in reducing the rise of the bilirubin level and hyperbilirubinemia from unconjugated / indirect bilirubin is still present, the second choice of treatment is the Exchange Transfusion.

• If maternal Ab is causing rapid destruction of fetal RBCs, Exchange Transfusion is performed, instead of phototherapy.

EXCHANGE TRANSFUSION - NEONATE TREATMENT

• Exchange Transfusion can occur when either conditions exists:

  • Hgb <10 g / dL

  • Bilirubin level is \geq 20 mg / dL

• Dependent on the age of the newborn, transfusion can occur either via the umbilical cord entry or extremities entry

EXCHANGE TRANSFUSION - NEONATE TREATMENT

• RBCs used for the exchange transfusion must be:

  • < 5 day old

  • CPDA-1 or AS-3 (lacks mannitol which is a diuretic; do not give diuretics to neonates)

  • RBCs must also be ag negative to a corresponding allo Ab present

  • O negative

  • CMV negative, if tested OR product is leuko-reduced

  • Hgb S negative

  • Irradiated

• Plasma must be "AB" - the Rh may be disregarded & may not be irradiated

EXCHANGE TRANSFUSION - NEONATE TREATMENT

Beneficial Effects of a 2-volume Exchange Transfusion:

1. Removes 50% of the bilirubin

2. Removes 80-90% of the infant's sensitized RBCs

3. Removes 80-90% maternal incompatible antibody

4. Replacement of incompatible RBCs with compatible RBCs

EXCHANGE TRANSFUSION - EXTREMITIES ENTRY

• The neonatal exchange transfusion process is very slow.

• Goal is to avoid:

  1. Any clinically significant hemodynamic shifts

  2. Any metabolic abnormalities