PART XX Diseases of the Blood OUTLINE Section 1 The Hematopoietic System Section 2 Anemias of Inadequate Production Section 3 Hemolytic Anemias Section 4 Polycythemia (Erythrocytosis) Section 5 The Pancytopenias Section 6 Blood Component Transfusions Section 7 Hemorrhagic and Thrombotic Diseases Section 8 The Spleen Section 9 The Lymphatic SystemSECTION 1 The Hematopoietic System OUTLINE Chapter 473 Development of the Hematopoietic System Chapter 474 The AnemiasCHAPTER 473 Development of the Hematopoietic System Stella T. Chou Hematopoiesis in the Human Embryo and Fetus Hematopoiesis is the process by which the cellular elements of blood are formed. In the developing human embryo and fetus, hematopoiesis has 3 developmental waves and is conceptually divided into 3 anatomic stages: mesoblastic, hepatic, and myeloid. Mesoblastic hematopoiesis occurs in extraembryonic structures, principally in the yolk sac, and begins between the 10th and 14th days of gestation. By 6-8 wk of gestation, the liver replaces the yolk sac as the primary site of blood cell production, and during this time the placenta also contributes as a hematopoietic site. By 10-12 wk, extraembryonic hematopoiesis has ceased. Hepatic hematopoiesis occurs through the remainder of gestation and then diminishes during the second trimester while bone marrow (myeloid ) hematopoiesis increases. The liver is the predominant erythropoietic organ through 20-24 wk of gestation. Each hematopoietic organ houses distinct populations of cells. The yolk sac predominantly produces erythrocytes, megakaryocytes, and macrophages. The fetal liver is primarily an erythropoietic organ, while the bone marrow produces erythrocytes, megakaryocytes, and leukocytes. The types of leukocytes present in the fetal liver and marrow differ with gestation. Macrophages precede neutrophils in the marrow, and the ratio of macrophages to neutrophils decreases as gestation progresses. Regardless of gestational age or anatomic location, production of all hematopoietic tissues begins with multipotent cells capable of both self-renewal and clonal maturation into all blood cell lineages. Progenitorcells differentiate under the influence of transcription factors and hematopoietic growth factors (Table 473.1 ). Table 473.1 Characteristics of Hematopoietic Growth Factors MOLECULAR CHROMOSOMAL GROWTH FACTOR Wass(kbay LOCATION PRINCIPAL TARGET CELL ERYTHROPOIETIN ] 30-39 Tqi-1z CFU, fetal BFU-E, endothelial calls, neurons, astrocytes, oligodendrocytes COLONY-STIMULATING FACTORS G-CSF TED TqliE2 CFU-G, CFU-MIX, mature neutrophils GM-CSF 18-30 3q2a31 CFU-MIX, CFU-GM, BFU-E, monocytes mature neutrophils NECSF F570 Dimer of | Sq ‘CFU-M, macrophages 2 subunits) SCF 36 TL az CFU-MIX, BFU-E, CFU-GM, mast calls TCFE 5 Homodimeric| 19q13.2 BL-CEC protein CFI 192 Amino acd | 1pI33 Monocytes, macrophages, dendritic cals, protein Langerhans cells, INTERLEUKIN tT 7 Alpha 2qi3 Beta] Hepatocytes, macrophages, lymphocytes 2g13-21 Lz 5-20 AqQo-27 T calls, cytotoxic Iymphocytes 1s 130 3q25-31 CFU-MIX, CFU-Meg, CFU-GM, BFU-E, macrophage 1 620 3qaT Tells, B cells, dendritic cells 1S (Diner of2 | Sq23-31 CFU-Eo, B calls subunits) 6 19-26 TpaEae CFU-MIX, CFU-GM, BFU-E, monocytes, EB calls, T cells, cytotoxic lymphocytes 17 Es Bqiz1 Bealls 1 aio aia Neutrophils, endothelial cals, T cells 1 16 Sqar-32 BFU-E, CFU-MIX 1-10 ie7 1q321 Macrophages; Iymphocytes Tr 2 Toqis CFU-Meg. B cells, Keratinocytes Tz 70-75 (Dimer of | p3s/p40 3 (pas) and 11 (p40) T cells, NK calls, 2 subunits) macrophages Taz g 3Q3T Pre-B lymphocytes macrophages Tad 3 Bealls USE} THIS Bells, Toalls Tis 2-14 Tealls Tar 20-30 Marrow svomal cells 1-18 2 CD4* Tells, NK cells Tar AQT Teals 1-25 Dimer of pIgIL-12pq0 Ce Tealis subunits 15 Tagiiz Tells monocytes, marrow Sromal calls 1-3 ‘FH bundle | 12q437 Tells, hematopoietic progenitors1-34 222 Amino acid | 16q22.1 Monocytes, macrophages protein THROMBOPOIETIN] 15-38 3qTe Magakaryocyte progenitors, megakaryocytes BFU-E, Burst-forming units-erythroid; BL-CFU, blast colony-forming cell; CFU-E, colony-forming units—erythroid; CFU-Eo, colony-forming units—eosinophil; CFU-G, colony-forming units— granulocyte; CFU-GM, colony-forming units-granulocyte-macrophage; CFU-M, colony-forming units-macrophage; CFU-Meg, colony-forming units~megakaryocyte; CFU-MIX, colony-forming units-mixed; CSF-1, colony-stimulating factor-1; G-CSF, granulocyte colony-stimulating factor, GM-CSF, granulocyte-macrophage colony-stimulating factor; IL, interleukin; M-CSF, macrophage colony-stimulating factor; NK, natural killer; SCF, stem cell factor; TGF-8, transforming growth factor-beta. ‘The classical model of hematopoietic differentiation involves differentiation into increasingly lineage-specific progenitors, although there may also be alternate pathways that are used separately or in combination with classical pathways (Fig. 473.1 ). In the classical pathway, long-term repopulating hematopoietic stem cells (LTR-HSCs) are characterized by their ability to self- renew and differentiate into cells that are multipotent. Multipotent progenitors (MPPs) have reduced self-renewal capacity and differentiate into common Iymphoid progenitors (CLPs) or common myeloid progenitors (CMPs). The CMP differentiates into all the blood lineages except for lymphoid. The commitment of hematopoietic cells to increasingly lineage-restricted cells requires cytokine stimulation and regulation by transcription factors.=a oe FIG. 473.1 Major eytokine sources and actions to promote hematopoiesis. Cells of the bone marrow microenvironment, such as macrophages, endothelial cells, and reticular fibroblasts, produce macrophage colony-stimulating factor (V-CSF), granulocyte ‘macrophage colony-stimulating factor (GM-CSF), and granulocyte colony-stimulating factor (G-CSF) after stimulation. These cytokines and others listed in the text have ‘overlapping interactions during hematopoietic differentiation, as indicated: for all lineages, optimal development requires a combination of early- and late-acting factors. BFU, Burst-forming unit; CFU, colony-forming unit; Epo, erythropoietin; MSC, myeloid ‘stem cells; PSC, pluripotent stem cells; SCF, stem cell factor; TNF, tumor necrosis factor; Tpo, thrombopoietin. (From Sieff CA, Daley GO, Zon Ll: The anatomy and physiology of hematopoiesis. In Orkin SH, Nathan DG, Ginsburg D, et al, editors: ‘Nathan and Oski's hematology of infancy and childhood , ed 8, Philadelphia, 2015, Elsevier) Erythrocytes in the fetus are larger than in adults, and at 22-23 wk gestation the mean corpuscular volume can be as high as 135 femtoliters (fL) (Fig. 473.2 , upper panel ). Similarly, the mean corpuscular hemoglobin is very high at 22-23wk and falls relatively linearly with advancing gestation (Fig. 473.2 , lower panel ). In contrast, the mean corpuscular hemoglobin concentration is constant throughout gestation at 34 +1 g/dL. While the size and quantity of hemoglobin in erythrocytes diminish during gestation, the hematocrit and blood hemoglobin concentration gradually increase (Fig. 473.3 , upper and lower panels , respectively). 140, 135 130] 125 120) 115 mev (fl) 110) 105 100 SSS 95 0 22°23 24 25 26 27 28 28.90 SI S299 04 9590 37 90940 AD Weeks Gest MCH (po) ——S Se 2225 24 25 25 27 2629 O'S 82 39S 36 7 GH BAO HT AD Weeks Gestation FIG. 473.2 Erythrocyte mean corpuscular volume (MCY, top ) and mean corpuscular hemoglobin (MCH, bottom ) from 22 wk gestation through term. The lines represent the ‘5th percentile, the mean, and the 95th percentile reference range. (From Christensen RD, Jopling J, Henry E, et al: The erythrocyte indices of neonates, defined using data from over 12,000 patients in a multihospital healthcare system, J Perinatol 28:24-28, 2008.)60 50 Hematocrit (%) & 20 40 A Time (wk) 6 s E64 g a 8 ee Bs = é 2 1 0 30 0 B Time (wk) FIG. 473.3 Reference ranges of fetal hematocrit (A) and fetal red blood cell count (B) by cordocentesis throughout gestations. Concentrations of platelets in the blood increase gradually between 22 and 40 wk gestation (Fig. 473.4 ), but the platelet size, assessed by mean platelet volume, remains constant at 8 +1 fL. No differences are observed between males and females in fetal and neonatal reference ranges for erythrocyte indices, hematocrit, hemoglobin, platelet counts, or mean platelet volume measurements.400,00, tee SPS ee aes ee ee ° Gestatonal age 22 25 24” 25" 26 27 28H) 31 BBs ww G7 a8 90 4 81 He ‘Sample Size 11 Ge (05 (0m as tga 216 364 317 G40 GHB 1Oa0 1708 2456 GaBs abtD 5178 7600 46s 1143.71 FG. 473.4. Platelet count from22 wk gest town ier Thelines represent the Sth percentile, the mean, andthe 86th percentile reference range. From Wedmeter SE, Henry € Sola Vianer My ta Platelet reference ranges or neonates, defined using data rover 47000 patent n'a matinosptalhealtnare system, 9 Perinat 29:130- 136, 2009) Fetal Granulocytopoiesis Neutrophils are first observed in the human fetus about 5 wk after conception as small clusters of cells around the aorta. The fetal bone marrow space begins to develop around the 8th wk, and from 8-10 wk the marrow space enlarges, but no neutrophils appear there until 10.5 wk. From 14 wk through term, the most common granulocytic cell type in the fetal bone marrow space is the neutrophil. Neutrophils and macrophages originate from a common progenitor cell, but macrophages appear before neutrophils in the fetus, first in the yolk sac, liver, Tung, and brain, all before the bone marrow cavity is formed. Granulocyte colony-stimulating factor (G-CSF) and macrophage colony- stimulating factor (M-CSF) are expressed in developing fetal bone as early as 6 wk after conception, and both are expressed in the fetal liver as early as 8 wk. Granulocyte-macrophage colony-stimulating factor (GM-CSF) and stem cell factor (SCF) also are distributed widely in human fetal tissues. However, no changes in expression of any of these factors, or of their specific receptors, appear to be the signal for fetal production of neutrophils or macrophages,because those signals have not yet been identified. Fetal blood contains few neutrophils until the third trimester. At 20 wk gestation the blood neutrophil count is 0-500/mm? . Although mature neutrophils are scarce, progenitor cells with the capacity to generate neutrophil clones are abundant in fetal blood. When cultured in vitro in the presence of recombinant G-CSF, they mature into large colonies of neutrophils. The physiologic role of G-CSF includes upregulating neutrophil production, and this appears to be the case for the fetus and neonate as well as for adults. Thus the low number of circulating neutrophils in the mid-trimester human fetus may be caused in part by low production of G-CSF. Monocytes isolated from the blood of adults produce G-CSF when stimulated with a variety of inflammatory mediators, such as bacterial lipopolysaccharide (LPS) or interleukin (IL)-1. In contrast, monocytes isolated from the blood or organs of fetuses up to 24 wk gestation generate only small quantities of G-CSF protein and messenger RNA (mRNA) after LPS or IL-1 stimulation. Despite this, G-CSF receptors on the surface of neutrophils of newborn infants are equal in number and affinity to those on adult neutrophils. In the fetus, actions of the granulocytopoietic factors (G-CSF, M-CSF, GM- CSF, and SCF) are not limited to hematopoiesis. Receptors for each of these are located in areas of the fetal central nervous system and gastrointestinal tract, where their patterns of expression change with development. Fetal Thrombopoiesis Several biologic differences exist between fetal/neonatal and adult megakaryopoiesis and thrombopoiesis. There is a developmentally unique pattern of fetal/neonatal megakaryopoiesis characterized by rapid proliferation, followed by full cytoplasmic maturation without polyploidization. Fetal and neonatal megakaryocytes are significantly smaller, exhibit lower ploidy, and produce fewer platelets. However, fetal and neonatal megakaryocytes have a higher proliferative potential than adult progenitors. These differences allow fetuses and neonates to populate their rapidly expanding bone marrow space and blood volume while maintaining normal platelet counts. Megakaryocyte progenitors are categorized as burst-forming unit— megakaryocytes (BFU-MK), which are primitive megakaryocyte progenitors, and colony-forming unit-megakaryocytes (CFU-MK), which are more differentiated. BFU-MK produce large multifocal colonies containing >50megakaryocytes, whereas CFU-MK generate smaller (3-50 cells/colony) unifocal colonies. Megakaryocytes are identified by their morphologic characteristics as they undergo endoreduplication, which results in large cells with polyploid nuclei. Megakaryocytes, unlike megakaryocyte progenitors, do not have the capacity to generate colonies. Rather, they undergo maturation, progressing from small mononuclear cells to large polyploid cells. The modal megakaryocyte ploidy (the number of sets of complete chromosomes) in normal adult marrow is 16N. In the fetus and neonate, ploidy is lower, primarily 2N and AN, and megakaryocyte size is smaller. Large megakaryocytes generate more platelets than do small megakaryocytes; it is assumed that megakaryocytes of neonates produce fewer platelets than do their adult counterparts. ‘The exact mechanisms by which megakaryocytes release platelets into the circulation remain incompletely understood. In situ examination of this process suggests that mature megakaryocytes migrate to a perivascular site and extend a process through the endothelium, giving rise to proplatelets, which then release platelets. An alternate mechanism is that platelets are released from megakaryocytes in the lungs as a result of shear forces. Thrombopoietin (TPO) is the dominant regulator of megakaryocyte development and platelet production (see Table 473.1 ). TPO is predominantly produced in the liver from early fetal to adult life but is also expressed by cells in the kidney, and to a lesser extent, by smooth muscle and marrow cells. TPO concentrations are higher in healthy neonates of any gestational age than in healthy adults. TPO is a primary stimulator of megakaryocyte and platelet production, but SCF, IL-3, IL-11, IL-6, and erythropoietin also stimulate megakaryopoiesis and thrombopoiesis in vitro and in vivo. Importantly, TPO also promotes expansion of hematopoietic stem cells (HSCs) and progenitor cells, and the TPO receptor is expressed on HSCs and erythroid progenitors in addition to megakaryocyte progenitors, megakaryocytes, and mature platelets. Fetal Erythropoiesis Similar to hematopoietic production of other cell lineages, fetal erythropoiesis is regulated by growth factors produced by the fetus, not by the mother. Erythropoietin (EPO) does not cross the human placenta. Stimulating maternal EPO production does not enhance fetal erythropoiesis, and suppressing maternal erythropoiesis by hypertransfusion does not suppress fetal erythropoiesis. EPO plays a central regulatory role on the proliferation and maturation oferythroid progenitors. Erythroid-committed progenitors consist of burst-forming unit-erythroid (BFU-E) and colony-forming unit-erythroid (CFU-E) cells. In colony-forming assays, human BFU-E are more proliferative, forming colonies of multiple clusters of erythroblasts, vs CFU-E, which form 1 or 2 clusters with each containing 8-100 hemoglobinized erythroblasts. EPO is essential for erythrocyte production from CFU-E cells by inducing survival and proliferation of erythroblasts. EPO binds to specific receptors on the surface of committed erythroid precursors, and its expression is regulated by an oxygen-sensing mechanism through the hypoxia-inducible factor (HIF) family of proteins. HIF- 1a and HIF-2a are regulated by oxygen tension, whereas HIF-1f is constitutively expressed. Together, HIF proteins maintain oxygen homeostasis and regulate erythropoiesis by inducing EPO under hypoxic conditions. EPO is produced by monocytes and macrophages in the fetal liver during the first and second trimesters. After birth, the anatomic site of EPO production shifts to the kidney. The specific stimulus for this shift is unknown but may involve the increase in arterial oxygen tension that occurs at birth. Epigenetic modification of gene expression may also play a role, since it appears that renal and hepatic EPO genes are methylated to different degrees. Although EPO mRNA and protein can be found in the human fetal kidney, it is not known whether this production is biologically relevant. It appears that renal production of EPO is not essential for normal fetal erythropoiesis, as evidenced by the normal serum EPO concentration and normal hematocrit of anephric fetuses. Hemoglobins in the Fetus and Neonate Hemoglobin is a tetramer of 4 globin chains with an iron-containing porphyrin ring called heme bound to each chain. A dynamic interaction between heme and globin gives hemoglobin its unique properties in the reversible transport of oxygen. The hemoglobin molecule consists of 2 alpha («)-like and 2 beta (B)-like polypeptide chains, with each chain having a heme group attached. The a-globin and B-globin gene clusters are located on chromosome 16 and 11, respectively (Fig. 473.5 ). There are 2 B-globin genes and 4 a-globin genes. Within erythrocytes of an early embryo, fetus, child, and adult, 6 different hemoglobins may normally be detected (Fig. 473.6 ): the embryonic hemoglobins (Gower-1, Gower-2, and Portland), fetal hemoglobin (HbF), and the adult hemoglobins (HbA and HbA, ). The electrophoretic mobilities of hemoglobins vary with their chemical structures.5 a iso TOTTI Tae Fetal: Hemoglobin a, v2 FIG. 473.5 Organization of the globin genes. The bottom line reflects the scale in Kdlobases. The upper segment represents the Bike globin genes on chromosome 11, {and the lower segment the alike genes on chromosome 16. Regions of the gene that code for primary globin proteins are shown as blue segments , and regions that code ‘for pseudogenes ("," nonexpressed remnants) are shown as pink segments . The composition of embryonic, fetal, and adult hemoglobins is listed. a, Alpha; B, beta; , ‘gamma; 6, delta; e, epsilon; Z, zeta.8 8 8 8 Hemoglobins: % of total Globin subunits: % of total > 6 T © aS rau mul 38 10 15 200m if FETUS NEWBORN FIG. 473.6 Changes in hemoglobin tetramers (A ) and in globin subunits (B)) during human development from embryo to early infancy. (From Polin RA, Fox WW: Fetal and neonatal physiology , ed 2, Philadelphia, 1998, Saunders, p 1769.) o Expression and quantitative relationships among the hemoglobins are determined by complex developmental processes. Globin chain expression is developmental stage specific and occurs through 2 hemoglobin switches, mediated primarily through changes of the B-globin genes expressed. There are 5 functional B-like globin chain genes: embryonic (HBEI ), 2 fetal (HBG1, HBG2 ), and 2 adult (HBD, HBB ); and 3 a-like globin chain genes: embryonic (HBZ ) and 2 adult (HBAL, HBA2 ). Primitive erythroid cells primarily express embryonic globins. The Ist B-globin switch occurs at approximately 6 wk gestation to fetal globin (HBG ), which coincides with the onset of definitive hematopoiesis. The major hemoglobin in the fetus (HbF) consists of 2 a and 2 gamma (y) globin chains (a) yp ). The 2nd globin switch is responsible for the expression of the major hemoglobin of a normal adult (HbA), consisting of 2 « and 2 B polypeptide chains (a B ) and is first expressed at mid-gestation. A key regulator of the fetal-to-adult hemoglobin switch is the transcription factor BCLIIA , which binds to the B-globin gene and acts to silence y-globinexpression and thus HbF. Embryonic Hemoglobins ‘The blood of early human embryos contains 2 slowly migrating hemoglobins, Gower-1 and Gower-2, and Hb Portland, which has HbF-like mobility. The zeta () chains of Hb Portland and Gower-1 are structurally quite similar to « chains. Both Gower hemoglobins contain the epsilon (e) B-like globin polypeptide chain. Hb Gower-I has the structure () €) , whereas Gower-2 has ay €) . Hb Portland has the structure ¢, 2 . In embryos up to 6 wk gestation, the Gower hemoglobins predominate but are no longer detectable by 3 mo of gestation. Fetal Hemoglobin By 6-8 wk gestation, HbF (a2 yp ) is the predominant hemoglobin; at 24 wk gestation it constitutes 90% of the total hemoglobin. HbF declines modestly in the third trimester, such that the HbF comprises 70-80% of the total hemoglobin. HDF production decreases rapidly postnatally (Fig. 473.7 ), and by 6-12 mo of age reaches adult levels of <2. Understanding the molecular basis of the fetal- to-adult hemoglobin switch is of interest because of the therapeutic benefits to patients with B-thalassemia and sickle cell disease, whose clinical severity is improved with modest elevation of HbF. The exact mechanisms by which BCLILIA acts to repress HbF are not fully elucidated, but erythroid-specific enhancers of BCLIIA have been identified and are potential targets for therapeutic HbF induction.© HF production 4 HOF production, after premature delivery eo a go 5 Fa 20 ° 2% a Postconceptional, 1 2 3 4 5 6 Ace ‘ ‘Term postnatal, mos FIG. 473.7 Pre- and postnatal changes in the percentage of total hemoglobin represented by fetal hemoglobin (HDF) (yellow ). The triangles represent postnatal production by reticulocytes in premature infants, and the cireles represent cord blood, {and postnatal reticulocyte production in terminfants. From Brown MS: Fetal and neonatal erythropoiesis. In Stockman JA, Pochedly C, editors: Developmental and ‘neonatal hematology , New York, 1988, Raven Press.) Adult Hemoglobins HDA constitutes 5~10% of total hemoglobin at 24 wk gestation and steadily increases, so that at term, HbA averages 30% of total hemoglobin. By 6-12 mo of age, individuals reach adult levels of HbA. The minor adult hemoglobin component, HbA , contains delta (6) chains and has the structure a 6) . At birth, <1.0% of HbAy is detected, but by 12 mo of age the normal level is 2.0— 3.4%. Throughout life, the normal ratio of HbA to HbA, is about 30 : 1. Alterations of the Hemoglobins by Disease HDF levels may be elevated with hemoglobinopathies, hereditary persistence of fetal hemoglobin, or bone marrow failure syndromes or may be associated with stress erythropoiesis. Since the HbF level is elevated during the Ist yr of life,knowledge of its normal pattern of decline is important (see Figs. 473.6 and 473.7 ). Two disorders resulting from mutations in the B-globin gene (HBB), B- thalassemia and sickle cell disease, become symptomatic postnatally as fetal y- globin expression decreases and adult B-globin increases. In both these disorders, elevated HDF levels persist in childhood and later. In patients with the most severe type, B° thalassemia, except for a small amount of HbA, , HDF is the only hemoglobin produced. At the other end of the spectrum, in individuals with B- thalassemia trait, the postnatal decrease of HbF is delayed and mildly elevated levels of HbF (>2%) may persist throughout life. Individuals with sickle cell disease, who also have a mutation in the HBB gene, typically demonstrate elevated levels of HbF, ranging from approximately 5% to up to 30%. In contrast, elevated HbF is not characteristic of a-thalassemia syndromes, but tetramers of y chains (y4 or Hb Barts) may be found in the neonatal period. Since a-globin chains are expressed in fetal and adult hemoglobin, 4 « gene mutations leading to functional deletions is not compatible with life. Fetuses die in utero or shortly after birth from the severe anemia and hydrops fetalis. Inheritance of only 1 normal gene of the 4 (« -/-) results in hemoglobin H disease , which is usually associated with a moderate anemia. Inheritance of 2 or 3 normal a genes results in a-thalassemia trait or carrier status, respectively. Hereditary persistence of fetal hemoglobin (HPFH) is a benign genetic condition caused by heterozygous deletions or nucleotide substitutions in regions of the B-globin locus that regulate transcription of HBG1 and HBG2 , causing persistent pancellular HbF expression levels of approximately 30% of total hemoglobins. Individuals with HPFH do not exhibit anemia. Preterm infants treated with human recombinant EPO increase HbF production during active erythropoiesis. Moderate elevations of HbF may also occur in many diseases accompanied by hematologic stress, such as hemolytic anemias, leukemia, and bone marrow failure syndromes such as Diamond Blackfan anemia. The normal adult level of HbA, (2.0—3.4%) is seldom altered. Levels of HbA) >3.4% are found in most persons with the B-thalassemia trait and in those with megaloblastic anemias secondary to vitamin By) and folic acid deficiency. Decreased HbA, levels are found in those with iron-deficiency anemia (see Chapter 482 ) and a-thalassemia (see Chapter 489.10 ).Red Cell Life Span in the Fetus and Neonate In general, the highest hematocrit during a person's lifetime occurs at birth, and the lowest hematocrit occurs at the physiologic nadir that occurs 8-10 wk postnatally. A shortened life span of fetal and neonatal red blood cells (RBCs) has been suggested as an important component. The average erythrocyte life span in normal adults is approximately 120 days. The life span of fetal/neonatal erythrocytes was once estimated to be considerably less, with an average of 60- 90 days suggested by chromium (°! Cr)-labeled erythrocyte studies. However, newer studies indicate that the life span of fetal/neonatal RBCs is similar to that of adults. Neocytolysis is the active removal of young erythrocytes that were generated in relatively hypoxic conditions, following normoxic or hyperoxic conditions. This process has also been suggested as an explanation for the physiologic nadir of neonates. Bibliography Christensen RD, Jopling J, Henry E, et al. The erythrocyte indices of neonates, defined using data from over 12,000 patients in a multihospital healthcare system. J Perinatol . 2008;28:24-28. Deutsch VR, Toner A. Megakaryocyte development and platelet production. Br J Haematol . 2006;134:453—-466. Jopling J, Henry E, Wiedmeier SE, et al. Reference ranges for hematocrit and blood hemoglobin concentration during the neonatal period: data from a multihospital healthcare system. Pediatrics . 2009;123:e333-€337. Julien E, Omar RE, Tavian M. Origin of the hematopoietic system in the human embryo. FEBS Lett . 2016;590:3987— 4001. Kaushansky K. Lineage-specific hematopoietic growth factors. N Engl J Med . 2006;354:2034-2045. Liang R, Ghaffari S. Advances in understanding themechanisms of erythropoiesis in homeostasis and disease. Br J Haematol . 2016;174:661-673. Liu A, Sola-Visner M. Neonatal and adult megakaryopoiesis. Curr Opin Hematol . 2011;18(5):330-337. Nandakumar SK, Ulrisch JC, Sankaran VG. Advances in understanding erythropoiesis: evolving perspectives. Br J Haematol . 2016;173:206-218. Sola-Visner MC, Christensen RD, Hutson AD, et al. Megakaryocyte size and concentration in the bone marrow of thrombocytopenic and nonthrombocytopenic neonates. Pediatr Res . 2007;61:479-484. Spangrude GJ, Perry SS, Slayton WB. Early stages of hematopoietic differentiation. Ann NY Acad Sci . 2003;996:186-194. Vats A, Bielby RC, Tolley NS, et al. Stem cells. Lancet . 2005;366:592-602. Wang Y, Hayes V, Jarocha D, et al. Comparative analysis of human ex vivo-generated platelets vs megakaryocyte- generated platelets in mice: a cautionary tale. Blood . 2015;125(23):3627—3636. Wiedmeier SE, Henry E, Sola-Visner MC, et al. Platelet reference ranges for neonates, defined using data from over 47,000 patients in a multihospital healthcare system. J Perinatol . 2009;29:130-136. Yoshimoto M, Yoder MC. Developmental biology: birth of the blood cell. Nature . 2009;457:801-813.CHAPTER 474 The Anemias Courtney D. Thornburg Anemia is defined as a reduction of the hemoglobin concentration or red blood cell (RBC) volume below the range of values occurring in healthy persons. “Normal” hemoglobin and hematocrit (packed red cell volume) vary substantially with age and sex (Table 474.1 ). There are also racial differences, with significantly lower hemoglobin levels in African American children than in white non-Hispanic children of comparable age (Table 474.2 ). Anemia is a significant global health problem affecting children and reproductive-age women (Figs. 474.1 and 474.2). Table 474.1 Normal Mean and Lower Limits of Normal for Hemoglobin, Hematocrit, and Mean Corpuscular Volume AGE gx) HEMOGLOBIN Gil) HEMATOCRIT (9, MEAN CORPUSCULAR VOLUME (iN) Mean LowerLimit Mean Lower Limit Mean Lower Limit vss ps [ao a7 [33 7 70 za 150 3a [st 7 73 57 13.0 [1s 39 [35 a 7 eit 135120 a0 [36 a5, 75 12-14 Female| 135120 a_[36 35, 78 Teidmale [140125 a a7 a 77 15-17 Female| 140 [ 120. a [36 a7, 7 TSA7male [150 [130 a6 _[38 36, 78 TeA9 Female] 140 [ 120, a [37 90) 30 TeA9male [160 [140 aa 30) 30 From Brugnara C, Oski FJ, Nathan DG: Nathan and Oski's hematology of infancy and childhood , ed 7, Philadelphia, 2009, Saunders, p 456.Table 474.2 NHANES-III Hemoglobin Values for Non-Hispanic Whites and African Americans Ages 2-18 Yr* AGE OD WHITE NON-HISPANIC ‘AFRICAN AMERICAN Mean 28D Mean 23D 2 1221 0s T1395 1037 610 1287 TT 12.40, 10.74 TIS male 1376 TL 76 13.05 10.88 THIS female 1332 TS, TZ 10.85 16-18 male 75.00 Ts2t THe Taz 16-18 Female 1330 ToT 1237 10.37 * Sample size is 5,142 (white, 2,264; African-American, 2,878) NHANES-IIl, Third National Health and Nutrition Examination Survey; SD, standard deviation, Adapted from Robbins EB, Blum S: Hematologic reference values for African American children and adolescents, Am J Hematol 82:611-614, 2007. FIG. 474.1 Global prevalence of anemia in children of preschool age (0-5 yr). (Adapted ‘from Worldwide prevalence of anaemia 1993-2005. In WHO global database on ‘anaemia, Geneva, 2008, World Health Organization.)Genetic hemoglobin disorders: Thalessemias Hemoglebin veriants Glucose-6-ohosphate dehydrogenase deficiency Ovalocytosis LE cre tne ‘Soiltransmited helminths ‘Nutrition Malar Iron doticioncy ‘Schistosomiasis, Flic acid deficiency “Tuberculosis Vitamin B12 deficiency ‘AIDS: Vitamin A deficiency Leishmaniasis Protein energy malnutrition ‘Topical sprue Malabsorption ard disorders f the sail intestine FIG. 474.2 Causes of anemia in countries with low or middle-income populations. (From Balarajan Y, Ramakrishnan U, Ozaltin E, et al: Anaemia in lowincome and ‘middle-income countries, Lancet 378:2123-2134, 2011, Fig 3.) Physiologic adjustments to anemia include increased cardiac output, augmented oxygen extraction (increased arteriovenous oxygen difference), and a shunting of blood flow toward vital organs and tissues. In addition, the concentration of 2,3-diphosphoglycerate increases within the RBC. The resultant “shift to the right” of the oxygen dissociation curve reduces the affinity of hemoglobin for oxygen and results in more complete transfer of oxygen to the tissues. The same shift in the oxygen dissociation curve can also occur at high altitude. Higher levels of erythropoietin (EPO) and consequent increased RBC production by the bone marrow further assist the body to adapt. History and Physical Examination As with any medical condition, a detailed history and thorough physical exam are essential when evaluating an anemic child. Important historical facts should include age, sex, race and ethnicity, diet, medications, chronic diseases, infections, travel, and exposures. A family history of anemia and associated difficulties (e.g., splenomegaly, jaundice, early-age onset of gallstones) is also important. Often, few physical symptoms or signs result solely from a lowhemoglobin, particularly when the anemia develops slowly. Clinical findings generally do not become apparent until the hemoglobin level falls to <7-8 g/dL. Clinical features can include pallor, sleepiness, irritability, and decreased exercise tolerance. Pallor can involve the tongue, nail beds, conjunctiva, palms, or palmar creases. A flow murmur is often present. Ultimately, weakness, tachypnea, shortness of breath on exertion, tachycardia, cardiac dilation, and high-output heart failure will result from increasingly severe anemia, regardless of its cause. Unusual physical findings linked to particular underlying disease etiologies are discussed in detail in sections describing the associated disorders and in Table 474.3 . Table 474.3 Physical Findings in the Evaluation of Anemia SYSTEM OBSERVATION ‘SIGNIFICANCE Skin Hyperpigmentation Fanconi anemia, congenital dyskeratoas Café au lait spats Fanconi anemia Vitiligo Vitamin Bp deficiency Partial oculocutaneous | Chédiak- Higashi syndrome albinism Taundice emalysis, hepatitis Petechiae, purpura ‘Bone marrow inflation, autoimmune hemoly’is with autoimmune thrombocytopenia, hemalytic-uremic syndrome Enythematous rash Parvovirus, Epstein-Barr virus Butterfly rash, ‘Systemic Tupus erythematosus Head Frontal bosing ‘Thalassemia major, severe iron deficiency, chronic subdural hematoma Microcephaly Fanconi anemia Diamond-Blackfan syndrome Eyes Microphihalmia Fanconi anemia Retinopathy Hemoglobin SS, SC disease ‘Optic atrophy, Blindness | OSeopetosis Blocked lacrimal gland | Dyskeratosis congenital Kayser-Fleischer ring | Wilson disease Blue sclera Tron deficiency Eas Deafness ‘Osteopetosis Mouth | Glossitis Vitamin Bip deficiency, iron deficiency ‘Angular Somattis Tron deficiency, Cleft lip, palate Diamond-BlackTan syndrome Pigmentation Peutz-Teghers syndrome (intestinal blood Toss) Telangiectasia ‘Osler-Weber-Rendu syndrome (blood loss) Teukoplakia ‘Dyskeratosis congenital Chest | Shield chest or Diamond-Blackfan syndrome ‘widespread nipples Murmur Endocarditis prosthetic valve hemolyas ‘Abdomen | Hepatomegaly Femolysis, infiltrative tumor, chronic disease, hemangioma, cholecysaus| ‘Splenomegaly Femolysis, sickle cell disease (early), thalassemia, malaria, lymphoma,Epstein-Barr virus, portal hypertension, hemophagocytic syndromes, Nephromegaly Fanconi anemia ‘Absent kidney Fanconi anemia Exiremities| Absent thumbs Fanconi anemia "Thenar eminence Diamond-Blackfan syndrome bypoplasia: triphalangeal thumb ‘Spoon nails Tron deficiency Beau Tine (nails) Heavy metal intoxication, severe Ines Mees Tine (nails) Fleavy metals, severe illness, sickle cell anemia Dystrophic nails Dyskeratosis congenital Edema ‘Milk-induced protein-Tosing enteropathy with ivan deficiency, renal failure Recal | Hemorrhoids Portal hypertension Heme positive stool | Tntestinal hemorhage Nerves | liritable, apathy Tron deficiency’ Peripheral neuropathy | Deficiency of vitamins B; and Byp , lead poisoning Dementia Deficiency of vitamins Biz and E ‘Alaxia, posterior column | Deficiency of vitamins By2 and E signs ‘Soke ‘Sickle cell anemia, paroxysmal nocturnal hemoglobinuria ‘Adapted from Scott JP: Hematology. In Behrman RE, Kliegman RM, editors. Nelson essentials of pediatrics, ed 2, Philadelphia, 1994, Saunders, p 520. Laboratory Studies Initial laboratory testing should include hemoglobin, hematocrit, and RBC indices as well as a white blood cell (WBC) count and differential, platelet count, reticulocyte count, and examination of the peripheral blood smear. The need for additional laboratory studies is dictated by the history, physical exam, and results of this initial testing. Differential Diagnosis Anemia is not a specific entity but rather can result from any of a number of underlying pathologic processes. To narrow the diagnostic possibilities, anemias may be classified on the basis of their morphology and physiology (Fig. 474.3 ).merooytio Normosytic acrooytic etieuteeye court] Reteulbeye court [Feteaoojie count ¥ ¥ ¥ ‘Lowtnaioquate igh Lewinadequate igh Lowinedequate shendelcory + Thaaseniagonee = * hasdymaiied = Fleder + Prepaninotose "eer oun © Macey ‘fen Stoop). + Neosho Peer sats) [eoresan Stine FIG. 474.3 Use of the mean corpuscular volume (MCV) and reticulocyte count in the diagnosis of anemia. (Adapted from Brunetti M, Cohen J: The Harriet Lane handbook , ed 17, Philadelphia, 2005, Elsevier Mosby, p 338.) Anemias may be morphologically categorized on the basis of red cell size (mean corpuscular volume [MCV]) and microscopic appearance. Anemias can be classified as microcytic, normocytic, or macrocytic based on whether the MCV is low, normal, or high, respectively. RBC size also changes with age, and normal developmental changes in MCV should be recognized before a designation is made (see Table 474.1 ). Examination of a peripheral blood smear often reveals changes in RBC appearance that will help to narrow further the diagnostic categories (Fig. 474.4 and Table 474.4 ). Details regarding morphologic changes associated with particular disorders are described in subsequent sections. D x E FIG. 474.4 Morphologie abnormalities of the red blood cell. A, Normal. B, Macrocytes (olic acid or vitamin B,, deficiency). C, Hypochromic microcytes (Iron deficiency). D,Target cells (HDC disease). E, Schizocytes (hemolytic-uremic syndrome). (Courtesy of Dr-E, Schwartz) Table 474.4 Peripheral Blood Morphologic Findings in Various Anemias Microcytes Iron deficiency Thalassemias Lead toxicity Anemia of chronic disease Macrocytes Newborns Vitamin By) or folate deficiency Diamond-Blackfan anemia Fanconi anemia Aplastic anemia Liver disease Down syndrome Hypothyroidism Spherocytes Hereditary spherocytosis Immune hemolytic anemia (newborn or acquired) Hypersplenism Sickled Cells Sickle cell anemias SS diseaseSC disease SB* thalassemia SB° thalassemia Elliptocytes Hereditary elliptocytosis Iron deficiency Megaloblastic anemia Target Cells Hemoglobinopathies (especially hemoglobin C, SC, and thalassemia) Liver disease Xerocytosis Basophil Stippling Thalassemia Lead intoxication Myelodysplasia RBC Fragments, Helmet Cells, Burr Cells Disseminated intravascular coagulation Hemolytic-uremic syndrome Thrombotic thrombocytopenic purpura Kasabach-Merritt syndrome Waring blender syndrome Uremia Liver disease Hypersegmented NeutrophilsVitamin By) or folate deficiency Blasts Leukemia (ALL or AML) Severe infection (rarely) Leukopenia, thrombocytopenia Fanconi anemia Aplastic anemia Leukemia Hemophagocytic histiocytosis Howell-Jolly Bodies Asplenia, hyposplenia Severe iron deficiency ALL, Acute lymphocytic leukemia; AML, acute myeloid leukemia. From Kliegman RM, Lye PS, Bordini BIJ, et al, editors: Nelson pediatric symptom-based diagnosis, Philadelphia, 2018, Elsevier (Table 37-7, p 666). Anemias may also be further divided on the basis of underlying physiology. ‘The 2 major categories are decreased production and increased destruction (or loss ). The 2 groups are not always mutually exclusive. Decreased RBC production may be a consequence of ineffective erythropoiesis or a complete or relative failure of erythropoiesis. Increased destruction or loss may be secondary to hemolysis, sequestration, or bleeding. The peripheral blood reticulocyte percentage or absolute number helps to distinguish between the 2 physiologic categories. The normal reticulocyte percentage of total RBCs during most of childhood is approximately 1%, with an absolute reticulocyte count of 25,000- 75,000/mm} . In the presence of anemia, EPO production and the absolute number of reticulocytes should rise. Low or normal numbers of reticulocytes generally represent an inadequate response to anemia that is associated with relative bone marrow failure or ineffective erythropoiesis. Increased numbers of reticulocytes represent a normal bone marrow response to ongoing RBCdestruction (hemolysis), sequestration, or loss (bleeding). Fig. 474.3 presents a useful approach to assessing the common causes of anemia in the pediatric age-group. Children with microcytic anemia and low or normal reticulocyte counts most often have defects in erythroid maturation or ineffective erythropoiesis. Iron deficiency is the most common cause (see Chapter 482 ). Thalassemia trait constitutes the primary differential diagnosis when iron deficiency is suspected (see Chapter 489 ). Distinctions between these entities are presented in Table 482.2 (see Chapter 482 ). Chronic disease or inflammation (more often normocytic), lead poisoning, and sideroblastic anemias should also be considered and are discussed in other chapters. Microcytosis and elevated reticulocyte counts are associated with thalassemia syndromes and hemoglobins C and E (see Chapter 489 ). Notably, thalassemias and hemoglobinopathies are most often seen in patients of Mediterranean, Middle Eastern, African, or Asian descent. Normocytic anemia and low reticulocyte count characterize a large number of anemias. The anemia of chronic disease/inflammation is usually normocytic (see Chapter 482 ). The anemia associated with renal failure, primarily a result of reduced EPO production, will invariably be associated with clinical and laboratory evidence of significant kidney disease. Decreased or absent RBC production secondary to transient erythroblastopenia of childhood, infection, drugs, or endocrinopathy usually results in a normocytic anemia, as does bone marrow infiltration by malignancy. In the case of invading leukemia or malignancy, abnormal leukocytes or tumor cells in association with thrombocytopenia or reduced or elevated WBC counts may be seen. Acute bleeding, hypersplenism, and congenital dyserythropoietic anemia type II are also normocytic (see Chapter 479 ). In children with normocytic anemia and an appropriate (high) reticulocyte response, the anemia is usually caused by bleeding, hypersplenism, or ongoing hemolysis. In hemolytic conditions, reticulocytosis, indirect hyperbilirubinemia, and increased serum lactate dehydrogenase are indicators of accelerated erythrocyte destruction. Many causes of hemolysis result from conditions that are extrinsic (usually acquired) or intrinsic (usually congenital) to the erythrocyte. Abnormal RBC morphology (e.g., spherocytes, sickle forms, microangiopathy) identified on the peripheral smear is often helpful in ascertaining the cause. ‘The anemia seen in children with macrocytic blood cells is sometimes megaloblastic, resulting from impaired DNA synthesis and nuclear development(see Chapter 481 ). The peripheral blood smear in megaloblastic anemias contains large macroovalocytes, and the neutrophils often show nuclear hypersegmentation. The major causes of megaloblastic anemia include folate deficiency, vitamin B,) deficiency, and rare inborn errors of metabolism. Other macrocytic anemias with low or normal reticulocyte counts include acquired and congenital (Diamond-Blackfan and Fanconi syndromes) aplastic anemias and hypothyroidism. Patients with trisomy 21 have macrocytic cells, although an accompanying anemia is generally not present. High MCV and reticulocytosis is seen in congenital dyserythropoietic anemias I and III and in situations where hemolysis results in such a large outpouring of young red cells that the mean MCV is abnormally high. Bibliography Noronha SA. Acquired and congenital hemolytic anemia. Pediatr Rev . 2016;37(6):235-246. Powell DJ, Achebe MO. Anemia for the primary care physician. Prim Care . 2016;43(4):527-542. Wang M. Iron deficiency and other types of anemia in infants and children. Am Fam Physician . 2016;93(4):270-278.SECTION 2 Anemias of Inadequate Production OUTLINE Chapter 475 Congenital Hypoplastic Anemia (Diamond-Blackfan Anemia) Chapter 476 Pearson Syndrome Chapter 477 Acquired Pure Red Blood Cell Anemia Chapter 478 Anemia of Chronic Disease and Renal Disease Chapter 479 Congenital Dyserythropoietic Anemias Chapter 480 Physiologic Anemia of Infancy Chapter 481 Megaloblastic Anemias Chapter 482 Iron-Deficiency Anemia Chapter 483 Other Microcytic AnemiasCHAPTER 475 Congenital Hypoplastic Anemia (Diamond-Blackfan Anemia) Courtney D. Thornburg Diamond-Blackfan anemia (DBA) is a rare, congenital bone marrow failure syndrome that usually becomes symptomatic in early infancy. More than 90% of cases are recognized in the Ist yr of life. The disorder is characterized by anemia, usually normochromic and macrocytic; reticulocytopenia; and insufficient or absent red blood cell (RBC) precursors in an otherwise normally cellular bone marrow. Up to 50% of affected individuals have additional, extrahematopoietic anomalies. Etiology DBA-associated mutations were initially identified in 1997 in RPS19, a gene that encodes a component protein of the small 40S ribosomal subunit. Such RPS19 mutations, all dominantly inherited, were found to be present in approximately 25% of patients. Additional ribosomal protein (RP) genes, each encoding a different small (40S) or large (60S) ribosomal subunit protein, have been identified. Mutations in RP genes were ultimately identified in up to 70% of cases, most with autosomal dominant inheritance. Novel mutations continue to be identified and reported. Since the majority of causative mutations are in RP genes, the disorder is often referred to as a ribosomopathy . GATAL , a non-RP gene, has also been implicated in DBA. The GATAL mutations are inherited as X-linked recessive and usually have no extrahematopoietic manifestations. It remains unclear whether 2 pathways, 1 related to ribosomal dysfunction and 1 to impaired GATAI production, independently cause the same phenotype, or alternatively, that DBA results fromproblems in a single pathway involving functional links between ribosomes and GATAI (Fig. 475.1). Rivesomal gene GATAY gene mutations Steroid ‘mutations: responsiveness, ; Increased oADA Seer neutropenia, exreheratopoietie hypoplasia, coyseryrropoxsis, anomalies: skeletal, ineraasea cv, } thrombocytopenia ronal, heart, increased fH, orofacial, bone, transtuson-dependent ersioctine iron everbad Malignant traneformetion? ‘Acivaton of psa? BM aplasia? FIG. 475.1 Common and distinet phenotypes in congenital red cell aplasia caused by ‘mutations in RP genes and in GATAI . BM, Bone marrow, eADA, erythrocyte adenosine deaminase activity; Hb, fetal hemoglobin; MCV, mean corpuscular volume. (Adapted from Weiss MJ, Mason PJ, Bessler Mt What's in a name? J Clin Invest 122:2346-2349, 2012.) Epidemiology DBA affects about 7 individuals per 1 million live births. It is primarily an autosomal dominant disease, although other inheritance patterns may yet be demonstrated. Notably, there is substantial phenotypic diversity in DBA, even in families whose members share the same mutation, suggesting that additional genetic modifiers affect phenotypic expression of the disease. International consensus recommendations suggest that a diagnosis of “nonclassical” DBA be applied to family members harboring an established mutation or those without a known mutation but with an associated anomaly or laboratory abnormality. Clinical Manifestations Profound anemia usually becomes evident by 2-6 mo of age, occasionally somewhat later. Approximately 25% of patients are anemic at birth, and hydropsfetalis occurs rarely; 92% are diagnosed within the Ist yr of life. Approximately 40-50% of patients have congenital anomalies, and >1 anomaly is found in 25% of DBA patients (Table 475.1 ). Craniofacial abnormalities are the most common (50% of patients) and include snub nose and high-arched palate. Skeletal anomalies, mostly upper limb and hand, affect 30%. Thumb abnormalities, including flattening of the thenar eminence and triphalangeal thumb, may be bilateral or unilateral. The radial pulse may be absent. Genitourinary (38%), cardiac (30%), ophthalmologic, and musculoskeletal anomalies have also been identified. Short stature is common, but it is often unclear whether this characteristic results from the disease itself, related therapies, or both. Table 475.1 Range of Congenital Anomalies Observed in Diamond- Blackfan Anemia TYPE/LOCATION ANOMALIES ‘Craniofacial Hypertelorism Broad, flat nasal bridge Cleft palate High-arched palate Microcephaly Micrognathia Microtia Low-set ears Low hairline Prosis ‘Ophihalmologic ‘Congenital glaucoma Suabismus Epicanthal folds Congenital cataract Neck Short neck, ‘Webbed neck Sprengel deformity Klippel-Feil deformity Thumbs “Triphalangeal Duplex or bifid Hypoplastic Flat thenar eminence Absent radial artery Urogenital ‘Absent kidney Horseshoe kidney Hypospadias Cardiac ‘Ventricular sepial defect Auial septal defect ‘Coarctation of the aorta ‘Complex cardiac anomalies ‘Oiher Tow birthweightShort stature Syndactyly Learning difficulties Multiple anomalies, most often including craniofacial, are present in up to 25% of affected individuals. At least 1 anomaly is present in 40-50%. From Viachos A, Ball S, Dahl N, et al: Diagnosing and treating Diamond-Blackfan anaemia’ results of an international clinical consensus conference, Br J Haematol 142(6): 859-876, 2008 (Table IV). Laboratory Findings ‘The RBCs are usually macrocytic for age, but no hypersegmented neutrophils or other characteristics of megaloblastic anemia are appreciated on the peripheral blood smear. RBC enzyme patterns are similar to those of a “fetal” RBC population, with increased expression of “i” antigen and elevated fetal hemoglobin (HbF). Erythrocyte adenosine deaminase (eADA) activity is increased in most patients with DBA, a finding that helps distinguish congenital RBC aplasia from acquired transient erythroblastopenia of childhood (TEC) (see Chapter 477 ). Because elevated eADA activity is not a fetal RBC feature, measurement of this enzyme may be particularly helpful when diagnosing DBA in very young infants. Thrombocytosis, or rarely thrombocytopenia, and occasionally neutropenia, may also be present. Reticulocyte percentages are characteristically very low despite severe anemia. Bone marrow erythrocyte precursors are greatly reduced in most patients; other marrow elements are usually normal. Serum iron levels are elevated. Unlike Fanconi anemia, there is no increase in chromosomal breaks when lymphocytes are exposed to alkylating agents. Table 475.2 outlines suggested diagnostic criteria. Table 475.2 Diagnostic Criteria for Patients With Diamond-Blackfan Anemia SUPPORTING CRITERIA DIAGNOSTIC CRITERIA Major Criteria Minor Criteria ‘Age younger than Tyr Pathogenic Elevated red cell adenosine deaminase ‘mutations, Macrocyiie anemia Positive family — | Congenital anomalies of Diamond-Blackfan anemia history Raticulocytopenia Elevated fetal hemoglobinPay bene mano ened No evidence for other inherited bone marrow failure precursors syndromes From Vlachos A, Ball S, Dah! N, et al: Diagnosing and treating Diamond-Blackfan anaemia: results of an international clinical consensus conference, Br J Haematol 142(6):859-876, 2008. Differential Diagnosis DBA must be differentiated from other anemias associated with low reticulocyte counts. The syndrome of TEC is often the primary alternative diagnosis. Table 477.1 shows a useful comparison of findings in these 2 disorders (see Chapter 477 ). TEC often is differentiated from DBA by its relatively late onset, although it occasionally develops in infants <6 mo of age. Macrocytosis, congenital anomalies, fetal RBC characteristics, and elevated eADA are generally associated with DBA and not with TEC. Other inherited macrocytic bone marrow failure syndromes, particularly Fanconi anemia and Shwachman-Diamond syndrome (see Chapter 495 ), should also be considered, as should myelodysplastic syndrome. Aase syndrome includes congenital RBC aplasia with triphalangeal thumb, congenital heart disease, and cleft palate. Hemolytic disease of the newborn can also mimic features of DBA because it can have a protracted course and can be coupled with greatly reduced erythropoiesis. The anemia in this disorder usually resolves spontaneously at 5-8 wk. of age. Several types of chronic hemolytic disease may be complicated by an aplastic crisis , characterized by reticulocytopenia and decreased numbers of RBC precursors. This event usually occurs after the Ist several mo of life and is often caused by parvovirus B19 infection (see Chapter 477 ). Infection with parvovirus B19 in utero also may be associated with pure RBC aplasia in infancy, and even with hydrops fetalis at birth (see Chapter 278 ). When diagnosing DBA in young infants, it is important to rule out parvovirus B19 infection using the polymerase chain reaction. Other infections, including HIV, as well as drugs, immune processes, and Pearson syndrome (see Chapter 476 ), should also be ruled out. Treatment Corticosteroids are a mainstay of therapy, and approximately 80% of patients initially respond. Because corticosteroids impair linear growth as well as physical and neurocognitive development, many hematologists maintain infants