Chapter 27: Alloimmune Hemolytic Disease of the Newborn

• A disease in which there is fetal to maternal transfer of red cells that results in immunization of the mother. Then, transplacental transfer of maternal anti–red cell antibodies to the fetus shortens the life span of fetal or newborn red cells.

• Manifestations include fetal anemia, jaundice, and hepatosplenomegaly; in more severe cases, anasarca and kernicterus also occur.

• Asymptomatic transplacental passage of fetal red cells occurs in 75 percent of pregnancies.

• If there is blood group incompatibility between mother and fetus, the chance of maternal immunization increases with the volume of any transplacental hemorrhage.

• Approximately 95 percent of pregnant women have fetomaternal hemorrhage of less than 1.0 mL at delivery.

• Intrapartum fetomaternal hemorrhage of more than 30 mL occurs in approximately 1.0 percent of deliveries.

• Larger volume transplacental hemorrhages are more likely to occur at delivery or during invasive obstetric procedures.

• The risk of sensitization increases with each trimester of pregnancy and is greatest (65%) at delivery.

• Fetomaternal transfusion can occur at the time of chorionic villous sampling, amniocentesis, therapeutic abortion, cesarean section, abdominal trauma, and other situations.

• Prior blood transfusions or abortions also can immunize the mother.

• Maternal red cell antibodies fall into three classes: antibodies directed against the D antigen in the Rh blood group, antibodies directed against the A or B antigens, and antibodies directed against the remaining red cell antigens.

• The D antigen of the Rh blood group system is involved in most serious cases.

• Without prophylaxis, immunization occurs in approximately 12 percent of those at risk with an RhD-positive, ABO-compatible fetus and 2 percent of these with an RhD-positive, ABO-incompatible fetus.

• Anti-D IgG crosses the placenta and leads to a positive antiglobulin test and hemolysis in the infant.

• In ABO hemolytic disease, the mother is usually type O and the fetus is type A or B.

• Anti-A and anti-B antibodies ordinarily cause mild and rarely severe hemolysis. Numerous other causative antibodies have been described but are less common (see Epidemiology).

• The distribution of blood group antigens among different ethnic groups determines their risk of alloimmune hemolytic disease.

• Approximately 16 percent of Americans of European descent are RhD-negative, compared to 8 percent of Americans of African ancestry, 5 percent of persons of Asian Indian ancestry, and 0.3 percent of those of Chinese ancestry.

• More than 50 different red cell antigens have been associated with maternal alloimmunization and with alloimmune hemolytic disease with varying degrees of severity.

• Women can have naturally occurring antigens to blood group A or B (e.g., Mother type O) or may develop other antibodies not screened for prior to blood transfusion.

• Antenatal screening programs detect antibodies in approximately 0.2 percent of pregnant women.

• After anti-RhD, the following antigens may be involved in alloimmunization: Rhc, C, e, cc, Ce, Kell, Duffy, Kidd, and the MNS antigen system.

• The presence of maternal antibodies is not predictive of alloimmune hemolytic disease because of the following: (a) they may be IgM antibodies and not traverse the placenta, (b) the antigens may not be present on fetal red cells or their density very low, (c) the concentration of antibody in maternal blood may be very low, (d) the antibody Ig subclass may not interact with fetal red cells, and other mitigating factors.

Distinctions between ABO and RhD Alloimmunization

• RhD and ABO hemolytic disease differ in several respects (see Table 27–1):

  — ABO hemolytic disease can occur in (a) mothers with O red cells and fetuses with blood group A or B red cells, (b) mothers of type B and fetuses of type A, (c) mothers of type A and fetuses of type B.

  — ABO incompatibility is present in 15 percent of O group pregnancies, but hemolytic disease of the fetus or newborn occurs in about 2 percent of births.

  — The low frequency of ABO hemolytic disease is the result of most anti-A and anti-B being IgM antibodies, which do not easily traverse the placenta.

  — Prenatal testing for maternal anti-A or anti-B antibodies is not predictive of occurrence of alloimmune hemolytic disease because of the unpredictable time of expression of A or B on fetal red cells and the sink for maternal antibodies provided by other fetal tissues that express A or B antigen.

  — ABO incompatibility may be observed during the first pregnancy because of preexisting anti-A and anti-B in the mother. Not so in RhD alloimmunization, unless the mother was previously immunized by transfusion or, rarely, by sharing needles with an RhD-positive intravenous drug abuser.

  — ABO alloimmune hemolytic disease usually results in early neonatal jaundice requiring phototherapy, but only rarely requires exchange transfusion. Moderate anemia and mild hepatosplenomegaly may also be evident.

  — ABO fetomaternal incompatibility rarely leads to severe disease (hydrops fetalis).

  — A somewhat higher degree of jaundice is seen in some ethnic groups (e.g., Americans of African, Southeast Asian, or Hispanic descent). This finding may have to do with variant glucuronyltransferase gene expression.

Hemolytic Disease

• Anemia, jaundice, and hepatosplenomegaly in the newborn are the major findings in alloimmune hemolytic disease.

• The spectrum of severity is wide. In RhD alloimmunization, half the newborn have mild disease and do not need intervention, one-quarter are born at term with moderate anemia and severe jaundice, and one-quarter of fetuses developed hydrops fetalis in utero prior to the availability of intrauterine intervention.

• With severe hemolysis, usually in RhD-sensitized mothers, profound anemia can lead to hydrops fetalis (anasarca caused by hypoproteinemia, cardiac failure), and such fetuses can die in utero.

• Hydrops fetalis is associated with marked extramedullary hematopoiesis in the liver, spleen, kidneys, and adrenal glands. Portal and umbilical vein hypertension, hypoproteinemia (liver dysfunction), and pleural effusions and ascites.

• With milder cases, hemolysis persists until incompatible red cells or the offending IgG is cleared (half-life of IgG is 3 weeks).

• If severe anemia is present, infant displays pallor, tachypnea, tachycardia; cardiovascular collapse and tissue hypoxia can occur if hemoglobin is less than 40 g/L.

• Most affected infants are not jaundiced at birth because of transplacental transport of bilirubin. Jaundice appears during the first postpartum day or in hours after birth if severe hemolysis is present.

• Generally, with mild disease, the bilirubin peaks at day 4 or 5 postpartum and declines slowly thereafter.

• Premature infants may have higher levels of bilirubin of longer duration because of decreased hepatic glucuronyltransferase activity.

• With marked elevation of the serum bilirubin level, kernicterus may develop from deposition of unconjugated bilirubin in the basal ganglia and brainstem nuclei.

• Acute bilirubin encephalopathy is marked initially by lethargy, poor feeding, and hypotonia. If unaddressed, may progress to high-pitched cries, fever, hypertonia, opisthotonus, and irregular respiration.

• Severe involvement can be fatal or lead to long-lasting severe neurologic defects (e.g., choreoathetoid cerebral palsy, sensorineural hearing loss, gaze abnormalities, cognitive abnormalities).

• Occasionally, severe thrombocytopenia or hypoglycemia also occurs and is a poor prognostic sign.

Historical Guideposts

• Obstetric history often guides the laboratory approach. Prior history of transfusions, alloimmunization, severity of prior alloimmune hemolytic disease, prior hydrops fetalis (recurs in 90% of immunized mothers), neonatal death, determining paternity in subsequent pregnancies since the fetus is at risk only if the father is positive for the antigen in question, and related factors may guide the timing and extent of fetal surveillance.

Maternal Red Cell Antigen Typing and Titering

• All pregnant patients should have ABO and RhD typing and testing for unusual red cell alloantibodies early in the pregnancy (10th to 16th week) (see Fig. 27–1).

• Whether RhD-positive or -negative, the mother should be tested again at 28 weeks gestation.

• If alloimmunized, the mother's titer should be determined at 4-week intervals from 20 to 28 weeks and every 2 weeks thereafter.

• Antibody titers are reported as the reciprocal of the highest dilution at which agglutination is observed. A difference of two dilutions is considered significant. If the titer becomes greater than 16 (varies from 8 to 32 in different laboratories), ultrasonography and amniocentesis can be performed to test for the bilirubin level, which predicts disease severity.

• In the United States and the United Kingdom, the anti-D level is compared to an international standard and reported in international units per milliliter (IU/mL). Levels above 4 IU/mL require prompt referral to fetomaternal specialist for monitoring and risk assessment. IU of 4 to 15/mL indicates moderate alloimmune hemolytic disease is likely and over 15 IU/mL implies a severe risk of alloimmune hemolytic disease is likely.

• The significance for antibody titer levels, if anti-D is not involved (e.g., anti-Kell antibodies), has not been determined.

Paternal Zygosity Testing

• In pregnancies in which maternal sensitization occurs or there is a past history of alloimmunization, determining paternal zygosity for all common antigens involved in alloimmune hemolytic disease may determine the risk to the fetus. If the father is homozygous for the antigen in question, one can assume the fetal red cells will carry that antigen. If the father is heterozygous, the fetus will have a 50 percent chance of carrying the antigen.

• If paternal zygosity is unknown, testing fetal red cell blood type early in pregnancy permits appropriate monitoring of the fetus or determines that invasive fetal monitoring is unnecessary.

Fetal DNA, Amniotic Fluid Bilirubin, and Middle Cerebral Artery Doppler Measurements

• Fetal DNA can be obtained from maternal plasma in the first trimester of pregnancy (as early as 5 weeks). Real-time quantitative polymerase chain reaction methods can distinguish maternal from fetal DNA and then amplify fetal exons that include RhD. Accuracy of RhD phenotyping using fetal DNA is 95 percent.

• Amniotic fluid spectrophotometry for bilirubin is an indirect measure of degree of hemolysis. Bilirubin is measured at an optical density (OD) of 450 nm and the elevation in OD₄₅₀ reflects the concentration of bilirubin derived from the fetus. A special nomogram is used to determine the bilirubin for gestational age and four zones in which the bilirubin may fall, providing the probability ranging from no risk of hemolysis to severe hemolytic anemia.

• Because of the risk of amniocentesis, this approach has been replaced by serial noninvasive middle cerebral artery Doppler ultrasound measurements for measuring fetal anemia. Measuring peak blood flow, usually at 1 to 2 week intervals after week 18 until week 35 is a more accurate assessment of anemia than amniotic fluid bilirubin levels. After week 38 of gestation, because of a high false-positive rate with Doppler measurements, amniocentesis and measurement of amniotic fluid OD₄₅₀ is required.

Ultrasonography

• Ultrasonography allows noninvasive description of the fetal condition, an estimate of the need for aggressive management, and a biophysical profile to determine fetal well-being.

• Ultrasonography can be done serially and can detect signs of hydrops such as polyhydramnios, placental enlargement, hepatomegaly, pericardial effusion, ascites, scalp edema, and pleural effusion in roughly that sequence of appearance.

Percutaneous Umbilical Blood Sampling

• More specific information can be obtained by percutaneous umbilical blood sampling (PUBS) (mortality less than 1%) or chorionic villus sampling.

• PUBS (syn. cordocentesis) allows direct measurement of fetal red cell antigens, blood hemoglobin, reticulocyte count, direct antiglobulin test, bilirubin level, blood gases, and lactate levels and can be done at 18 weeks gestation, if severe fetal anemia is indicated by amniotic fluid bilirubin levels or by middle cerebral artery Doppler peak flow measurements.

• PUBS can be done through a 22 gauge spinal needle inserted into the umbilical vein at the site of the cords insertion into the placenta under ultrasonic guidance. If necessary, red cell transfusion can be given by this route.

• Complications of PUBS include umbilical cord bleeding, chorioamnionitis, fetomaternal hemorrhage and maternal red cell sensitization, and fetal death. The latter reported as approximately 3 percent.

Neonatal Assessment

• After delivery, cord blood of the neonate should be sampled for hemoglobin and bilirubin concentrations, the ABO and Rh type, and the direct antiglobulin test. The blood film may show nucleated red blood cells, microspherocytes, and polychromatophilia, if alloimmune hemolytic anemia has occurred. Such testing with a blood group O RhD-positive mother is useful to detect ABO alloimmunization before the newborn is discharged.

• One hour after delivery, blood should be drawn from the mother to evaluate the degree of fetomaternal hemorrhage, so that an appropriate dose of anti-Rh IgG can be given (see "Therapy, Course, and Prognosis" section below).

• Hematopoiesis may be suppressed in the newborn, but marrow recovery is usually complete by 2 months.

• Other diseases can cause hydrops but are distinguished from alloimmune hemolysis by the absence of maternal antibodies.

Fetus

• PUBS (see "Laboratory Evaluation" section above) can be used to deliver a red cell transfusion to a severely affected fetus, based on level of anemia, development of ascites, or a rising bilirubin concentration. This approach has replaced exchange transfusion in some centers because it is a more rapid procedure.

• Packed red cells are transfused to the fetus to achieve a hematocrit of 40 to 45 percent. The red cells are packed to about 75 percent and a calculation made as to what volume of red cells should be necessary to achieve the desired hematocrit in the fetus.

• Intraperitoneal fetal transfusions may be necessary if (a) intravascular access is not possible because the umbilical vessels are too narrow in early pregnancy or (b) fetal size blocks access to the cord later in pregnancy.

• For a woman alloimmunized in a previous pregnancy, fetal transfusions should begin 10 weeks before the time of the earliest prior fetal death or transfusion, but not before 18 weeks gestation unless hydrops is present. Transfusions are given to keep the hematocrit of the fetus in the 20 to 25 percent range and to prevent hydrops.

• O-negative, antigen-negative for any other identified antibody, CMV-negative, irradiated packed red cells are used, cross-matched against the mother's blood.

• The decision about when to deliver the fetus is complex; if possible, transfusions are given up to 34 weeks with delivery at 36 weeks gestation.

• Other treatments to desensitize the mother (maternal immunomodulation) have included intravenous immunoglobulin with or without plasmapheresis, glucocorticoids, or administration of recombinant D-specific antibodies that do not destroy RhD-positive red cells. The nonhemolytic anti-D enters the fetal circulation and competes with natural, hemolytic anti-D for red cell sites, ameliorating the hemolysis.

Neonatal

• The aim of treatment is to prevent bilirubin neurotoxicity.

• Indications for immediate exchange transfusion:

  — The cord blood hemoglobin level is significantly less than normal (perhaps a threshold of ≤110 g/L).

  — The bilirubin level is greater than 4.5 mg/dL.

  — Cord blood bilirubin is rising rapidly (> 0.5 mg/dL per hour).

• If the infant is premature or has unstable vital signs, less stringent criteria are used to give an exchange transfusion. After the first exchange, the rate of rise of bilirubin is used to guide to subsequent transfusions.

• Double volume exchanges will remove, perhaps, 85 percent of sensitized red cells and greater than 50 percent of intravascular bilirubin, and also some maternal anti-D antibody.

• In some centers, prior to exchange, intravenous albumin is given to mobilize extravascular, interstitial bilirubin. Removal of sensitized red cells and prevention of bilirubin formation is the most efficient approach.

• ABO-compatible, RhD-negative, irradiated blood is used, cross-matched against the mother.

• Potential newborn complications of exchange transfusion include hypocalcemia, hypoglycemia, thrombocytopenia, dilutional coagulopathy, neutropenia, disseminated intravascular coagulation, umbilical venous or arterial thrombosis, enterocolitis, and infection. Permanent serious sequelae or neonatal death has been reported in a rate as high as 12 percent in sick infants compared to <1 percent in healthy infants over a period of observation from 1981 to 1995.

• Recombinant human erythropoietin, 200 U/kg, subcutaneously, three times per week for 6 weeks, has been used to enhance recovery of the hemoglobin concentration and decrease the need for postnatal exchange transfusions. It is also useful in Kell antigen-mediated alloimmune disease, since in that case, erythroid hypoplasia is an important factor.

• Phototherapy is used prophylactically in any patient with moderate or severe hemolysis or in infants with bilirubin levels rising at >0.5 mg/dL per hour and is the mainstay of treatment for unconjugated hyperbilirubinemia. The object is to prevent bilirubin neurotoxicity.

• Intensive phototherapy (≥30 microWatts/cm²) in the 430–490 nm band delivered to as much of the infants surface area as possible.

• In full-term infants (at least 38 weeks gestation) with alloimmune hemolytic disease, intensive phototherapy should be instituted if total serum bilirubin is ≥5.0 mg/dL at birth, ≥10 mg/dL at 24 hours after birth, or ≥13 mg/dL 48 to 72 hours after birth.

• Phototherapy is recommended at lower bilirubin levels for preterm or ill infants or infants with a positive direct antiglobulin test, often at serum bilirubin less than 5.0 mg/dL to lessen the need for exchange transfusions.

• Other treatments have been applied. For example, administration of high-dose intravenous immunoglobulin as soon as possible after diagnosis of alloimmune hemolysis is made, decreases the need for phototherapy or exchange transfusion by nonspecific blockade of macrophage Fc receptors and, thereby, a decrease in hemolysis.

• Perinatal survival is greater than 90 percent with intrauterine transfusions in non-hydropic fetuses with severe alloimmune hemolytic disease. The overall survival for hydropic fetuses is approximately 85 percent despite intrauterine transfusion.

• A first trimester screening program increased survival in Kell antigen induced alloimmune hemolytic disease from 61 to 100 percent in the Netherlands.

• Transfusion of red cells matched for RhD, other Rh antigens, and for Kell antigens should be used in premenopausal women.

• RhIg immunoprophylaxis is standard practice for an RhD-negative mother (Table 27–2).

• Intramuscular doses of 100 to 300 μg of Rh immune globulin to nonsensitized RhD-negative mothers within 72 hours of delivery have decreased Rh immunization by greater than 90 percent.

• If the mother is Rh-D-negative with an RhD-positive newborn, administration of antepartum Rh immunoglobulin at 28 weeks has decreased immunization to about 0.1 percent. Rarely, sensitization may occur before the 28th. This approach is standard practice in the United States.

• The standard dose of 300 μg of RhIg (1500 IU) affords protection for a fetomaternal transfusion of 15 mL of RhD-positive red cells or 30 mL or RhD-positive whole blood.

• Larger fetomaternal transfusions can occur in certain circumstances. The blood of RhD-negative women should be tested 1 hour after delivery of an RhD-positive infant. If abruptio placenta or abdominal trauma occurred, the testing can be done after 20 weeks gestation. Testing uses a rosette test requiring very small amounts of maternal blood, followed by a Kleihauer-Betke test for fetal red cells in the maternal blood. Flow cytometric methods are particular useful for quantification of fetal red cells in the maternal blood.

• In patients found to have large fetomaternal transfusions, larger doses of RhIg can be calculated to try to prevent maternal immunization.

• Although immunoprophylaxis has greatly reduced the incidence of alloimmune hemolytic disease, alloimmune sensitization still occurs in 10.6 per 10,000 births in the United States.

• Because the only adequate prophylaxis is for the D-antigen, other less common antibodies will continue to cause hemolytic disease.