Pediatrics

Hemolytic disease of the newborn (alloimmunization)

OVERVIEW: What every practitioner needs to know

Are you sure your patient has hemolytic disease of the newborn? What are the typical findings for this disease?

Early and excessive jaundice: The most common symptoms are early (especially during the first 24 hours of life) and excessive jaundice, and positive antibody testing (Coombs, direct antibody test [DAT]). Most commonly, the mother is blood type O and her baby is A or B; less often the mother will be Rh negative and the baby Rh-positive. Occasionally there is an antibody to a so-called minor blood group antigen such as Duffy, Kell, or others. Although called "minor," these incompatibilities can also be very significant, especially anti-Kell.

Anemia: The next most common finding is anemia, although this usually is less severe than the hyperbilirubinemia, slower to develop, and will persist for a much longer time.

Edema: The most severely affected infants (almost always Rh incompatibility) will also have severe intrauterine anemia with generalized edema (hydrops fetalis, erythroblastosis fetalis), including pleural effusions and ascites leading to severe respiratory distress and cardiovascular instability. Fortunately, this is much less common today with modern antenatal diagnosis and treatment, including intrauterine intravascular transfusions, which prevent and even reverse these findings before birth.

What other disease/condition shares some of these symptoms?

Other causes of hemolysis, for example, hereditary spherocytosis or glucose-6-phosphate dehydrogenase deficiency may present with early and/or excessive jaundice, but the Coombs or DAT test is not positive. Early and excessive jaundice can also occur from polycythemia (hematocrit >60%-65%) or from a cephalohematoma or extensive bruising, all resulting in an excessive amount of hemoglobin to be broken down and processed; again test results for red blood cell antibodies are negative. Anemia without hyperbilirubinemia is not likely to be due to a hemolytic process, but rather to blood loss or other more unusual problems such as bone marrow failure. Anemia caused by hemolysis in the immediate newborn period is typically associated with severe hyperbilirubinemia.

What caused this disease to develop at this time?

Hyperbilirubinemia does not occur before birth because bilirubin is ordinarily not conjugated before birth. The purpose of conjugation is to make bilirubin water soluble for excretion into bile and ultimate removal from the body. In order for bilirubin to be removed from the fetus it must remain unconjugated and lipid soluble so that it can be transported across the placenta for excretion by the mother. This is effective even if hemolysis is severe. Anemia, in contrast, can be present and produce symptoms before birth.

Women who are blood type O have naturally occurring anti-A and anti-B antibodies, mostly of the IgM class but some IgG. IgM antibodies do not cross the placenta, but IgG antibodies, if present, can cross and bind to the infant's red blood cells, causing removal by the infant's reticuloendothelial system. This excessive red blood cell destruction leads to hyperbilirubinemia because of slow conjugation of bilirubin by the liver. This process (ABO incompatibility) is generally mild, and with unpredictable severity from one pregnancy to the next, although occasionally infants are severely affected.

Although approximately 15% of pregnancies are a "set-up" for ABO incompatibility (mother O, baby A or B), a positive DAT result is seen in only about 30% of these (4%-5% of pregnancies), and significant hemolysis is seen in only about 15% of those, or 1% of all pregnancies. ABO incompatibility is less severe than Rh incompatibility because there are fewer group A or B antigen sites on neonatal red blood cells compared with Rh antigens, allowing sensitized A or B cells to survive longer in the infant's circulation than with anti-Rh antibodies. O-A incompatibility is most common, but O-B incompatibility is more likely to be problematic.

Women who are Rh-negative must first be sensitized against the Rh antigen before producing anti-Rh antibodies (generally anti-D, but anti-C/c or anti-E/e are possible). Anti-Rh antibodies are of the IgG class, and will cross the placenta. The usual way for a pregnant woman to be sensitized is through a previous pregnancy or unrecognized miscarriage, or through a blood transfusion of incompatible blood.

To prevent this, women who are Rh blood group negative are tested for antibody both at the start of the pregnancy and at 28 weeks' gestation; if still negative at 28 weeks, they are given passive immunization with anti-Rh globulin to prevent them from becoming sensitized (actively immunized) later in pregnancy when the placental barrier may be less effective in preventing fetal cells from entering the maternal circulation. They are then given an additional dose of anti-Rh globulin after delivery to further protect them from sensitization.

Before the use of anti-Rh globulin, the incidence of Rh isoimmunization was 10% of Rh-negative mothers after one pregnancy, which decreased to 1.8% with a single postpartum prophylactic dose of Rh immunoglobulin, and to 0.14% with an additional prophylactic dose at 28 weeks. Other factors predisposing to Rh sensitization are a woman's refusal of anti-Rh globulin (e.g., Jehovah's Witness) or a large fetomaternal hemorrhage (wherein the amount of blood entering the maternal circulation exceeds the capacity of the usual dose of anti-Rh globulin to prevent sensitization).

The usual dose of anti-Rh globulin given to the mother (300 µg) protects against up to 30 mL fetal whole blood (15 mL fetal red blood cells); if a larger hemorrhage is suspected because of apparent acute blood loss and shock in the infant, estimation of the amount transfused by a Kleihauer-Betke test on the mother's blood is indicated to estimate the dose of anti-Rh globulin needed. Once sensitized, the Rh-negative mother will have an anamnestic response in future incompatible pregnancies that will lead to a more severe hemolysis in each successive pregnancy involving an Rh-positive infant.

What laboratory studies should you request to help confirm the diagnosis? How should you interpret the results?

  1. 1. Blood type and antibody screen on the mother's blood.

  2. 2. Blood type and DAT on the cord blood or infant blood, looking for antibody on the infant's red blood cells.

Although in the case of ABO incompatibility, the indirect antibody test may be positive (antibody present in the infant's plasma); if the DAT result is negative, these infants are not at increased risk of hemolysis.

Complete blood count and reticulocyte count in the infant (to evaluate hematocrit, smear, and number of nucleated red blood cells). The smear will frequently show microcytic spherocytes if an isoimmune hemolytic process is present.

In the case of known Rh sensitization (or sensitization to the minor blood group antigens) diagnosed antenatally, the bilirubin level should be determined on the cord blood, and at frequent intervals after birth (every 4-6 hours), because the rate of rise of the bilirubin level is an indicator and predictor of the severity of hemolysis. The bilirubin should be fractionated one time to ensure that the hyperbilirubinemia is unconjugated rather than mixed. The total bilirubin, however, should be used for all decisions regarding therapy.

A rate of rise greater than 5 mg/dL/24 h (or >0.5 mg/dL/h) is suggestive of hemolysis in anyinfant; therefore, clinical jaundice(bilirubin >5 mg/dL needed to be clinically visible) in the first 24 hours strongly suggests a hemolytic process.

Although the rate of rise of bilirubin is fairly steady and predictable in Rh disease, in ABO incompatibility it is not; the bilirubin often rises quickly to 10-15 mg/dL during the first 24 hours, then plateaus at 15-20 mg/dL during the second day.

In the case of ABO incompatibility (mother O, baby A or B), the strength of the positive DAT correlates somewhat, but not completely, with the severity of the hemolysis expected in the infant. Most infants with a 4+ DAT result will have significant hyperbilirubinemia requiring phototherapy; a negative DAT result essentially eliminates ABO incompatibility-related hemolysis as the cause of excessive jaundice, and other causes should be sought.

In the case of Rh incompatibility, the DAT result will likely be strongly positive, and the infant will likely be significantly affected. The exception is that up to 15% of infants born to mothers who have received anti-Rh globulin at the 28th week may have a false-positive antibody screen, related to transplacental passage of the anti-Rh globulin given to the mother.

The serum albumin level is measured because unconjugated bilirubin binds to albumin. Free bilirubin (i.e., unconjugated bilirubin not bound to albumin) is thought to be the neurotoxic form of bilirubin. Therefore, treatment (phototherapy or exchange transfusion) would be instituted earlier in an infant with a low serum albumin concentration than in one with a robust concentration.

If you are able to confirm that the patient has hemolytic disease of the newborn, what treatment should be initiated?

Phototherapy: Phototherapy should begin as soon as the diagnosis of Rh incompatibility is confirmed. Since ABO incompatibility is much less predictable, serum bilirubin concentrations can be followed to see if significant hemolysis is actually occurring, and phototherapy can then be started in accordance with published guidelines. Phototherapy (light in the 425- to 475-nm wavelength range, peak effect at 460 nm, in the blue spectrum) acts by changing the isomeric structure of the bilirubin molecule, resulting in a more water-soluble stereoisomer that can be excreted in urine and bile without first being conjugated in the liver, the rate-limiting step for bilirubin excretion in the bile.

Intensive phototherapy with maximal exposure of body surface is most effective, including the combined use of conventional phototherapy with a supplemental fiberoptic phototherapy blanket under the infant. A fiberoptic phototherapy blanket alone is not sufficient for a term or late preterm infant with hemolysis because of the inadequate body surface area exposed to the light. Similarly, double-bank phototherapy will expose more surface area than a single bank of lights and is therefore more effective.

When to institute phototherapy in the case of ABO incompatibility has been outlined by the American Academy of Pediatrics (AAP) Subcommittee on Hyperbilirubinemia: Management of Hyperbilirubinemia in the Newborn Infant 35 or More Weeks of Gestation. The risk of neurologic injury (bilirubin encephalopathy or kernicterus) is dependent on gestational age, postnatal age, and associated illness (presence of hemolysis, acidosis, sepsis, asphyxia, and perhaps the serum albumin concentration). For this reason the limits of acceptable bilirubin concentrations change with age and overall status. The nomogram published by the AAP is quite useful.

Feedings: Most infants should be fed while receiving phototherapy, as this will stimulate intestinal motility and biliary drainage and help with bilirubin excretion. If, however, exchange transfusion is needed, feedings should be held temporarily because intestinal perfusion may be abnormal during that procedure. Breast-feeding can be supplemented with expressed breast milk or formula if the infant is having difficulty latching on; use of a breast shield may also be helpful. Intravenous fluid administration for hydration is not generally needed unless the infant is being fed nothing by mouth for possible exchange transfusion or other reasons, or if albumin or intravenous immunoglobulin (IVIG) is to be administered.

Albumin administration: Since bilirubin is primarily bound to albumin in the bloodstream, and it is free or unbound bilirubin that is available to enter and damage the central nervous system (bilirubin encephalopathy or kernicterus), ensuring an adequate serum albumin concentration is essential. A bilirubin-albumin ratio of 7:1 is considered safe under most circumstances. If the ratio is approaching this limit, albumin 1 g/kg (25% albumin, 4 mL/kg) can be given intravenously over 1-2 hours to improve binding while the blood for exchange transfusion is being prepared.

IVIG: If hemolysis is severe, and there is increasing hyperbilirubinemia despite intensive phototherapy, or the bilirubin is within 2-3 mg/dL of the designated exchange level, IVIG can be administered at a dose of 0.75 g/kg over 2-3 hours. Although recommended in the 2004 AAP Committee statement, a Cochrane review in 2002 recommended trials of higher quality be performed before endorsing this therapy. A randomized controlled trial by Smits-Wintjens et al suggests that IVIG therapy is not helpful.

Exchange Transfusion: The final intervention for severe hemolysis that is unresponsive to the above procedures, or in the presence of signs of acute bilirubin encephalopathy, is exchange transfusion. Exchange transfusion is considered when the bilirubin concentration continues to rise despite intensive phototherapy and is indicated in a full term infant when the bilirubin is 20 mg/dL or more in the first 48 hours, 25 mg/dL or more after 48 hours, or whenever clinical symptoms of bilirubin encephalopathy are thought to be present.

A double-volume exchange transfusion (160 mL/kg, with type O, Rh-negative blood compatible with the mother, irradiated and leukodepleted) will remove approximately 85% of the infant's cells, as well as much of the antibody, and will reduce the total bilirubin load by approximately 25% by removing that which is circulating in the blood. The serum bilirubin concentration will rebound after exchange transfusion. Pretreatment with 25% albumin has been shown to increase the amount of bilirubin removed, and to decrease the need for further exchange transfusion and the length of subsequent phototherapy needed.

Exchange transfusion is invasive, requires central venous access (generally through the umbilical vein, or vein and artery), and should only be undertaken in a neonatal intensive care unit (NICU) setting with experienced personnel, as there are risks associated with the procedure, even in an otherwise healthy infant. The risks include metabolic disturbances, thrombocytopenia, infection, necrotizing enterocolitis, and death. The rate of severe morbidity is approximately 5%, whereas the mortality rate in otherwise healthy infants is less than 0.5%. As with criteria for phototherapy, criteria for exchange transfusion vary with infant age, gestation, associated illness, and albumin concentration, and is also outlined by the AAP in nomogram form.

What are the adverse effects associated with each treatment option?

Phototherapy: This is a benign procedure, although some infants develop diarrhea, and occasionally require intravenous hydration to supplement their enteral intake. In infants with direct or conjugated hyperbilirubinemia (cholestasis), phototherapy results in bronze baby syndrome, which resolves when the cholestasis improves. Severe blistering of the skin and agitation during phototherapy may be a sign of congenital porphyria, which is very rare.

Albumin, IVIG: An intravenous line is required for administration. These products are made from human plasma, and there is a theoretical risk for transmission of infections (although not reported) or other serious side effects (rare in newborn).

Exchange transfusion: Central access is necessary as is the use of blood products. There is the possibility of infection, metabolic derangements during and after the procedure, thrombocytopenia, necrotizing enterocolitis, and death. The risk of serious morbidity is approximately 5%, whereas the risk of mortality in an otherwise healthy term infant is less than 0.5%. Nevertheless, when signs of acute bilirubin encephalopathy are present or the bilirubin continues to rise relentlessly despite other measures, this is the best option for lowering the bilirubin level and interrupting the hemolytic process.

What are the possible outcomes of this disease?

Since the maternal antibodies persist in the newborn for 2-3 months, there will be ongoing destruction of red blood cells (hemolysis) for the first 8-12 weeks of life. For this reason, anemia will continue to be a problem long after the hyperbilirubinemia has resolved, and may require one or more transfusions of packed red blood cells (top-up transfusion) during this time. This may occur whether or not the infant had exchange transfusion, IVIG, or intrauterine transfusion. Packed red blood cell transfusion is needed in approximately 85% of Rh-sensitized infants, and less often in ABO-incompatible infants.

The hematocrit should be followed weekly, with transfusion performed when the hematocrit is less than 22%-25% and/or the infant is symptomatic (e.g., poor weight gain, easily fatigued). Folic acid 50 µg orally daily may be helpful in maintaining the maximal hematocrit possible. Iron supplementation is not likely to be needed immediately, especially if there have been previous, including intrauterine, transfusions. In general, by several weeks of age, some iron supplementation (e.g., 2-4 mg/kg/d) would be appropriate. The use of erythropoietin has been recommended, especially if the anemia is associated with a low reticulocyte count at several weeks of age, consistent with continued marrow suppression after intrauterine transfusions.

No long-term effects are expected unless there has been neurologic injury from the hyperbilirubinemia. The goal of therapy is to avoid this outcome.

Signs of acute bilirubin encephalopathy include lethargy, poor feeding, and hypotonia, progressing to irritability alternating with stupor, high-pitched cry, and hypertonicity with retrocollis and opisthotonus. Ultimately, there is apnea, coma, seizures, and death. Kernicterus/chronic bilirubin encephalopathy is a permanent and nonprogressive condition manifesting clinically as choreoathetoid (dyskinetic) cerebral palsy, limited upward gaze, deafness (auditory dyssynchrony, which is abnormal brainstem auditory evoked response in the face of normal otoacoustic emissions), and enamel dysplasia of the deciduous teeth. Cognitive deficits are rare.

What causes this disease and how frequent is it?

Epidemiologic features of Rh incompatibility: Hemolytic disease of the newborn is more common in whites (15% Rh negative) than in blacks (7% Rh negative), and is rare in IndoEurasians (2% Rh negative). If Rh immunoglobulin prophylaxis is not given during pregnancy, there is an 8%-16% risk that an Rh-negative woman with an Rh-positive, ABO-compatible infant would be sensitized; with antenatal and postpartum prophylaxis, the risk decreases to 0.14%.

Epidemiologic features of ABO incompatibility: Although 15% of pregnancies are a "set-up" for ABO incompatibility (mother O, baby A or B), a positive DAT result is seen in only about 30% of these cases, and significant hemolysis in only about 15% of those, or 1% of all pregnancies.

Other clinical manifestations that might help with diagnosis and management

In the case of severe erythroblastosis fetalis, there may be severe clinical manifestations, including generalized edema, respiratory failure due to surfactant deficiency with or without bilateral pleural effusions, hypotension and cardiac failure due to severe anemia (sometimes worsened by pericardial effusion), ascites, hepatosplenomegaly (due to congestive heart failure as well as extramedullary hematopoiesis), and skin lesions (blueberry muffin spots, palpable purpura), also due to extramedullary hematopoiesis.

These infants are extremely ill and require massive support. Exchange transfusion with O-negative blood compatible with the mother to increase the hematocrit without further volume overload is preferred over simple transfusions; exchange transfusion will also decrease the bilirubin load. Despite maximal support, mortality remains high in these infants. Such an infant should be transferred as soon as possible to a high-level NICU for care if antenatal transfer to a perinatal center has not been possible.

What complications might you expect from the disease or treatment of the disease?

The complication to be avoided is bilirubin encephalopathy. Adverse effects of treatments are described above.

How can this disease be prevented?

Prophylaxis for maternal Rh isoimmunization is possible through administration of anti-Rh globulin to Rh-negative women after miscarriage or abortion, during each pregnancy at 28 weeks' gestation, and after delivery of an Rh-positive infant. Prophylaxis of ABO incompatibility is not possible. Prophylaxis of isoimmunization involving the minor blood group antigens is also not possible.

What is the evidence?

"Management of hyperbilirubinemia in the newborn infant 35 weeks or more of gestation". Pediatrics. vol. 114. 2004. pp. 297-316.

(Extensive review of available literature regarding management of hyperbilirubinemia)

Eder, AF. "Update on HDFN: new information on long-standing controversies". Immunohematology. vol. 22. 2006. pp. 188-195.

(Discussion of minor blood group antigens and antenatal monitoring for severity of fetal involvement)

Harkness, UF, Spinnato, JA. "Prevention and management of RhD isoimmunization". Clin Perinatol. vol. 31. 2004. pp. 721-742.

(Extensive review of antenatal management of Rh-sensitized mother and use of intrauterine transfusion)

Iskander, I, Gamaleldin, R, Houchi, SE, Shenawy, AE. "Serum bilirubin and bilirubin/albumin ratio as predictors of bilirubin encephalopathy". Pediatrics. vol. 134. 2014. pp. e1330-1339.

(Although both total bilirubin and bilirubin/albumin ratio are strong predictors of acute and longterm outcome, bilirubin/albumin ratio does not improve prediction over total serum bilirubin alone)

Kaplan, M, Hammerman, C, Vreman, HJ. "Hemolysis and hyperbilirubinemia in antiglobulin positive, direct ABO blood group heterospecific neonates". J Pediatr. vol. 157. 2010. pp. 772-777.

(Description of relative proportions and severity of O-A and O-B incompatibility from a prospective population cohort)

Kuzniewicz, MW, Wickremasinghe, AC, Wu, YW, McCulloch, CE. "Incidence, etiology, and outcomes of hazardous hyperbilirubinemia in newborns". Pediatrics. vol. 134. 2014. pp. 504-509.

(Large population reviewed for instances of bilirubin concentration 30 mg/dL or more for determination of etiology, incidence and outcome)

Maisels, MJ, McDonagh, AF. "Phototherapy for neonatal jaundice". N Engl J Med. vol. 358. 2008. pp. 920-928.

(Extensive review of mechanism of action of phototherapy and bilirubin metabolism)

Murray, NA, Roberts, IAG. "Haemolytic disease of the newborn". Arch Dis Child Fetal Neonatal Ed. vol. 92. 2007. pp. F83-88.

(Review of all forms of hemolytic disease in the newborn, including antibody-mediated)

Ross, MB, Alarcon, P. " Hemolytic disease of the fetus and newborn". NeoReviews. vol. 14. 2013. pp. e83-e88.

(Updated review of hemolytic disease of the newborn)

Smits-Wintjens, VEHJ, Walther, FJ, Rath, MEA. "Intravenous immunoglobulin in neonates with Rhesus hemolytic disease: a randomized controlled trial". Pediatrics. vol. 127. 2011. pp. 680-686.

(Questions the efficacy of IVIG in hemolytic disease)

Stevenson, DK, Wong, RJ. "Metalloporphyrins in the management of neonatal hyperbilirubinemia". Semin Fetal Neonat Med. vol. 15. 2010. pp. 164-168.

(Review of potential utility of heme oxygenase inhibitors to prevent excessive heme catabolism in susceptible neonates)

Watchko, JF. "Identification of neonates at risk for hazardous hyperbilirubinemia: emerging clinical insights". Pediatr Clin North Am. vol. 56. 2009. pp. 671-687.

(ABO hemolytic disease as an important cause of exaggerated hyperbilirubinemia)

Ongoing controversies regarding etiology, diagnosis, treatment

There is controversy about the utility of IVIG for hemolytic disease. It is currently recommended by the AAP as of their 2004 publication, but its efficacy has been questioned.

For many years, the use of metalloporphyrins, competitive inhibitors of heme oxygenase (the initial and rate-limiting enzyme involved in heme degradation) has been proposed for the prevention of bilirubin production in infants with known hemolytic processes. Although used in Europe and studied in the United States, no metalloporphyrin is currently approved by the US Food and Drug Administration for clinical use. Examples are tin mesoporphyrin, zinc protoporphyrin, chromium mesoporphyrin. and zinc deuteroporphyrin bis glycol.

The heme oxygenase system has protean effects, similar to nitric oxide pathways, so positive and negative consequences are difficult to predict. Although metalloporphyrins appear to be very effective in preventing bilirubin production, the preferred compound, dose, and method of administration is not known.

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