127: Rh Incompatibility

Isoimmune hemolytic anemia of variable severity may result when Rh incompatibility develops between an Rh-negative mother previously sensitized to the Rh (D) antigen and her Rh-positive fetus. The onset of clinical disease begins in utero as the result of active placental transfer of maternal immunoglobulin (Ig)G-Rh antibody. It is manifested as a partially compensated, moderate to severe hemolytic anemia at birth, with unconjugated hyperbilirubinemia developing in the early neonatal period.

Historically, Rh hemolytic disease of the newborn accounted for up to a third of symptomatic cases seen and was associated with detectable antibody in ∼15% of Rh-incompatible mothers. The use of Rh immunoglobulin (RhoGAM) prophylaxis has reduced the incidence of Rh sensitization to <1% of Rh-incompatible pregnancies. Other alloimmune antibodies have become relatively more important as a cause of hemolysis. Anti-c, Kell (K and k), Duffy (Fya), Kidd (Jka and Jkb), MNS (M, N, S, and s), and less commonly anti-C and anti-E may cause severe hemolytic disease of the newborn. This cannot be prevented by the use of D antigen–specific Rh immunoglobulin.

Initial exposure of the mother to the Rh antigen occurs most often during parturition, miscarriage, abortion, and ectopic pregnancy. Invasive investigative procedures such as amniocentesis, chorionic villus sampling, and fetal blood sampling also increase the risk of fetal transplacental hemorrhage and alloimmunization. Recognition of the antigen by the immune system ensues after initial exposure, and reexposure to the Rh antigen induces a maternal anamnestic response and elevation of specific IgG-Rh antibody. Active placental transport of this antibody and immune attachment to the Rh-antigenic sites on the fetal erythrocyte are followed by extravascular hemolysis of erythrocytes within the fetal liver and spleen. The rate of the hemolytic process is proportionate in part to the levels of the maternal antibody titer but is more accurately reflected in the antepartum period by elevation of the amniotic fluid bilirubin concentration and in the postpartum period by the rate of rise of unconjugated bilirubin. In contrast to ABO incompatibility, the greater antigenicity and density of the Rh-antigen loci on the fetal erythrocyte facilitates progressive, rapid clearance of fetal erythrocytes from the circulation. A demonstrable phase of spherocytosis will be absent. Compensatory reticulocytosis and shortening of the erythrocyte generation time, if unable to match the often high rate of hemolysis in utero, results in anemia in the newborn infant and a risk of multiple systemic complications.

1. Birth order. The firstborn infant is at minimum risk (<1%) unless sensitization has occurred previously. Once sensitization has occurred, subsequent pregnancies are at a progressive risk for fetal disease.

2. Fetomaternal hemorrhage. The volume of fetal erythrocytes entering the maternal circulation correlates with the risk of sensitization. The risk is ∼8% with each pregnancy but ranges from 3 to 65%, depending on the volume of fetal blood (3% with 0.1 mL compared with 22% with >0.1 mL) that passes into the maternal circulation.

3. ABO incompatibility. Coexistent incompatibility for either the A or B blood group antigen reduces the risk of maternal Rh sensitization to 1.5–3.0%. Rapid immune clearance of these fetal erythrocytes after their entry into the maternal circulation exerts a partial protective effect. It confers no protection once sensitization has occurred.

4. Obstetric factors. Cesarean delivery or trauma to the placental bed during the third stage of labor increases the risk of significant fetomaternal transfusion and subsequent maternal sensitization.

5. Gender. Male infants are reported to have an increased risk of more severe disease than females, although the basis for this observation is unclear.

6. Ethnicity. Approximately 15% of whites are Rh negative compared with 7% of blacks and almost 0% in Asiatic Chinese and Japanese. The risk to the fetus varies accordingly.

7. Maternal immune response. A significant proportion of Rh-negative mothers (10–50%) fail to develop specific IgG-Rh antibody despite repeated exposure to Rh antigen.

1. Symptoms and signs

   1. Jaundice. Unconjugated hyperbilirubinemia is the most common presenting neonatal sign of Rh disease, usually appearing within the first 24 hours of life.

   2. Anemia. Low cord blood hemoglobin at birth reflects the relative severity of the hemolytic process in utero and is present in ∼50% of cases.

   3. Hepatosplenomegaly. Enlargement of the liver and spleen is seen in severe hemolysis, sometimes occurring in association with ascites, with an increased risk for splenic rupture.

   4. Hydrops fetalis. Severe Rh disease has a historical association with hydrops fetalis and at one time was its most common cause. Clinical features in the fetus include progressive hypoalbuminemia with ascites, pleural effusion, or both; severe chronic anemia with secondary hypoxemia; and cardiac failure. There is an increased risk of late fetal death, stillbirth, and intolerance of active labor. The neonate frequently has generalized edema, notably of the scalp, which can be detected by antepartum ultrasonography, cardiopulmonary distress often involving pulmonary edema and severe surfactant deficiency, congestive heart failure, hypotension and peripheral perfusion defects, cardiac rhythm disturbances, and severe anemia with secondary hypoxemia and metabolic acidosis. Currently, nonimmune conditions are more commonly associated with hydrops fetalis. Secondary involvement of other organ systems may result in hypoglycemia or thrombocytopenic purpura.

Obligatory screening in an infant with unconjugated hyperbilirubinemia includes the following studies:

1. Blood type and Rh type (mother and infant). These studies establish the likelihood of Rh incompatibility and exclude the diagnosis if the infant is Rh negative, with one exception (see direct antiglobulin test [direct Coombs test], Section VI.C).

2. Reticulocyte count. Elevated reticulocyte levels, adjusted for the degree of anemia and gestational age in preterm infants, reflect the degree of compensation and support a diagnosis of an ongoing hemolytic process. Normal values are 4–5% for term infants and 6–10% for preterm infants (30–36 weeks' gestational age). In symptomatic Rh disease, expected values are 10–40%.

3. Direct antiglobulin test (direct Coombs test). A strongly positive direct Coombs test indicates that fetal red blood cells (RBCs) are coated with antibodies and is diagnostic of Rh incompatibility in the presence of the appropriate setup and an elevated reticulocyte count. If Rh immunoglobulin was given at 28 weeks' gestation, subsequent passive transfer of antibody will result in a false-positive direct Coombs test without associated reticulocytosis. Very rarely, a strongly positive direct Coombs test is associated with a falsely Rh-negative infant when all fetal RBC Rh-antigenic sites are covered by a high titer of maternal antibodies.

4. Blood smear. Polychromasia and normoblastosis proportionate to the reticulocyte count are typically present. Spherocytes are not usually present. The nucleated RBC count is often >10 per 100 white blood cells.

5. Bilirubin levels. Progressive elevation of unconjugated bilirubin on serial testing provides an index of the severity of the hemolytic process. An elevated direct fraction is most likely to be secondary to a laboratory artifact in the first 3 days of life and should not be subtracted from the total bilirubin when making management decisions. In the most severely affected infants, particularly those who are hydropic, the intense extramedullary erythropoiesis may cause hepatocellular dysfunction and biliary canalicular obstruction with significant elevated direct bilirubin by 5–6 days of age.

6. Bilirubin-binding capacity tests. Correlation between measurements of serum albumin, free bilirubin, bilirubin saturation index, and reserve binding capacity and outcome has been variable. The role of these values in directing the management of patients remains unclear.

7. Glucose and blood gas levels. These should be monitored closely.

8. Supplementary laboratory studies. Supportive diagnostic studies may be required when the basis of the hemolytic process remains unclear.

   1. Direct Coombs test in the mother. This study should be negative in Rh disease. This test can be positive in the presence of maternal autoimmune hemolytic disease, particularly collagen vascular disease.

   2. Indirect antiglobulin titer (indirect Coombs test). This test detects the presence of antibodies in the maternal serum. Rh-positive RBCs are incubated with the serum being tested for the presence of anti-D. If present, the RBCs now coated with anti-D are agglutinated by an antihuman globulin serum reflecting a positive indirect antiglobulin (Coombs) test result. The reciprocal of the highest dilution of maternal serum that produces agglutination is the indirect antiglobulin titer.

   3. Carbon monoxide (CO). The severity of Rh disease may be determined by measurement of endogenous CO production. When heme is catabolized to bilirubin, CO is produced in equimolar amounts. Hemoglobin binds the CO to form carboxyhemoglobin (CO Hb) and then is finally excreted in the breath. CO Hb levels are increased in neonates with hemolysis. CO Hb levels >1.4% have been correlated with an increased need for exchange transfusion.

1. Antepartum treatment. Verification of the Rh-negative status at the first prenatal visit may be obtained by the following measures:

   1. Maternal antibody titer. Once an IgG-Rh antibody has been identified, it is important to determine the titer. Serial antibody titer determinations are required every 1–4 weeks (depending on the gestational age) during pregnancy. Invasive fetal testing becomes indicated when the titer is above a critical level, usually between 1:8 and 1:16. A negative antibody screen (indirect Coombs test) signifies absence of sensitization. This test should be repeated at 28–34 weeks' gestation.

   2. RhoGAM. Current obstetric guidelines suggest giving immunoprophylaxis at 28 weeks' gestation in the absence of sensitization.

   3. Amniocentesis. If maternal antibody titers indicate a risk of fetal death (usual range, 1:16–1:32), amniocentesis should be performed to assess fetal Rh genotype and assess severity. Fetal Rh-genotype determination can also be made from fetal cell free DNA found in maternal plasma. To reasonably predict the risk of moderate to severe fetal disease, serial determinations of amniotic fluid bilirubin levels present photometrically at 450 nm are plotted on standard graphs according to gestational age (known as the Liley curve). Readings falling into very high zone II or zone III indicate that hydrops will develop within 7–10 days. Zone I indicates no fetal hemolytic disease or no anemia.

   4. Ultrasonography. As a screening study in pregnancies at risk, serial fetal ultrasound examinations allow detection of scalp edema, ascites, or other signs of developing hydrops fetalis. Peak systolic middle cerebral artery velocity can reliably detect moderate and severe fetal anemia and thus reduce the need for more invasive diagnostic procedures such as amniocentesis and cordocentesis.

   5. Intrauterine transfusion. Based on the studies just mentioned, intrauterine transfusion may be indicated because of possible fetal demise or the presence of fetal hydrops. This procedure must be performed by an experienced team. The goal is maintenance of effective erythrocyte mass within the fetal circulation and maintenance of the pregnancy until there is a reasonable chance for successful extrauterine survival of the infant.

   6. Glucocorticoids. If premature delivery is anticipated, glucocorticoids should be given to accelerate fetal lung maturation and reduce the risk of intraventricular hemorrhage.

   7. Reduction of maternal antibody level. Intensive maternal plasma exchange and high-dose intravenous immunoglobulins (IVIGs) have been reported of value in the severely alloimmunized pregnant woman to reduce circulating maternal antibodies levels by >50%.

2. Postpartum treatment

   1. Resuscitation. Moderately to severely anemic infants with or without hydropic features are at risk for high-output cardiac failure, hypoxemia secondary to decreased oxygen-carrying capacity or surfactant deficiency, and hypoglycemia. These infants may require immediate single-volume exchange blood transfusion at delivery to improve oxygen-carrying capacity, mechanical support of ventilation, and an extended period of monitoring for hypoglycemia.

   2. Cord blood studies. A cord blood bilirubin level >4 mg/dL, a cord hemoglobin <12 g/dL, or both usually suggests moderate to severe disease. The cord blood is used for these and initial screening studies, including blood typing, Rh typing, and Coombs test.

   3. Serial unconjugated bilirubin studies. Determination of the rate of increase in unconjugated bilirubin levels provides an index of the severity of the hemolytic process and the need for exchange transfusion. Commonly used guidelines include a rise of >0.5 mg/dL/h or >5 mg/dL over 24 hours within the first 2 days of life or projection of a serum level that will exceed a predetermined “exchange level” for a given infant (usually 20 mg/dL in term infants).

   4. Phototherapy. In severe Rh hemolytic disease, phototherapy is used only as an adjunct to exchange transfusion. Phototherapy decreases bilirubin levels and reduces the number of total exchange transfusions required. See Chapters 58 and 100 for phototherapy details.

   5. Exchange transfusion. (For the procedure, see Chapter 30.) Exchange transfusion is indicated if the unconjugated bilirubin level is likely to reach a predetermined “exchange level” for that patient. Optimally, exchange transfusion is done well before this exchange level is reached to minimize the risk of entry of unconjugated bilirubin into the central nervous system. Consideration should be given to irradiation of blood before the transfusion is given, particularly in preterm infants or infants expected to require multiple transfusions, to reduce the risk of graft-versus-host disease. The process removes 70–90% of the fetal red cells, but only 25% of the total bilirubin because most of the bilirubin is in the extravascular space. A rapid rebound of serum bilirubin is common after reequilibration, and thus additional exchange transfusions may be required.

   6. Heme oxygenase inhibitors (stannsoporfin). Tin (Sn) porphyrin can decrease the production of bilirubin and reduce the need for exchange transfusion and duration of phototherapy. It is an inhibitor of heme oxygenase, which is the enzyme that allows the production of bilirubin from heme. The dose of stannsoporfin is 6 μmol/kg IM as a single dose given within 24 hours of birth with severe hemolytic disease, and it is available via compassionate use protocol.

   7. IVIG. By blocking neonatal reticuloendothelial Fc receptors, and thus decreasing hemolysis of the antibody-coated RBCs, high-dose IVIG (1 g/kg over 4 hours) reduces serum bilirubin levels and the need for blood exchange transfusion with ABO or Rh hemolytic diseases. (See Chapters 80 and 127.) Caution should be used when considering treatment with IVIG, as there are emerging reports of increased incidence of NEC in term and late-preterm infants with hemolytic disease of the newborn and isoimmune neonatal thrombocytopenia who were treated with IVIG.

3. RhoGAM prophylaxis. Most cases of incompatibility involve the D antigen. RhoGAM given at 28 weeks' gestation, within 72 hours of suspected Rh-antigen exposure, or both reduces the risk of sensitization to <1%; the recommended dosage (300 mcg) should be well in excess of the amount of Rh antigen transfused (300 mcg for every 25 mL of fetal blood in maternal circulation). The amount of fetal blood entering the maternal circulation may be estimated using the Kleihauer-Betke acid elution technique Fetomaternal hemorrhage during the immediate postpartum period.

   No treatment equivalent to RhoGAM is available for maternal Rh sensitization to non-D antigens, notably C and E antigens. These antigens, however, are significantly less antigenic than the D antigen, clinical manifestations of incompatibility are frequently milder, and the risk of severe disease is considerably less.

4. Hydrops fetalis. Skilled resuscitation and anticipation of selective systemic complications may prevent early neonatal death.

   1. Isovolumetric partial exchange transfusion with type O Rh-negative packed erythrocytes. Raises the hematocrit and improves the oxygen-carrying capacity (see Chapter 30).

   2. Central arterial and venous catheterization. May be performed to provide the following measures:

      1. Isovolumetric exchange transfusion

      2. Monitoring of arterial blood gas levels and central venous and systemic blood pressures

      3. Monitoring of fluid and electrolyte balance, particularly renal and hepatic function, calcium-to-phosphorus ratio, and serum albumin levels, as well as appropriate hematologic studies and serum bilirubin levels

   3. Positive-pressure mechanical ventilation. This measure may include increased levels of positive end-expiratory pressure, if pulmonary edema is present, as a means of stabilizing alveolar ventilation. Treatment with exogenous surfactant may be considered in particular when the infant is judged to be not fully mature.

   4. Therapeutic thoracentesis or paracentesis. May be performed to remove fluid that may further compromise respiratory effort. Excessive removal of ascitic fluid may lead to systemic hypotension. (See Chapters 27 and 37.)

   5. Volume expanders. May be necessary, in addition to erythrocytes, to improve peripheral perfusion defects. This should be done with caution because most hydropic infants are hypotensive or poorly perfused because of hypoxic heart failure rather than hypovolemia, or both.

   6. Drug treatment. May include diuretics such as furosemide for pulmonary edema and pressor agents such as dopamine (for dosing, see Chapter 148). In the case of cardiac rhythm disturbances, appropriate drugs may be used if indicated.

   7. Electrocardiography or echocardiography. May be needed to determine whether cardiac abnormalities are present.

Prenatal mortality for infants at risk of anti-D Rh isoimmunization is currently ∼1.5% and has decreased significantly over the past 2 decades. Antenatal immune prophylaxis and improved management techniques, including amniotic fluid spectrophotometry, intrauterine transfusion, and advances in neonatal intensive care, have been largely responsible for this reduction. Isolated cases of severe isoimmunization still occur because of isoimmunization by other than anti-D antibody or failure to receive immune prophylaxis and may exhibit the full spectrum of disease, including an increased risk of stillbirths and early neonatal morbidity and mortality.