Hematologic and Oncologic Problems in the Fetus and Neonate

Fetal–Maternal Hemorrhage

Fetal cells may be found in the maternal circulation in about half of all pregnancies. At 20 weeks’ gestation, the fetoplacental volume is 30 mL, and it is rare for transplacental hemorrhage to exceed 1 mL of fetal red blood cells. By term, fetal–maternal hemorrhage (FMH) during delivery can exceed 30 mL of fetal blood, although in only 0.3% of pregnancies. Fetal–maternal hemorrhage is associated with procedures such as external cephalic version or traumatic amniocentesis. The diagnosis is made by demonstrating fetal RBCs, which contain mostly Hb F, in a maternal blood sample; therefore, the analysis must be done before the fetal cells are cleared from the maternal circulation. The test usually is performed within the first few hours after delivery. In cases of ABO blood group incompatibility, the red blood cells may be cleared more rapidly, and the test may be falsely negative.

The prevention of maternal Rh D alloimmunization requires accurate detection and quantification of fetal blood cells in the maternal circulation. If a D-negative mother has evidence of FMH, Rh immune globulin (Rh IgG) can reduce D-sensitization and the resultant hemolytic disease of the newborn. Vaginal or peripheral blood samples from women with vaginal bleeding late in pregnancy may also be evaluated for the presence of fetal blood cells. The rosette screen is a sensitive, FDA-approved screening test for FMH when the mother is D-negative. Maternal red blood cells are incubated with anti-D and then washed. Indicator D-cells are added, and in the presence of fetal D-positive cells, aggregates or rosettes will be seen by light microscopy. Confirmation of a positive screening test is performed with a quantitative method. The acid elution technique, or Kleihauer-Betke test, is the most widely available quantitative test for FMH. This method exploits the stability of Hb F–containing RBCs in acid solution relative to cells containing Hb A. False-positive test results are seen in women with any condition (including many of the hemoglobinopathies) that elevates their own Hb F level. The Kleihauer-Betke test suffers from problems with standardization and is labor intensive. Some centers now offer flow cytometry–based quantification of FMH by detecting either Hb F or Rh D or a combination. Flow cytometry can distinguish adult F cells from fetal RBCs.

Twin–Twin Transfusion Syndrome

In monochorionic diamniotic multiple gestations, twin–twin transfusion syndrome (TTTS) (see Chapter 21 ) can be diagnosed by ultrasound-demonstrating oligohydramnios in the donor sac and polyhydramnios in the recipient sac. Twin–twin transfusion syndrome occurs in 8%-10% of twin pregnancies with monochorionic diamniotic placentation and accounts for about half of the perinatal deaths. Twin anemia polycythemia sequence (TAPS) is defined as the presence of anemia in the donor and polycythemia in the recipient twin. Prenatal middle cerebral artery peak systolic velocity criteria have been established for diagnosis in the absence of oligohydramnios–polyhydramnios. Blood can be exchanged unequally between the fetuses through placental vascular interfetal connections. Transfusion can be problematic for the hypovolemic donor who develops oligohydramnios, but the recipient often experiences greater difficulties with hyperbilirubinemia, hypervolemia, and hyperviscosity that arise from the increased RBC mass. Cardiac, neurologic, and developmental disorders have been associated with TTTS.

Alloimmune Hemolytic Anemia

Alloimmune hemolytic disease of the newborn (HDN) (see Chapter 23 ) is caused by the destruction of fetal or neonatal RBCs by maternal immunoglobulin G (IgG) antibodies. Alloimmune HDN involves the major blood group antigens of the Rhesus (Rh) and the A, B, AB, and O systems, but minor blood group incompatibilities (Kell, Duffy, MNS, and P systems) can also be associated with clinically significant disease. Particularly for Rh (D), anti-c, anti-E, and anti-K (Kell) antibodies, intrauterine or direct fetal transfusion may be indicated prenatally, and exchange transfusion may be necessary postnatally. Maternal IgG antibodies can cross the placenta, enter the fetal circulation and cause hemolysis, anemia, hyperbilirubinemia, and hydrops fetalis. In utero, the process is called erythroblastosis fetalis , whereas postnatally it is called hemolytic disease of the newborn (HDN). Maternal sensitization occurs through a prior transfusion of incompatible RBCs after fetal–maternal hemorrhage of incompatible fetal RBCs or from production of a pre-existing antibody developed against antigens from bacteria, viruses, or food. Transfer of antibodies across the placenta depends on the F _c component of the IgG molecule. Because both IgM and IgA antibodies lack this component, only IgG antibodies cause hemolytic disease of the newborn. Immunoglobulin G ₁ antibodies cross the placenta earlier and are responsible for more prenatal hemolysis. Immunoglobulin G ₃ antibodies cross the placenta later in gestation and are responsible for more severe hemolysis postnatally.

Other Immune Hemolytic Anemias

The other immune hemolytic diseases of early childhood are relatively rare. Autoimmune hemolytic anemia occurs, as do anemias associated with infections, drugs, and immunodeficiency syndromes. IgG antibodies, with or without complement, are often directed against one of the Rh erythrocyte antigens. These antibodies are most active at 37°C and often are called warm autoantibodies . The IgG-coated cells, with or without the assistance of complement, are cleared by the spleen and, to a lesser extent, by the liver. Congenital infections (syphilis, cytomegalovirus, rubella, toxoplasmosis, herpes, HIV, hepatitis) and acquired infections can cause hemolytic anemia and bone marrow suppression with reticulocytopenia (see Chapter 48, Chapter 49, Chapter 50 ). Immunoglobulin M autoantibodies can cause disease and are usually referred to as cold agglutinins , because they are most active at between 0°C and 30°C. These antibodies, with complement, coat RBCs and are usually cleared in the liver. Intravascular hemolysis is less common. The best-known causes of cold hemagglutinin disease are Mycoplasma pneumoniae , adenovirus, and Epstein-Barr virus. Immunoglobulin A–mediated hemolysis is quite rare but is remarkable for its severity ( Table 79.10 ).

The natural history of autoimmune hemolytic disease in infancy is that of a rapid onset of anemia with hyperbilirubinemia, splenomegaly or hepatomegaly, and hemoglobinuria (with intravascular hemolysis). Initially, reticulocytopenia may be noted, especially if antibodies are directed against RBC progenitors in the bone marrow, but a brisk reticulocytosis usually follows. Resolution within 3-6 months is common. Anemia with reticulocytosis and spherocytosis may be seen on the peripheral blood smear, but the diagnosis depends on demonstration of antibody-coated cells, with or without complement, in the DAT and antibodies in the serum by the IAT. Therapy includes treatment of underlying infection, removal of the offending drug, and supportive measures to limit hyperbilirubinemia. Response to corticosteroids is common, as is complete recovery. Steroids are less effective in IgM-mediated disease. A subset of patients younger than 2 years, and with a slower onset of disease at presentation, develops chronic hemolytic anemia. Difficulties with identifying compatible blood during a hemolytic crisis, as well as the usually self-limited nature of the disease, restrict blood transfusions to cases in which severe anemia impairs tissue oxygenation.

Erythrocyte Structural Defects

During its lifetime, a normal RBC traverses the circulation thousands of times, enduring tremendous mechanical and metabolic stress with each passage. The smallest capillaries have an internal diameter that is smaller than the diameter of the RBC; thus the blood cells must deform to squeeze through the capillaries and then return to their normal shape as they enter distal venules. During their rapid transit, the RBCs must endure tremendous fluctuations in pH, P o ₂ , and osmotic pressure. The lipid bilayer that forms the cell membrane is the site of numerous biologic functions mediated by integral proteins. It also is the attachment site for the proteins of the cytoskeleton, which confer the shape and stability necessary for proper membrane function. Some abnormalities in the membrane-cytoskeleton unit result in morphologic defects of RBCs ( Fig. 79.6 ). Because abnormally shaped RBCs are removed from the circulation by the reticuloendothelial system, many of these defects cause some degree of hemolytic anemia. Analysis of these congenital defects has led to an understanding of the erythrocyte membrane and cytoskeleton. The more common erythrocyte cytoskeletal defects originally were described by morphologic and clinical criteria. With better understanding of the cytoskeleton at a molecular level, it became clear that various mutations in genes coding for a few structural proteins are responsible for a family of inherited hemolytic anemias with overlapping morphologic, clinical, and genetic features. Mutations in the genes encoding α-spectrin, β-spectrin, ankyrin, protein 4.1, glycophorin C, protein 4.2, and band 3 have been reported in patients with hereditary defects of the red blood cell membrane/cytoskeleton. Inherited hemolytic anemias caused by RBC membranopathy, enzymopathy, and hemoglobinopathy are generally diagnosed by using conventional methods, including RBC morphology, membrane protein analysis, Hb electrophoresis, and measurement of RBC enzyme levels. In the genetic era, Sanger sequencing has been useful for detecting genetic mutations that cause inherited hemolytic anemias. Sanger sequencing is widely applied to identify disease-causing mutations in cases where traditional testing has failed or when a patient has been extensively transfused, leading to confounding biochemical and other testing findings caused by mixed RBC populations. Recent advances in genetic technology using next generation sequencing have enabled better identification of various genetic mutations that can cause these conditions.

Membrane defects associated with hemolytic anemia may present in the neonate with hepatosplenomegaly, hyperbilirubinemia, and hemolytic anemia. Aplastic crisis, attributed to drugs or an infectious agent such as parvovirus B19, or splenic sequestration, may cause life-threatening anemia in patients with hemolytic anemia. The more severe cases of hemolytic anemia can require splenectomy, although surgery generally is delayed beyond the first few years of life.

Hereditary Elliptocytosis and Hereditary Pyropoikilocytosis.

The findings on the peripheral blood smears in hereditary elliptocytosis and hereditary pyropoikilocytosis are distinct ( Fig. 79.7 ). Hereditary elliptocytosis (HE) is an autosomal dominant condition characterized by elliptical RBCs called ovalocytes, which is more common in people of African and Mediterranean origin. In hereditary pyropoikilocytosis (HPP), the RBC morphology is bizarre, marked by fragments, elliptocytes, and spiculated cells. The disorders are caused by defects in cytoskeletal proteins, most commonly α- or β-spectrin, which weaken the horizontal interactions necessary for cytoskeletal stability and reversible deformability (see Fig. 79.6 ). The deformed RBCs are prematurely cleared by the reticuloendothelial system. Hemolysis in HE generally is milder than in hereditary spherocytosis. The majority of those with HE are symptomatic. Hereditary pyropoikilocytosis is caused by homozygous or compound heterozygous defects. Some neonates with HE may exhibit hemolysis with HPP morphology on blood smear, but later in life their red blood cells will appear as typical elliptocytes. Hereditary pyropoikilocytosis is associated with more severe hemolysis and hyperbilirubinemia, often requiring exchange transfusion or phototherapy in the neonatal period. Examination of blood smears from each parent may be helpful in making this diagnosis.

Southeast Asian ovalocytosis (SAO), or stomatic elliptocytosis, is a mild hemolytic variant of HE, which is common in Southeast Asia, likely because it confers malaria resistance. Defects in band 3 are the most common.

Spherocytic elliptocytosis , characterized by spherocytes and elliptocytes and a mild hemolysis, occurs in those of European ancestry.

Membrane Lipid Defects

Membrane lipid abnormalities can be inherited but most often are acquired. Their presence, detected by morphologic abnormalities on the peripheral blood smear, is most important as the signal of an underlying disease. Target cells form when the surface-to-volume ratio of the red blood cell increases, either because of poorly hemoglobinated cells (iron deficiency, thalassemias, Hb C) or because lipids are added to the RBC membrane (liver disease with intrahepatic cholestasis, lecithin-cholesterol acyltransferase deficiency). Acanthocytes form when the membrane lipid composition between the inner and outer leaflets of the bilayer is altered, as in liver disease and abetalipoproteinemia.

Inherited Enzymatic Defects

The mature RBC, lacking a nucleus, polyribosomes, and mitochondria with which to carry out protein synthesis, must circulate through extremes of pH, P o _2, and osmotic gradients while maintaining deformability and integrity. Red blood cells metabolize glucose through the Embden-Meyerhof pathway and hexose monophosphate shunt to provide energy for maintaining ionic pumps, to reduce methemoglobin, and to synthesize small molecules such as adenine, guanine, pyrimidine nucleotides, glutathione, and lipids. The most commonly defective enzyme, glucose-6-phosphate dehydrogenase (G6PD), has been extensively evaluated at the clinical, biochemical, and molecular level. Many other enzymatic defects have been reported, but the rarity of the disorders has hindered investigation. Enzymatic defects of the RBC known to be associated with hemolytic anemia are listed in Box 79.3 . A few of the enzyme deficiencies cause abnormalities in other tissues. The end result of these defects is hemolytic anemia of varying severity, sometimes called hereditary nonspherocytic anemia . The RBCs may be morphologically normal and usually produce normal results on osmotic fragility testing, but they have a shorter life span. The milder cases cause little difficulty in the neonatal period. Some defects are associated with chronic and/or severe hemolysis, necessitating intermittent or chronic blood transfusions.

Glucose-6-phosphate dehydrogenase deficiency is the only common defect of RBC enzymes and can be associated with severe hemolysis, anemia, and hyperbilirubinemia in the neonate, although many are asymptomatic. This enzymatic defect is common in infants with bilirubin encephalopathy. Occasionally, the hyperbilirubinemia is severe, because of an interaction of G6PD deficiency with the (TA) ₇ promoter polymorphism of the UDP-glucuronosyltransferase 1A1 (UGT1A1) bilirubin-conjugating enzyme. The prevalence among some ethnic groups suggests a selective advantage, because deficient cells are resistant to malaria. As an enzyme in the pentose phosphate pathway, G6PD supplies NADPH, and by producing hydrogen ions, promotes reduction of glutathione. Deficiency limits the cell's ability to recover from oxidative stress. The gene is inherited as an X-linked recessive. Males are affected most commonly. Females may also be affected, either because of homozygosity or compound heterozygosity, or in cases in which X-inactivation is unbalanced. In the United States, African-American males are the most commonly affected. The A-minus variant, common in Africans, results in a mild reduction in both catalytic activity and stability. The B variant, common among Mediterranean, some Asian, and Ashkenazi Jewish populations, involves a severe reduction of enzyme activity, resulting in chronic, moderate-to-severe hemolytic anemia that could prove fatal in the face of severe oxidant challenge. In affected patients, the oldest RBCs are most deficient, resulting in a normal or minimally shortened erythrocyte life span, but exposure to oxidant drugs and toxins or a febrile episode can precipitate severe hemolysis. Hemolysis occurs within 24-48 hours of exposure and is accompanied by abdominal pain, vomiting, diarrhea, low-grade fever, jaundice, splenomegaly, hemoglobinuria, and anemia.

Neonates may present with hemolytic anemia and hyperbilirubinemia within 24 hours of life and are susceptible to developing methemoglobinemia. Management involves avoidance of precipitating agents, hydration, phototherapy and, if needed, transfusion (partial volume exchange or simple). Common drugs to be avoided or used with caution in this disorder include antimalarials, sulfonamides and sulfones, nitrofurans, anthelminthics, ciprofloxacin, methylene blue, acetaminophen, aspirin, and vitamin K analogues. An excellent resource for complete food and drug lists, as well as family education, is at https://www.g6pd.org (accessed May 4, 2018). Enzyme levels are highest in younger cells. Evaluation immediately after a hemolytic episode could be inconclusive because the older, abnormal cells have been destroyed. Repeat quantitative testing at a later time, as well as evaluation of maternal enzyme levels, can be useful. Some states and countries include GPD deficiency screening as part of their newborn screening program, using biochemical qualitation, biochemical quantitative assays, and DNA-based PCR screening for common mutations.

Vitamin E Deficiency

Vitamin E is a fat-soluble antioxidant that reduces the peroxidation of polyunsaturated fatty acids by reactive oxygen species during oxidative enzyme activity. Preterm infants and infants with low birth weight have low serum and tissue levels of this vitamin. Patients who have chronic fat malabsorption are the most likely to develop symptomatic vitamin E deficiency. A deficiency state in preterm infants and those with very low birth weight has been described, characterized by hemolytic anemia, reticulocytosis, thrombocytosis, chronic lung disease, intracranial hemorrhage, and retinopathy of prematurity. However, present evidence does not support routine supplementation with intravenous or high-dose vitamin E in neonates.

α-Thalassemia

Because the production of α-globin chains predominates from midgestation onward, defects in synthesis can be detected at birth. Before the debut of molecular analysis, the various α-thalassemia syndromes were assigned according to clinical criteria ( Table 79.12 ). Hydrops fetalis, the most severe and rarest form of α-thalassemia, occurs when an infant inherits deletions of both α-globin genes from each of the parents (see Chapter 23 ). Such an infant produces small quantities of Hb Portland to maintain in utero viability. The excess γ and β chains form tetramers with themselves, resulting in functionally useless Hb Barts (γ ₄ ), present in the newborn, and Hb H (β ₄ ) present in the older infant. The fetus with four α-globin gene defects experiences congestive heart failure in utero secondary to severe anemia and, in the absence of fetal transfusion, becomes grossly hydropic (hydrops fetalis). There is evidence of extensive extramedullary hematopoiesis and placental hypertrophy. Most are stillborn at 30-40 weeks’ gestation or die shortly after delivery. Infants with hepatosplenomegaly, jaundice, and a moderate hypochromic, microcytic hemolytic anemia should be evaluated for deletion of three α-globin genes, Hb H disease. At birth, large amounts of Hb Barts may be seen as well as some Hb H. Incubation of the RBCs with brilliant cresyl blue reveals many small inclusions. Treatment is supportive and includes supplementation with folic acid, avoidance of oxidant drugs, transfusion of RBCs, and attention to iron hyperabsorption and transfusional iron overload. Hypersplenism, characterized by leukopenia, thrombocytopenia, and worsening anemia, may necessitate splenectomy, although this usually is deferred beyond the first few years of life and is associated with infectious risk as well as increased thrombosis.

α-Thalassemia trait is characterized by a mild anemia with microcytosis, hypochromia, and erythrocytosis. Hemoglobin Barts is mildly elevated at birth. After the neonatal period, there is no simple laboratory evaluation that is diagnostic for α-thalassemia trait. Alpha globin gene analysis can be ordered to confirm clinical suspicion. The diagnosis usually is made in an iron-replete patient with a normal hemoglobin electrophoresis and appropriate family history for α-thalassemia. As the name implies, silent carriers of α-thalassemia could have slightly microcytic RBCs but usually are asymptomatic. Several states now use molecular testing for HBA1 and HBA2 , the genes coding for α ₁ - and α ₂ -globin, respectively.

β-Thalassemia

β-Thalassemia is not usually diagnosed at birth unless blood loss or RBC destruction has created an unusually high demand for replacement of fetal RBCs with adult. The disease generally manifests after 6 months of age, when a microcytic anemia persists beyond the time course for physiologic anemia. The more severe manifestations of the disease traditionally were called Cooley anemia , but these have been separated into thalassemia major and thalassemia intermedia (see Table 79.12 ). Affected patients exhibit splenomegaly, poor growth, microcytic hemolytic anemia with ineffective erythropoiesis, target cells on the peripheral blood smear, and extramedullary expansion of erythropoiesis. An abnormal hemoglobin analysis reveals elevations of Hb F and Hb A ₂ and a decrease in Hb A.

Patients are categorized as having thalassemia intermedia if they are able to maintain a hemoglobin level greater than 7 g/dL without routine transfusion. Thalassemia major patients require regular transfusions and iron chelation therapy to maintain a hemoglobin value adequate for growth and development. Iron overload is a significant cause of morbidity and mortality starting in the second decade of life. Evidence of hypersplenism may necessitate splenectomy. Bony deformities from expansion of extramedullary hematopoiesis occur in incompletely transfused patients.

β-Thalassemia trait is characterized by abnormalities on the blood smear: microcytosis, hypochromia, erythrocytosis, and the appearance of elliptocytes and target cells. Hemoglobin analysis reveals a mild elevation of Hb A ₂ and Hb F, although this is not observed if the genetic mutation also interferes with production of δ- or γ-globin or if Hb A ₂ and Hb F production are depressed because of coexistent iron deficiency anemia. The silent carrier state of β-thalassemia is associated with normal findings on the peripheral blood smear and hemoglobin electrophoresis and usually is identified through family studies of patients with a more severe β-thalassemia syndrome.

Hemoglobin E

Hemoglobin E causes one of the most common hemoglobinopathies in the world. This thalassemia syndrome, characterized as a qualitative and quantitative hemoglobinopathy, is particularly abundant among people of Southeast Asian ancestry, especially individuals from Laos, Cambodia, and Thailand. Hemoglobin E is generated by a single nucleotide substitution, Glu26Lys, in the coding region of the β-globin gene. The β-globin monomers associate normally with a pair of α-globin proteins to form a functional hemoglobin tetramer with essentially normal stability and oxygen-dissociation characteristics. Persons with Hb E are anemic because the mutation creates a cryptic splice site in the β-globin message. During RNA processing, a portion of the message is spliced into an abnormal, unstable transcript. Consequently, the total level of β-globin messenger RNA is reduced, resulting in a thalassemia phenotype. Hemoglobin E trait results in mild microcytosis. Those who are homozygous for Hb E have chronic, mild to moderate, microcytic anemia but are otherwise well; however, co-inheritance of an Hb E allele from one parent and a β-thalassemia allele from the other produces a moderate to severe anemia that can be transfusion dependent. Hemoglobin levels in these patients range from 2-7 g/dL, and children with this condition can develop hepatosplenomegaly and growth failure. Hemoglobin E-β ⁰ thalassemia disease now is the most common form of transfusion-dependent thalassemia in the United States and other parts of the world. Hemoglobin E co-inherited with Hb S results in a sickle phenotype.

Inefficient erythropoiesis is the main mechanism underlying the complications in thalassemia major, such as anemia, splenomegaly, bone malformations, and pulmonary hypertension. New knowledge has become available on pathways that contribute to the pathogenesis and clinical manifestations of this disease. Clinical trials with JAK2 inhibitors (ruxolitinib) and TGF-β family member inhibitors, sotatercept and luspatercept, are currently studying the benefits of these agents in decreasing ineffective erythropoiesis and splenomegaly.

Another modality of treatment for hemoglobinopathies is gene therapy, which is showing its safety and potential efficacy in both preclinical and early clinical studies. Gene therapy is now a reality, with seven patients cured of their β0/βE thalassemia or with significant amelioration from β0/β0 thalassemia and one patient with SCD, while others are showing modest transgene expression.

Unstable Hemoglobins

Most mutations causing unstable hemoglobin affect β-globin and involve amino acid substitutions in hydrophobic residues around the heme pocket. These amino acid replacements alter the hydrophobic interior of the molecule, predisposing the hemoglobin to instability and precipitation as small round inclusions known as Heinz bodies. Because the amino acid substitutions often involve replacing one neutral amino acid with another, the hemoglobin electrophoretic mobility may not change. Patients usually develop anemia and jaundice in late infancy as β-globin synthesis increases. Mutations in fetal or embryonic globins can cause symptoms at birth but not later in life. The diagnosis is confirmed by supravital staining for Heinz bodies, which are usually present, and hemoglobin analysis after isopropanol precipitation.

Sickle Cell Anemia

Sickle cell disease is a common medical problem in the United States, where 1 in 400 newborns is affected. By definition, sickle cell anemia refers to the doubly heterozygous inheritance of abnormal genes that code for the substitution of valine in place of glutamic acid at position six in the β chain of hemoglobin (Hb SS). There are also a number of sickle hemoglobinopathies in which one gene coding for Hb S is inherited along with a second abnormal β-globin gene coding for other hemoglobins, such as Hb C, D, O ^Arab , or β-thalassemia. The clinical course of affected patients may be indistinguishable from, or milder than, those of Hb SS disease. As with the thalassemias, prenatal diagnostic techniques are available. Newborn screening programs have been instituted in many parts of the world, including most of the United States. Samples of cord or capillary blood are subjected to testing, usually hemoglobin electrophoresis, high pressure liquid chromatography, or isoelectric focusing. Infants with abnormal results on the newborn screen are referred for confirmatory testing and disease counseling ( Fig. 79.8 ; Box 79.5 ).

The clinical hallmark of Hb SS disease is hemolytic anemia with reticulocytosis and the appearance of irreversibly sickled cells on the peripheral blood smear. The onset of signs and symptoms correlates with the decrease in Hb F levels and increasing expression of the mutant β-globin gene products during the first year of life. Neonates generally are asymptomatic, although older infants are at risk for several disease-related complications. Because of splenic dysfunction, infections with certain bacteria, such as pneumococcus, are a major cause of mortality and morbidity in infants and young children. A suddenly pale, listless infant may be experiencing life-threatening anemia caused by hepatic or splenic sequestration. Aplastic crisis, triggered by infection or drugs, also may occur. Vaso-occlusive crisis of the infant's hands and feet, the hand–foot syndrome (dactylitis), manifests with pain, warmth, and swelling of the hands and/or feet.

Identification of affected newborns through newborn screening programs with enhancements in parent education and supportive care such as penicillin prophylaxis, immunization against S. pneumoniae and H. influenzae , and early identification of hepatic or splenic sequestration have decreased the death rate for infants and children. The natural history of sickle cell anemia includes painful vaso-occlusive events, acute and chronic lung disease, stroke, overwhelming sepsis from splenic dysfunction, avascular necrosis of joints, retinopathy, and transfusion-related iron overload. The National Heart, Lung and Blood Institute (NHLBI) issued management guidelines for sickle cell disease in 2002, which are available at https://www.nhlbi.nih.gov/files/docs/guidelines/sc_mngt.pdf . Treatment of the sickle hemoglobinopathies has been largely supported using penicillin prophylaxis; vaccinations; blood transfusions with extended RBC phenotyping; cross-matching to decrease the incidence of transfusion-associated alloimmunization; and hydration, narcotics, and nonsteroidal anti-inflammatory drugs for painful crises. Hydroxyurea decreased the number and severity of vaso-occlusive pain events in infants and young children with Hb SS and Hb SB ⁰ thalassemia in the randomized, placebo-controlled BABY HUG study. Stem cell transplantation, the only established cure for sickle cell anemia, has been limited by the availability of suitable donors and the uncertainty in predicting which children will express a more severe phenotype. Ongoing studies using reduced intensity conditioning regimens and alternative donor options, as well as risk stratification of sickle cell patients, may allow larger number of patients to undergo stem cell transplantation in the future.

Sickle Cell Trait

Heterozygotes for Hb S (sickle cell trait) were thought to be clinically unaffected, but there are reports of hematuria, decreased urinary concentrating ability, and occasional reports of sudden death related to high altitude and extreme exercise. Associations have been made between sickle cell trait and venous thrombosis, renal medullary carcinoma, and a more severe course of diabetes mellitus.

Iron Deficiency Anemia

Although the prevalence of iron deficiency anemia is decreasing in the United States, it remains a major cause of anemia in infants and children, and it has been associated with adverse neurocognitive outcomes. Factors associated with the declining rates of iron deficiency anemia include the increasing number of infants receiving breast milk, the use of standard iron-supplemented infant formulas and iron-supplemented baby cereals during the first year of life, and the delay of the introduction of cow milk into the infant diet until after 1 year of age.

Iron is an essential nutrient. Most iron is bound to the heme proteins, hemoglobin, and myoglobin. The storage proteins ferritin and hemosiderin contain most of the rest of the iron. A small percentage is bound in enzyme systems such as cytochromes and catalase. In healthy adults, most of the body's iron is recycled from the breakdown of RBCs. Little iron needs to be absorbed from the intestines, and little is lost through fecal or urinary excretion. Because neonates must rapidly expand muscle mass and blood volume, they require a much higher percentage of dietary iron intake. The amount of iron stored in the neonate's body is proportional to birth weight. Eighty percent of a term infant's iron is acquired during the third trimester. For a full-term infant, this should be sufficient for the first 4-6 months of life. Preterm infants and those born small for gestational age have a much faster rate of postnatal growth and are at risk for iron deficiency within the first 3 months of life ( Box 79.6 ). Infants born anemic and those who have accelerated iron losses from repeated laboratory testing, trauma, surgery, or anatomic abnormalities may also become deficient. Although both breast milk and cow milk contain iron, the bioavailability of the iron in breast milk is much superior. The American Academy of Pediatrics (AAP) Committee on Nutrition updated its recommendations for diagnosing and preventing iron deficiency and iron deficiency anemia in 2010 ( Table 79.13 ). Full-term infants who are breastfed should not require iron supplementation until 4-6 months of age. At 4 months of age, human milk–fed infants should receive iron supplementation until they are eating iron-containing foods. Two daily servings of an iron-fortified cereal will meet this requirement. Infant cereal fortified with iron should be one of the earliest solid foods introduced. After 6 months of age, one daily serving of a vitamin C–rich food should be introduced. Because the protein in cow milk can cause occult gastrointestinal bleeding in the neonate and because of the poor bioavailability of iron in cow milk, it is recommended that babies avoid cow milk until after 1 year of age. In general, preterm infants who are fed human milk require iron supplementation after 1 month of age, either by a supplement added to the breast milk or iron given directly to the infant. An exception may be made for infants who have received multiple blood transfusions and are thus at risk for iron overload rather than deficiency. A thought-provoking, randomized, placebo-controlled trial of daily oral iron supplementation in very low birth weight infants (<1500 g) of less than 32 weeks’ gestational age, who were managed on a liberal transfusion strategy, did not demonstrate an increase in the hematocrit, reticulocyte count, or transfusion requirements at week 36 postmenstrual age.

With improved survival in the smallest patients, additional knowledge gaps and opportunities for investigation are identified in diagnosing, treating, and preventing iron deficiency and iron deficiency anemia ( Box 79.7 ). Although severe iron deficiency in children is fairly simple to diagnose, milder deficiency may not yet have caused anemia or may be difficult to distinguish from other causes of microcytic anemia, especially in patients with chronic or acute illness. The AAP recommends that testing for anemia by hemoglobin estimation should occur at approximately 12 months of age. In addition, there should be an assessment of risk factors for iron deficiency/iron deficiency anemia at that time. Earlier and/or more frequent assessments should occur in situations of high concern. A careful dietary history should be taken, and consideration should be given to special circumstances such as ongoing blood loss, malabsorption, early or excessive cow milk intake, and birth history. In early iron deficiency, the bone marrow stores are depleted and the RBC distribution width increases. Subsequently, the iron transporter levels fall, resulting in lower serum levels of iron, ferritin, and transferrin in the term infant. Finally, erythrocyte production is affected. A hypochromic, microcytic anemia becomes apparent. Screening test results such as serum iron, total iron-binding capacity, ferritin, and transferrin saturation may be inconclusive because of acute or chronic illness. It is unclear what testing is optimal in the premature infant. The AAP favors the reticulocyte Hb concentration (CHr) and the serum transferrin receptor 1 (TfR1), because these tests are not affected by inflammation or infection, but these tests are not widely available. A therapeutic trial of iron for presumptive iron deficiency anemia is a cost-effective strategy. Ferrous sulfate, in a dose of 2-6 mg/kg per day of elemental iron, may be given for 1 month. If the hemoglobin rises by at least 1 g/dL (in the absence of ongoing blood loss), the diagnosis of iron deficiency is made, and the iron supplementation is usually continued for 2-3 more months or until 1-2 months after the hemoglobin is in the normal range. Screening for lead exposure should be done as part of routine well-baby care and particularly considered if a hypochromia and microcytosis are detected.

Iron Transport Proteins and Iron Homeostasis

Our knowledge of iron biology has been enriched substantially by the recent discovery of mammalian iron transport proteins (DMT1, ferroportin, mitoferrin 1, and ZIP14), heme transporters (HRG1and FLVCR1), iron chaperones (PCBPs), ferrireductases and ferroxidases (DCYTB, STEAP3, PrPc, and hephaestin), and the iron-regulatory hormone hepcidin. The physiologic function of these proteins has been studied to a great extent and the implications of their up-and-down regulation.

We have learned in depth the mechanisms through which iron is transported in the gut. Iron is transported out of the enterocyte and into portal blood via ferroportin (SLC40A1) located on the basolateral membrane. Ferroportin transports only Fe2+, whereas transferrin in portal blood will bind only Fe3+. Efficient transfer of iron to portal blood transferrin is thought to involve an oxidation step catalyzed by a ferroxidase. The best characterized intestinal ferroxidase is hephaestin, a membrane-anchored homologue of the plasma ferroxidase, ceruloplasmin.

Iron Metabolism in Erythrocyte Precursors

Greater than 95% of iron in plasma is bound to its circulating transport protein transferrin, which delivers most of its iron to erythrocyte precursors (i.e., erythroid progenitor cells of the bone marrow that differentiate into mature RBCs). Each day, ~25 mg of iron is taken up into these cells to support the daily production of 200 billion new RBCs. Erythrocyte precursors take up iron nearly exclusively from transferrin via transferrin receptor 1 (TFR1). The quantitative importance of this uptake is illustrated by the fact that an estimated 80% of total body cellular TFR1 is located in the erythroid marrow of an adult human. When transferrin binds to TFR1 at the cell surface, the complex is internalized into endosomes, which become acidified, causing iron to dissociate from transferrin. The endosomal ferrireductase STEAP3 (six-transmembrane epithelial antigen of the prostate 3) reduces the liberated Fe3+ to Fe2+, which is subsequently transported into the cytosol via DMT1. Despite being the most avid consumers of iron in the body, erythrocyte precursors abundantly express ferroportin at the plasma membrane and are therefore able to export nonheme iron. Systemic iron depletion has been shown to increase ferroportin expression in erythrocyte precursors, leading to the hypothesis that upregulation of ferroportin in iron deficiency serves to provide iron to nonerythropoietic tissues.

The hepatically produced peptide, hepcidin, is an important regulator of iron homeostasis. It inhibits the transport of iron across the gut mucosa as well as the transport of iron out of macrophages. Hepcidin is an acute-phase reactant: in inflammation, increased hepcidin results in decreased iron absorption. Overexpression of hepcidin in genetically modified mice results in iron deficiency anemia. This has led to interest in human defects of hepcidin overexpression that may result in the inability to absorb iron (i.e., refractoriness to oral iron). Conversely, the intestinal hyperabsorption of iron in thalassemias is associated with low hepcidin levels.

Physiologic Anemia of Infancy and Physiologic Anemia of Prematurity: The Role of Erythropoietin

The physiologic anemia of infancy can be exaggerated in the sick or premature infant by frequent blood sampling: the smaller the infant, the proportionally greater the volume of blood that is withdrawn for laboratory testing. Most RBC transfusions in neonates occur within the first 3-4 weeks of life, with the majority being in the first 2 weeks. Physiologically lower levels of EPO in neonates provided the rationale for the pharmacologic use of erythropoietin to reduce the volume and risks of blood transfusions. Recombinant erythropoietin products are commercially available and have been used for decades in the treatment of adults with renal disease and in cancer-associated anemia. The Food and Drug Administration (FDA) mandated black box warnings for EPO because of increased risks of death, myocardial infarction, stroke, venous thrombosis, and cancer progression in adult patients. These complications have not been seen in neonates. Two approaches to EPO therapy have been systematically reviewed in neonates and reported in a 2017 Cochrane database systematic review. The early approach, defined by the use of EPO before day 8 of life, has been shown to reduce (but not eliminate) RBC transfusions and donor exposures but did not impact morbidity and mortality measures. The later use of EPO (on day 8 of life or after) resulted in a reduction in the number of blood transfusions but not the total volume of blood, per infant, so that meaningful clinical outcomes were not affected by the use of EPO. Because most infants received RBC transfusions very early and before being enrolled in the clinical trials evaluating this question, the authors concluded that the critical effort should be aimed at preventing donor exposures in the very first few days of life. Stable and larger preterm infants have a better response to EPO therapy when compared with ELBW infants. Erythropoiesis requires iron. Even though EPO administration has been shown to reduce hepcidin levels, thereby increasing intestinal iron absorption, ferritin levels have been reported to drop with EPO use. Supplementation of between 1 mg/kg per day and 10 mg/kg per day elemental iron has been used to lessen the risk of iron deficiency.

Transient Erythroblastopenia of Childhood

Transient erythroblastopenia of childhood (TEC) is an acquired, transient, normocytic, hypoplastic anemia with a peak incidence at ages 2-3 years. It is exceedingly rare in neonates. The etiology of TEC is unknown but may be the result of viral injury to erythroid precursors. Familial cases have been reported. The degree of anemia can range from moderate to severe. Transient erythroblastopenia of childhood is a diagnosis of retrospect. Treatment consists of supportive measures, including RBC transfusion if needed, until bone marrow recovery. Most patients recover in 1-2 months. The condition may be confused with Diamond-Blackfan anemia and parvovirus-induced anemias.

Parvovirus-Induced Anemia

Parvovirus B19, which proliferates in human erythroid precursors, causes several diseases. The majority of adults in the United States have antibodies to the virus. The virus produces fifth disease. The rash and joint symptoms associated with this generally minor illness are caused by vascular deposition of antibody complexes generated in response to the infection. In patients with underlying hemolytic anemias, parvovirus infection can cause a transient, severe aplastic crisis. It can also cause chronic anemia from a persistent infection in erythroblasts in immunocompromised hosts. Chronic parvovirus anemia responds to administration of intravenous immunoglobulin. The virus binds to blood group P-antigen on the surface of erythroid precursors, is internalized, and then replicates, disrupting normal erythroid differentiation. Endothelial cell P-antigen is postulated to participate in placental transmission of the virus. This antigen is also found on fetal cardiomyocytes, a finding consistent with the fact that the fetus infected with parvovirus can develop myocarditis. Infection rate during pregnancy is estimated to be in the 3% range. Parvovirus infection during pregnancy can result in anemia, hydrops fetalis, fetal loss, or congenital infection. The rate of transplacental transmission of the virus has been estimated at 33%. Initial reports of high rates of stillbirth and fetal loss have been modified; it is recognized that most of the mortality occurs during the first 20 weeks of gestation. Severe thrombocytopenia occurs in more than one-third of cases and can complicate intrauterine transfusion. Parvovirus-induced hydrops can be detected by ultrasound, and treatment of suspected hydropic infants includes intrauterine RBC transfusion. Parvovirus infection of the fetus may persist after birth as a cause of congenital RBC aplasia.