H



Problems in the Fetus and
Neonate
YASER DIAB • LORI LUCHTMAN-JONES
Hematopoietic Development
 HEMATOPOIETIC STEM CELLS
Blood cells arise from the differentiating embryonic
mesoderm. Human erythroid and macrophage progeni-
tor cells have been observed in the yolk sac by days 16
to 19 and at day 19 in the aortic-gonad-mesonephros
(AGM). After the development of the circulatory system
on day 21, pluripotent hematopoietic cells localize in the
AGM, placenta and liver by days 24 to 28. Myeloid, lym-
phoid and megakaryocytic precursors have been noted in
fetal liver at this stage. Definitive erythropoiesis occurs in
the fetal liver, thymus, spleen and bone marrow. A knowl-
edge gap exists about the details of in situ hematopoiesis
between weeks 3 and 12, but fetal liver is believed to be
the major site of hematopoiesis between weeks 6 and
16. The bone marrow assumes this role by week 24
(F 88-1)3
P blood cell formation is necessary for survival.
Much has been learned about the origin and regulation
of blood cells through studies of congenital and acquired
defects in hematopoiesis. Studies of mouse hematogen-
esis identified pluripotent cells in the murine inner cell
mass. These embryonic stem (ES) cells are capable of self-
renewal as well as differentiation into hematopoietic
cells. Human ES cells were isolated in 1998, fueling
research efforts to generate pluripotent stem cells from
early embryos and to perform genetic manipulation of
differentiated somatic cells. Somatic cells can be repro-
grammed into an ES-like state, and these induced plu-
ripotent stem (iPS) cells are another tool researchers are
using both to understand and manipulate progenitor
cells.3
T earliest blood cells produced by colony-forming
cells (CFCs) in the yolk sac are very large, primitive ery-
throid cells expressing embryonic globins. Subsequently,
CFCs produce definitive erythrocytes expressing fetal
globins and macrophages. Later still, multipotent CFCs
and lymphoid progenitor cells arise. The adult-type
globins are not expressed until just before birth and
rapidly assume primacy afterward (T  !!"#; F
!!"8) With each transition from primitive to definitive
to adult erythroid cells, the mean corpuscular volume
(MCV) decreases.
1

REGULATION OF HEMATOPOIESIS
Few hematopoietic stem cells enter the cell cycle at any
given time. Most are in a resting state. Proliferation and
differentiation occur within a suitable microenvironment
of stroma and humoral factors. The WNT/β-catenin and
Notch-δ signaling pathways drive stem cell development.
Transcription factors involved in stem cell development
include GATA2, RUNX1, TEL/ETV6, SCL/TAL1, and LM02.
Other transcription factors such as PU.1, GFIX, C/EBPα,
and GATA1 are considered to be more lineage-specific,
but most also participate in lineage priming, where stem
cells differentiate along a pathway depending upon cel-
lular and environmental stimuli. Multiprotein complexes
assemble and bind DNA regulatory elements to modulate
transcription. Epigenetic regulatory mechanisms of tran-
scription include DNA methylation on CpG residues and
histone modification.3
F$%& '%( '%&& ($ * +,P-.
VEGF, WNT, and FGF. Hematopoietic cytokines such
as stem cell factor, fms-like tyrosine kinase receptor-3
ligand, interleukin-6, thrombopoietin, erythropoietin,
and granulocyte colony-stimulating factor (G-CSF) play
critical roles in the maintenance and differentiation of
human hematopoietic cells. Some hematopoietic growth
factors are produced in the vicinity of hematopoietic
progenitors, and others are synthesized remotely (T 
!!"8) Few of the glycoprotein growth factors are avail-
able for clinical use, but that number is expected to
increase.
Some of the earliest hematopoietic growth factors dis-
covered were referred to as colony-stimulating factors (CSFs),
because in culture they stimulate progenitor cells to form
colonies of recognizable maturing blood cells. The pre-
fixes refer to the maturing cell produced. GM-CSF is
granulocyte-macrophage CSF. The interleukins (ILs) were
named for the fact that they are derived from, or act upon,
leukocytes. Other factors are named for the cell surface
receptor to which they bind, such as thrombopoietin
receptor agonists that stimulate megakaryocyte differen-
tiation and platelet production. Growth factors such as
IL-3 and GM-CSF stimulate proliferation, differentiation,
and survival of a broad range of precursors, including
stem cells. Others such as erythropoietin and granulocyte
CSF (G-CSF) are lineage restricted.
 // • HEMATOLOGIC AND ONCOLOGIC PROBLEMS IN THE FETUS AND NEONATE 1295

S024 52667 C:44<002= >?:@2A<0:?7 Mature cells

T-lymphocyte
Pre-T cell
B-lymphocyte/
L9mphoid plasma cell
stem cell
Pre-B cell

Erythrocyte
BFU-E CFU-E
Megakaryocyte/
platelets
Meg-CFC
Basophil/
Hematopoietic Multi- mast cell
stem cell CFC Mast-CFC
Eosinophil
Eo-CFC
Neutrophil
Self-renewal
G-CFC
Monocyte/
GM-CFC macrophage
M-CFC
Osteoclast
Oc-CFC
Figure 88-1 Hematopoietic stem cell differentiation. BFU-E, Burst-forming units, erythroid; CFC, colony-forming cells; CFU-E, colony-forming
unit, erythroid; Eo-CFC, eosinophil colony-forming cells; G-CFC, granulocyte colony-forming cells; GM-CFC, granulocyte-macrophage colony-forming
cells; M-CFC, macrophage colony-forming cells; Meg-CFC, megakaryocyte colony-forming cells; Oc-CFC, osteoclast colony-forming cells.

100
F—α2γ2

80
Hemoglobins: % of total

A
60

40
F
ζ2ε2
20
A—α2β2
ζ 2 γ2
α2ε2 A2—α2δ2
0
α
Globin subunits: % of total

50

40
Gγ β
30

20 ζ Aγ
10 ε
β
δ
0
J K0 20 30 10 20 30 40 Weeks
Birth
Crown-rump
3 5 10 15 20 =cm

Fetus Newborn
BDEGIe 88-2 Changes in expression of hemoglobin tetramers (upper panel) and individual globin chains (lower panel) during development. (Adapted
from Bunn HF, et al. Hemoglobin: molecular, genetic, and clinical aspects. Philadelphia: Saunders; 1969.)
 1296 MNOQ 14 • THE BLOOD AND HEMATOPOIETIC SYSTEM

TABLE 88-1 Human Hemoglobins Expressed Red Blood Cells

During Development
]^_`ab`cde AND
Globin OXYGEN-CARRYING CAPACITY
Z[\Xglobin Predominates Composition
The function of red blood cells (RBCs) is to transport
Hb Gower 1 Embryonic (yolk sac) ζ2ε2 oxygen (O2) to tissues to meet metabolic demands.
Hb Gower 2 Embryonic (yolk sac) α2ε2
Hemoglobin (Hb), the most abundant protein in eryth-
Hb Portland Embryonic (yolk sac) ζ2γ2
rocytes, facilitates oxygen delivery by reversibly binding
Hb F Fetal (liver) α2γ2
Hb A Adult (bone marrow) α2β2 O2 molecules. The binding of oxygen to hemoglobin tet-
Hb A2 Minor adult (bone marrow) α2δ2 ramers is cooperative, resulting in the familiar sigmoidal
Hb Barts Fetal-alpha thalassemia γ4 oxygen dissociation curve (fghijk 88-3lm The affinity of
Hb H Adult-alpha thalassemia β4 Hb molecules for oxygen is influenced by a variety of
factors, including temperature, pH, carbon dioxide pres-
sure (PCO2), and the concentration of red blood cell
organic phosphates (2,3-biphosphoglycerate or 2,3-BPG,
also known as 2,3-diphosphoglycerate or 2,3-DPG). In
the case of adult hemoglobin (Hb A), oxygen affinity for
the molecule varies directly with pH and inversely with
temperature and the concentration of 2,3-BPG. Fetal

TABLE 88-2 Hematopoietic Growth Factors

RUVWXY Source Receptor Target Cells Effects
Erythropoietin (EPO) Kidney, hepatocytes EPO-R E, Meg Stimulates growth and differentiation of
erythroid precursors
Stem cell factor (SCF) (also Ubiquitous KIT E, mast cells, Stimulates growth and differentiation of
known as steel factor [SF], melanocytes, erythroid and myeloid precursors;
KIT ligand [KL], and mast germ cells enhances growth of mast cells
cell growth factor [MCGF])
Granulocyte colony- Stromal cells, G-CSF-R N Stimulates growth and differentiation of
stimulating factor (G-CSF) macrophages neutrophil precursors; activates
phagocytic function of mature neutrophils
Granulocyte-macrophage Stromal cells GM-CSF-R (α M, N, Eo, Endo Stimulates growth and differentiation of
colony-stimulating factor and β chains) neutrophils, eosinophils, and monocytes;
(GM-CSF) activates endothelial cells; induces
cytokine expression by monocytes
Macrophage colony- Mesenchymal cells FMS M Stimulates growth and differentiation of
stimulating factor (M-CSF) monocytes; induces phagocytic function
in monocytes and macrophages; is
involved in bone remodeling
Interleukin 1 (IL-1) Ubiquitous IL-1RI, IL-1RII T, E, B, M, S Induces production of cytokines and
prostaglandins by stromal cells, T cells,
and many other cell types; induces fever
Interleukin 2 (IL-2) T cells P55, P75 B, T, NK Induces proliferation and activation of T, B,
and NK cells; induces IL-1 expression by
monocytes
Interleukin 3 (IL-3) T cells IL-3Rα, M, N, Eo, Meg Stimulates growth and differentiation of
GM-CSF-Rβ myeloid and erythroid precursors,
induces cytokines
Interleukin 4 (IL-4) T cells, mast cells, IL-4R M, Ba, B, T Induces proliferation and activation of B
basophils and T cells
Interleukin 6 (IL-6) Ubiquitous IL-6R/GP130 B, N Induces activation of neutrophils; induces B
cell maturation, synergistic with IL-3
Interleukin 7 (IL-7) Stromal cells IL-2R B, T, meg Stimulates T cells; induces monocytes
Interleukin 8 (IL-8) Stromal cells, IL-8R T, N Induces neutrophils and chemotaxis
macrophages, T cells
Interleukin 10 (IL-10) T cells, macrophages IL-10R Meg, E Induces B and mast cells; inhibits T cells
Interleukin 11 (IL-11) Stromal cells IL-11R, GP130 Meg Stimulates megakaryocytes
Interleukin 12 (IL-12) Neutrophils, monocytes IL-12R T, NK Induces differentiation of cytotoxic T cells
Thrombopoietin (TPO) Unknown MPL Meg Stimulates megakaryocytes

B, B cells; Ba, basophil; E, erythroid precursors; Endo, endothelial cell; Eo, eosinophil; M, monocyte; Meg, megakaryocyte; N, neutrophil; NK, natural killer cell;
S, stroma cell; T, T cell.
Adapted from Bagby CC. Hematopoiesis. In: Stamatoyannopoulos G, et al, eds. The molecular basis of blood diseases. Vol 2. Philadelphia: Saunders; 1994:76.
 // • HEMATOLOGIC AND ONCOLOGIC PROBLEMS IN THE FETUS AND NEONATE
Š‹‹
90
80
↑ pH
—˜– 70 ↓ DPG ¯ pH
•“” ↓ Temp ↑ DPG
60 ↑ Temp
“ 7-50
’‘ 50

Ž 40
 151-200
Π30
20
Š‹
0 10 20 30 40 50 60 70 80 90 100
P50
™2 Tension (mm Hg)
Figure 88-3 Oxyhemoglobin dissociation curve. Factors that influ-
ence the position of the curve are indicated. DPG, Diphosphoglycerate.
(Adapted from Bunn HF, et al. Hemoglobin: molecular, genetic, and clinical
aspects. Philadelphia: Saunders; 1969.)
hemoglobin (Hb F) has a high oxygen affinity. Some
mutations of hemoglobin affect oxygen affinity, as
explained later.
HEMOGLOBIN SWITCHING
The hemoglobin tetramer is composed of two heterodi-
heterodi
mers, consisting of an α-- and β-type-type globin. Different
globin genes are sequentially expressed in RBC precur-
sors, a process known as hemoglobin switching (see
fghijk 88-2 o€ nopqk rrsƒlm During development, α- and
β-type globin gene clusters are activated sequentially
from the 5′ (embryonic) end to the 3′ (adult) end. The
α-type globin genes, ζ-globin, α1-globin, and α2-globin,
are located on chromosome 16 with the ζ gene 5′ to a
pair of duplicated α-globin genes. The β-type genes on
chromosome 11 are oriented 5′ to 3′ as ε-, Gγ, Aγ, δ, and
β (see fghijk 88-2lm The protein products of the Aγ- and
G
γ-globin genes are functionally similar and differ by a
single amino acid residue. Globin gene expression is
controlled by cis-elements of individual globin gene pro-
moters, proximal and distal enhancer regions, and posi-
tively acting transcription factors, such as GATA1, GATA2,
NFE2, MYB, EKLF, RBTN2, and SCL. Other mechanisms
that also regulate globin switching are silencers, DNA
conformational changes, and DNA methylation.„…†‡ˆ ‰
fact, discovery of the ability to chemically demethylate
CpG residues in the silenced γ-globin gene promoter
launched translational research efforts to enhance fetal
hemoglobin production in patients with β-globin defects
such as β thalassemia and sickle cell anemia.
During yolk sac hematopoiesis, RBCs produce the
embryonic hemoglobins (see nopqk rrsƒlm ). Hb F is the
predominant hemoglobin in the fetus and neonate. Pro Pro-
duction of the major adult hemoglobin (Hb A) increases
significantly between birth and 6 months of age, as Hb F
production declines. Synthesis of HbA2, a minor adult
globin, also increases gradually over the first months of
fghijk 88-2
life (see Figure lm
88-2).
). After 6 months of age, Hb F usually
constitutes less than 1% of the total hemoglobin and is
unevenly distributed among red blood cells. Each of the
1297
TABLE 88-3 Serum Erythropoietin Levels
During Infancy
šX›WœUWU žŸ[ Serum EPO Level
(Days) (mU/Ml) Sample Size
0-6 33.0 ± 31.4 11
11.7 ± 3.6 7
51-100 21.1 ± 5.5 13
101-150 15.1 ± 3.9 5
17.8 ± 6.3 6
>200 23.1 ± 9.7 10
Data from Yamashita H, Kukita J, Ohga S, et al. Serum erythropoietin levels in
term and preterm infants during the first year of life. Am J Pediatr Hematol
Oncol. 1994;16:213-218.
different types of globin exhibits distinctive functional
properties. Fetal erythrocytes, which contain mostly Hb
F, have a higher oxygen affinity than adult red blood cells.
This allows the transport of oxygen from maternal Hb
A–containing erythrocytes across the placenta to fetal red
blood cells. The increased O2 affinity of fetal hemoglobin
has been ascribed to the diminished interactions of Hb F
with red blood cell 2,3-BPG. Embryonic erythrocytes also
display a greater affinity for oxygen than adult cells.
ERYTHROPOIETIN, ERYTHROPOIESIS, AND
THE PHYSIOLOGIC ANEMIA OF INFANCY
Erythropoietin (EPO), the essential glycoprotein growth
factor for erythropoiesis, binds to erythropoietin recep recep-
tors on early erythroid progenitor cells and via the JAK2
signaling pathway ay regulates RBC production by protect
protect-
ing them from apoptosis. Erythropoietin is produced pri- pri
marily in the fetal liver and later in the cortical peritubular
cells of the kidney, so that in adults renal production of
EPO is the most important. Erythropoiesis is highly
responsive to blood oxygenation. Hypoxia inducible
factors (HIFs), constitutively expressed EPO transcription
factors,, are destroyed in the presence of oxygen. Under
hypoxic conditions, EPO production increases. Levels of
EPO in cord blood are higher than in adult blood samples
nopqk rrstlu
((Table 88-3),
), but there is a dramatic decrease after birth
in response to higher levels of tissue oxygenation. By 1
month of age, serum levels in healthy term infants reach
their nadir. This is followed by a rise to maximal levels at
2 months of age and then a slow drift down to adult
values.
The postnatal changes in tissue oxygenation and erytheryth-
ropoietin production result in a physiologic anemia of
infancy with a mean minimal hemoglobin concentration
in healthy term infants of about 11 g/dL at 8 to 12 weeks
(fghijk 88-4
of life (Figure 88-4;v nopqk
Table rrswlm
88-4).
). Because of the shorter
life span of RBCs in preterm infants with low EPO levels,
the nadir is noted by 6 weeks of age and ranges from 7
to 10 g/dL. In VLBW and ELBW infants, the nadir is more
than 20% below the value of the Hb at birth. In ELBW
infants whose nadir falls below 7 g/dL, this so-called
physiologic anemia of prematurity can be associated with
pallor, tachypnea, tachycardia, poor feeding, and poor
weight gain.x2 yz{kj |oi}k} ~ pq~~€ q~}} o€ }i‚‚jk}}g~
of erythropoiesis in the ill neonate can contribute to
more severe and earlier anemia. Although preterm infants
will respond to hypoxia with a rise in EPO levels, the
 1298 MNOQ 14 • THE BLOOD AND HEMATOPOIETIC SYSTEM

increase is lower than that expected for term infants. The term infants, the mean capillary hemoglobin at birth is
suboptimal EPO response may be due to developmental 19.3 g/dL (see nopqk
Table rrswlm
88-4).
). The Hct has a mean of 61 g/
changes in transcription factors or to the site of fetal EPO dL. Premature infants have lower Hb levels than do full-
production. The use of recombinant EPO in premature term infants. In addition to gestational age, Hb levels are
and sick newborn infants is discussed later. influenced by a variety of factors that must be kept in
mind when analyzing the neonate with anemia or poly poly-
RED BLOOD CELL INDICES cythemia. One important determinant is the site of sam sam-
DURING PRENATAL AND pling: Capillary Hb values are higher than peripheral
POSTNATAL DEVELOPMENT venous samples, and umbilical venous Hb results are the
The RBC count, Hb concentration, and hematocrit (Hct) lowest. The interval between delivery and clamping of
increase throughout gestation, as shown in nopqk rrs¤m. ‰
In the umbilical cord and the height of the baby relative to
the placenta can significantly affect a newborn’s blood
volume and total RBC mass. The placenta contains about
100 mL of blood. The mean blood volume of a full-term
20 infant is about 85 mL/kg. Early or delayed clamping of
the umbilical cord alters this mean blood volume by
about 10% lower or higher, respectively. The average Hb
15 at birth is relatively unchanged; however, 48 hours later,
Hb (g/dL)

after redistribution of plasma volume, Hb values will

reflect the lower or higher red cell mass. Racial differences
10 also occur. One study reported significantly higher Hb,
Hct, and MCV in white infants compared with black
5 infants of similar gestational ages.„3  kzg|iq~|¡zk
Reticulocyte |~iz}
counts g
in
the cord blood of infants average 4% to 5%, and nucle nucle-
ated RBCs are evident in most cord blood samples
£ (40,000/μ μL).
(40,000/μL).L). These findings are presumed to reflect high
0 25 50 75 100 125 EPO production secondary to low oxygen retention in
Days utero. Infants who experience placental insufficiency and
Figure 88-4 Hemoglobin concentrations in full-term and premature
intrauterine growth restriction have higher than normal
infants. (●¢ Full-term infants; (▲¢ premature infants, birth weight 1200 EPO production and an even greater degree of erythrocy
erythrocy-
to 2350 g; (■¢ premature infants, birth weight less than 1200 g. (Adapted tosis. The mean MCV of RBCs in the newborn is increased.
from Oski F. The erythrocyte and its disorders. In: Nathan D, et al, eds. The RBCs of the neonate have an increased Hb content,
Hematology of infancy and childhood. 4th ed. Philadelphia; Saunders; but the mean corpuscular hemoglobin concentration
1993:18.) (MCHC) is comparable to that of adultsadults.

TABLE 88-4 Red Blood Cell Values (Capillary Samples) for Term Infants
During the First 12 Weeks of Life
Hb (g/dL) RBC (×1012/L) Hematocrit (%) MCV (fl) MCHC (g/dL) Reticulocytes (%)
žŸ[ ± SD ± SD ± SD ± SD ± SD ± SD
Days
1 19.3 ± 2.2 5.14 ± 0.7 61 ± 7.4 119 ± 9.4 31.6 ± 1.9 3.2 ± 1.4
2 19.0 ± 1.9 5.15 ± 0.8 60 ± 6.4 115 ± 7.0 31.6 ± 1.4 3.2 ± 1.3
3 18.8 ± 2.0 5.11 ± 0.7 62 ± 9.3 116 ± 5.3 31.1 ± 2.8 2.8 ± 1.7
4 18.6 ± 2.1 5.00 ± 0.6 57 ± 8.1 114 ± 7.5 32.6 ± 1.5 1.8 ± 1.1
5 17.6 ± 1.1 4.97 ± 0.4 57 ± 7.3 114 ± 8.9 30.9 ± 2.2 1.2 ± 0.2
6 17.4 ± 2.2 5.00 ± 0.7 54 ± 7.2 113 ± 10.0 32.2 ± 1.6 0.6 ± 0.2
7 17.9 ± 2.5 4.86 ± 0.6 56 ± 9.4 118 ± 11.2 32.0 ± 1.6 0.5 ± 0.4
Weeks
1-2 17.3 ± 2.3 4.80 ± 0.8 54 ± 8.3 112 ± 19.0 32.1 ± 2.9 0.5 ± 0.3
2-3 15.6 ± 2.6 4.20 ± 0.6 46 ± 7.3 111 ± 8.2 33.9 ± 1.9 0.8 ± 0.6
3-4 14.2 ± 2.1 4.00 ± 0.6 43 ± 5.7 105 ± 7.5 33.5 ± 1.6 0.6 ± 0.3
4-5 12.7 ± 1.6 3.60 ± 0.4 36 ± 4.8 101 ± 8.1 34.9 ± 1.6 0.9 ± 0.8
5-6 11.9 ± 1.5 3.55 ± 0.4 36 ± 6.2 102 ± 10.2 34.1 ± 2.9 1.0 ± 0.7
6-7 12.0 ± 1.5 3.40 ± 0.4 36 ± 4.8 105 ± 12.0 33.8 ± 2.3 1.2 ± 0.7
7-8 11.1 ± 1.1 3.40 ± 0.4 33 ± 3.7 100 ± 13.0 33.7 ± 2.6 1.5 ± 0.7
8-9 10.7 ± 0.9 3.40 ± 0.5 31 ± 2.5 93 ± 12.0 34.1 ± 2.2 1.8 ± 1.0
9-10 11.2 ± 0.9 3.60 ± 0.3 32 ± 2.7 91 ± 9.3 34.3 ± 2.9 1.2 ± 0.6
10-11 11.4 ± 0.9 3.70 ± 0.4 34 ± 2.1 91 ± 7.7 33.2 ± 2.4 1.2 ± 0.7
11-12 11.3 ± 0.9 3.70 ± 0.3 33 ± 3.3 88 ± 7.9 34.8 ± 2.2 0.7 ± 0.3

Hb, Hemoglobin; MCHC, mean corpuscular hemoglobin concentration; MCV, mean corpuscular volume; RBC, red blood cells.
Data from Matoth Y, Zaizov R, Varsano I. Postnatal changes in some red cell parameters. Acta Paediatr Scand. 1971;60:317-323.
 // • HEMATOLOGIC AND ONCOLOGIC PROBLEMS IN THE FETUS AND NEONATE 1299

TABLE 88-5 Red Blood Cell Values (Arterial Samples) on First Postnatal Day at Different
Gestational Ages®
Group 1 Group 2 Group 3
23-25 wk 26-28 wk 29-31 wk
°UY±U²[› (N = 40) (N = 60) (N = 88)
Hematocrit (%) 43.5 ± 4.2³ ´µ¶· ± 4.5¸ ´¹¶· ± 5.0¸†
º»¼¶·½ 43.8, 51.0) (37.5, 45.0, 54.3) (39.4, 47.6, 56.0)
Hemoglobin (g/dL) 14.5 ±1.6 15.1 ± 1.6¸ ¾¼¶¿ ± 1.7¸†
º¾¿¶·½ 14.7, 17.4) (12.5, 15.0, 18.3) (13.2, 16.1, 18.8)
Mean corpuscular hemoglobin (pg) 38.6 ± 2.2³ »¹¶» ± 2.0 37.3 ± 2.5³
º»µ¶·½ 38.6, 43.0) (33.4, 38.4, 43.2) (32.0, 37.5, 40.6)
Mean corpuscular volume (fl) 115.6 ± 5.6³ ¾¾´¶· ± 7.6¸ ¾¾·¶´ ± 6.6³‡
º¾·À¶·½ 114.5, 125.7) (98.4, 114.0, 126.6) (97.3, 111.2, 120.0)
Mean corpuscular hemoglobin concentration (g/dL) 33.4 ± 0.9 33.6 ± 0.6 33.7 ± 0.7
(32.3, 33.3, 34.6) (32.3, 33.6, 34.6) (32.5, 33.6, 34.9)
Red cell distribution width 15.9 ± 1.4 16.5 ± 1.9 16.4 ± 1.5
(14.2, 15.6, 18.5) (14.5, 16.0, 21.0) (14.6, 16.0, 19.4)

*Values are reported as mean ± standard deviation and 5th, 50th, and 95th percentiles in parentheses.
†
P value of <.01 between groups 1 and 3.
‡
P value of <.01 between groups 2 and 3.
Data from Alur P, Devapatla SS, Super DM, et al. Impact of race and gestational age on red blood cell indices in very low birth weight infants. Pediatrics. 2000;106:306-310.

Delayed (30-90 seconds) cord clamping continues to TABLE 88-6 ÁÂÃÃerences in Neonatal Red
generate considerable interest because it has been shown Blood Cells Compared with
to prevent hypotension, raise hematocrit, and decrease
the need for transfusions in preterm infants. In term
Adult Red Blood Cells
infants who have had delayed cord clamping, there has ÄÅÆ (mean corpuscular volume) ↑
been a reduction in iron deficiency anemia in the first RBC count ↑
year of life, but an increased risk of early jaundice.…§¨ MCHC (mean corpuscular hemoglobin concentration) ↑
Surface area ↑
©^ª BLOOD CELL SURVIVAL Reticulocyte count ↑
The normal life span of adult RBCs is about 120 days. Resting cell diameter ↑
Hemoglobin F content ↑
The life span of RBCs in newborns at term is 60 to 80
Whole cell deformability ↔
days and 30 to 50 days in ELBW infants. In general, red Suction pressure for complete aspiration ↑
blood cell survival is affected by changes related to aging Sensitivity to osmotic lysis ↑
(senescence) and by random hemolysis of red blood ATP utilization ↑
cells, or portions of red blood cells, in the spleen and the Glucose utilization ↑
rest of the reticuloendothelial system. Some of the Catalase glutathione peroxidase ↓
changes in neonatal RBCs compared with adult RBCs Susceptibility to oxidant injury ↑
listed in nopqk rrs« ok|z }ij¬g¬oqm ­hgh kj¡z{j~|¡zk} Phospholipid, lipid, cholesterol content ↑
with declining RBC enzyme activity become progressively Loss of volume, surface area and deformability with age ↑
less tolerant of oxidative challenges during the transpor- Permeability to sodium and potassium ↑
tation of oxygen molecules and exposure to circulating I antigen expression on cell surface ↑
oxidants. Any additional deficiencies in the enzymatic A, B, H blood group antigens on cell surface ↓
Life span ↓
pathways of the RBC may affect the ability of the eryth-
rocyte to tolerate oxidative challenges and further reduce ↑, Increased; ↓, decreased; ↔, same.
red blood cell survival.
al. With transit through the kidneys Adapted from Linderkamp O, Nash GB, Paul YK, et al. Deformability and intrinsic
and lungs, the RBCs experience cycles of osmotic swelling material properties of neonatal red blood cells. Blood. 1966:67:1244-1250.
and shrinkage. Shear forces in high-pressure areas of the
circulation buffet the erythrocytes. Each passage through
the cords of Billroth within the spleen requires the RBCs ok¥go because of decreased erythropoiesis, enhanced
to deform and squeeze through tiny slits in the walls of splenic filtration, and activation of phagocytes.
the cords or face destruction if they cannot. Congenital
or acquired defects in membrane stability or decreases in
the ratio of surface area to red blood cell volume will also Red Blood Cell Disorders
decrease erythrocyte survival. Alterations in the deform-
ability of neonatal erythrocytes and relative intolerance ¦e^_d¦
to oxidative challenges result in shorter survival for neo- Anemia is defined by a hemoglobin or hematocrit value
natal red blood cells. Random hemolysis can be increased that is more than two standard deviations below the
with splenic enlargement or activation of the phagocytic mean for age. In the neonate, the causes of anemia can
system. Infants with hemolysis may have exaggerated be divided into two broad categories: anemia resulting
1300 MNOQ 14 • THE BLOOD AND HEMATOPOIETIC SYSTEM

BOX 88-1 ANEMIAS BY ETIOLOGY BOX 88-2 CLASSIFICATION OF THE

ÊËËÌÍÌÎÊÏÌÐ ÍÑÒÒ ANEMIAS ACCORDING TO
MEAN CORPUSCULAR VOLUME
• Hemorrhage
• Fetal ÓÊËÎÑËÔÏÕË ÊÖÌÓÕÊ
• Fetal-maternal • Reticulocytosis
• Placental
• Traumatic delivery • Folic acid deficiency
• Coagulation defects • B12 deficiency
• Early umbilical cord clamping • Bone marrow failure syndromes
• Twin-twin transfusion • Diamond-Blackfan anemia
• Pearson syndrome
• Excess phlebotomy losses • Fanconi anemia
ACCELERATED DESTRUCTION • Down syndrome
• Hemolytic anemia • Myelodysplastic syndrome
• Immune • Liver disease
Alloimmune: Rh, ABO, minor blood group • Drugs (phenytoin, mercaptopurine)
Autoimmune • Hypothyroidism
• Nonimmune
• Hemoglobinopathy MICROCYTIC, HYPOCHROMIC ANEMIA
• Thalassemia • Iron deficiency
• Unstable hemoglobin • Thalassemia
• Red blood cell enzyme defect • Chronic infection
• Structural defect of red blood cell membrane • Hb E trait
• Mechanical destruction • Sideroblastic anemia
• Microangiopathic hemolytic anemia • Copper deficiency
• Infection • Defects of iron metabolism
• Vitamin E deficiency • Lead poisoning
DIMINISHED RED BLOOD CELL PRODUCTION NORMOCYTIC ANEMIA
• Congenital • Low reticulocyte count
• Diamond-Blackfan anemia • Acute blood loss
• Pearson syndrome • Infection
• Fanconi anemia • Parvovirus B19
• Congenital dyserythropoietic anemias • Transient erythroblastopenia of childhood
• Anemia of prematurity • Chronic disease
• Acquired • Drugs
• Parvovirus B19 • Leukemia
• Transient erythroblastopenia of childhood • Bone marrow infiltration
• Human immunodeficiency virus • Liver disease
• Syphilis • Renal failure
• Iron deficiency • Aplastic anemia, acquired
• Lead toxicity • Normal or high reticulocyte count
• Infection • Blood loss
• Sequestration
• Red blood cell enzyme defects
• Immune hemolytic anemia
from accelerated loss or destruction of red blood cells and • Mechanical hemolytic anemia
anemia caused by a defect at some stage of red blood cell • Red cell membrane defects
88-1lm
Ç~È 88-1).
production ((Box ). The defects may be congenital or • Unstable hemoglobin
• Hemoglobinopathy
acquired, and the abnormality may be intrinsic to the
RBCs or extrinsic. Anemias also may be categorized on a
morphologic basis. Using the normal range of the MCV
for age and gestation, the anemia may be characterized age but often include the medical and dietary history
as microcytic, normocytic or macrocytic (Ç~È 88-2lm of the pregnancy, the estimated gestational age at birth,
Hypochromicity, abnormal RBC shapes (poikilocytes), the chronologic age, the infant’s diet, and details of any
polychromasia, and cell inclusions (e.g., basophilic stip- previous anemia, blood loss,, transfusions, medications,
pling or Howell-Jolly bodies) also provide clues to the and illnesses,, as well as the family history of anemia. The
etiology of the anemia (nopqk rrsÉlm physical examination should evaluate the infant’s general
health, growth, and development. Identification of any
Evaluation of Anemia dysmorphic features, abnormal masses, or skin lesions
The clinician begins the evaluation of anemia by taking a can aid the diagnosis ((Tablenopqk rrsrlm
88-8).
). The patient also
thorough history. Appropriate data vary with the patient’s should be assessed for jaundice, hepatosplenomegaly,
 // • HEMATOLOGIC AND ONCOLOGIC PROBLEMS IN THE FETUS AND NEONATE 1301

TABLE 88-7 Morphologic Findings on Peripheral Blood Smears

×XYØÙXXgic Abnormality Etiology
Acanthocytes Alteration of lipid bilayers
Liver disease
Abetalipoproteinemia
Blister cells G6PD deficiency
Basophilic stippling Ineffective erythropoiesis: iron deficiency, lead poisoning, thalassemia, nonimmune hemolytic anemias
Elliptocytes Structural defects of red cell membrane: hereditary elliptocytosis
Heinz bodies Precipitated hemoglobin: normal in newborn; nonimmune hemolytic anemias
Howell-Jolly bodies Splenic hypofunction or post splenectomy
Hypochromia Iron deficiency, thalassemias, lead poisoning
Nucleated red blood cells Normal in newborn; hemolytic anemias, semi-acute blood loss
Polychromasia Normal in newborn; reticulocytosis
Pyropoikilocytosis Neonates with hereditary elliptocytosis, hereditary pyropoikilocytosis, thermal injury of red cells (burn)
Rouleaux Increased fibrinogen, inflammation
Schistocytes Microangiopathic hemolytic anemias
Sickle cells Hemoglobin SS and sickle variants
Spherocytes Decreased cell membrane: volume—IgG+ hemolytic anemia, hereditary spherocytosis, artifact of area of
blood smear
Target cells Increased red blood cell surface: volume ratio
Alteration in lipid structure of red blood cell membrane
Hemoglobin C, hemoglobin S, thalassemias, liver disease, abetalipoproteinemia

TABLE 88-8 Physical Findings in Neonatal Anemia

šÙڛ±VU R±œÛ±œŸ Etiology
“Blueberry muffin” spots Extramedullary hematopoiesis, replacement of bone marrow by tumor, congenital infection
Cardiac disease/mechanical heart valve “Waring blender” syndrome
Congestive heart failure Chronic anemia
Dysmorphic features Bone marrow failure syndromes: Diamond-Blackfan anemia, Fanconi anemia, Shwachman-Diamond
syndrome, Pearson syndrome
Down syndrome
Myelodysplastic syndrome
Failure to thrive Pearson syndrome
Shwachman-Diamond syndrome
Hepatosplenomegaly Congenital infection, storage disorder, malignancy, hypersplenism, hemangioma, hemolytic anemia,
transient abnormal myelopoiesis
Jaundice Hemolytic anemia
Kaposiform hemangioendothelioma Kasabach-Merritt syndrome
Microcephaly Congenital infection
Bone marrow failure syndrome
Short stature Congenital bone marrow failure syndrome

vascular malformations, cardiovascular function, and
lymphadenopathy.
The initial laboratory evaluation includes a complete
blood count (CBC) with RBC indices, a reticulocyte
count, and evaluation of the peripheral blood smear
(fghijk 88-5
(Figure lm
88-5).
). The results of the preliminary laboratory
testing, combined with information from the history and
physical examination, should dictate the need for further
tests, such as hemoglobin analysis of the infant or parents,
CBCs and blood smears of the parents, analysis of hepatic
or renal function, direct or indirect antiglobulin (Coombs)
testing, cultures or titers to identify infectious agents, a
bone marrow aspirate or biopsy, osmotic fragility tests,
and quantitative or qualitative testing for glucose-6-phos-
phate dehydrogenase (G6PD) deficiency.
The neonatal patient presents the diagnostician with a
number of unique challenges. Because of the small total
blood volume of the infant, who already is anemic,
testing must be limited. Major pediatric medical centers
can perform many of the necessary tests on very small
quantities of blood, especially if the tests are appropri-
ately batched when they are submitted. Because of the
many morphologic and biochemical differences in neo-
natal and adult RBCs (see nopqk rrs«lu some diagnoses
are best made by testing the parents for evidence of
disease or carrier states. At times, a definitive diagnosis
can be made only with repeat testing later in infancy,
when the infant would be expected to have a much higher
percentage of adult-type RBCs or when the infant has
recovered from an acute hemolytic crisis that may have
destroyed the older, more biochemically or morphologi-
cally abnormal cells.
Rationale for Transfusion Therapy
See Chapter 89. Because the primary function of the RBC
is to transport oxygen from the pulmonary bed to other
 1302 MNOQ 14 • THE BLOOD AND HEMATOPOIETIC SYSTEM

Reticulocyte count

ÜÝÞ Normal or high

ßÝàáâàãäåæ åçæåèäãé Ýê ëìçÝçæåèäãé
åàâíãå
îéïðãêâñ åçæåèäãé åàâíãå
òàñÝéêãàâ ñâóâéäè
Toxin
Chronic disease
Infection (B19 parvovirus)
Nutritional deficiency (iron)
ôÝàâ íåêêÝÞ êâçæåéâíâàä
õÜâðöâíãå÷ àâðêÝøæåèäÝíåù
Direct and indirect antiglobulin test

Negative Positive
Immune hemolytic anemia

MCV

Low Normal or high

α-Thalassemia

Peripheral blood smear

Normal Abnormal
Congenital deficiency red Hemoglobinopathy
cell enzymes Microangiopathic hemolytic anemia
Blood loss Congenital structural defect of
Infection red cell membrane Figure 88-5 Algorithm for diagnosis of
Splenic sequestration G6PD deficiency anemia in the neonate. MCV, Mean corpuscular
Mechanical hemolysis volume.

zg}}ik} for release, anemia diminishes oxygen-carrying
capacity and can compromise tissue oxygenation. Tissue
oxygenation is a complex concept involving not only the
Hb concentration but also the oxygen affinity of the Hb
in the patient’ss red blood cells, blood viscosity, and the
patient’ss cardiorespiratory status. The only absolute indi-
indi
cations for rapidly correcting anemia by RBC transfusion
are to restore tissue oxygenation and to expand blood
volume after severe, acute loss. In most pediatric centers,
the sicker patients, especially those with cardiopulmo-
nary dysfunction, receive transfusions to maintain the Hb
closer to normal for age. Neonatal exchange transfusions
are also performed with the goal of replacing “doomed”
infant RBCs with healthy adult RBCs, which have superior
oxygen-transporting ability. This has the triple benefit of
limiting hyperbilirubinemia and other byproducts of RBC
breakdown, reducing the body load of maternal antibod
antibod-
ies, and supplementing with cells that contain Hb A.
Anemia Caused by Blood Loss
Blood loss can occur in the fetus, at birth, or in the post
post-
natal period. The bleeding can be acute or chronic.
Anemia caused by chronic blood loss generally is better
tolerated because the neonate will at least partly compen
compen-
sate for a gradual reduction in RBC mass. Chronic blood
loss can be diagnosed by identifying signs of compensa
compensa-
tion. Doppler assessment of the fetal middle cerebral
artery peak systolic velocity is a noninvasive method for
determination of fetal anemia, independent of the etioletiol-
ogy. The infant may be pale and may exhibit signs and
symptoms of congestive heart failure. Anemia will be
present, often with reticulocytosis, hypochromia, and
microcytosis.
An infant with acute blood loss may not be anemic if
blood sampling is done soon enough after the acute
event so that hemodilution has not yet occurred. Anemia
usually develops within 3 to 4 hours after blood loss;
repeat testing 6 to 12 hours after the event should reveal
the true extent of the loss. In acute blood loss, the infant
may exhibit signs and symptoms of hypovolemia and
hypoxemia (e.g., tachycardia, tachypnea, hypotension).
The RBCs should be morphologically normal. With either
kind of hemorrhage, infants tend to have fewer problems
with hyperbilirubinemia because they have a reduced
RBC mass. Jaundice can result if entrapped RBCs in a
hematoma breakdown.
 // • HEMATOLOGIC AND ONCOLOGIC PROBLEMS IN THE FETUS AND NEONATE
Internal bleeding can occur if the fetus has anatomic
abnormalities or defects in the hemostatic system, or with
interventional obstetric procedures. A surprisingly large
amount of blood can be lost within a cephalohematoma,
subaponeu
and even greater bleeding can occur in the subaponeu-
rotic area of the scalp (subgaleal hemorrhage), where
Trau
bleeding is not limited by periosteal attachments. Trau-
matic or assisted deliveries and vitamin K deficiency are
commonly associated with such bleeding. Full-term
infants may have intracranial bleeding, which usually
occurs in the subarachnoid or subdural regions. Full-term
infants with intracranial hemorrhage should be evaluated
bleed
for hemostasis abnormalities because this type of bleed-
ing is associated with qualitative and quantitative platelet
defects and with abnormalities of several of the coagula-
tion proteins. Hemorrhage into the adrenals, kidneys,
liver, spleen, or retroperitoneum also can occur after dif-
ficult or breech deliveries. Splenic or hepatic rupture can
occur after trauma, especially if the organs are enlarged
as a result of extramedullary hematopoiesis. Occult or
superficial vascular tumors can bleed and sequester large
volumes of red blood cells and platelets.
Maternal factors can cause prenatal blood loss.
Maternal history of vaginal bleeding, placenta previa,
abruptio placentae, nonelective cesarean delivery, and
cord compression are associated with anemia. Hemor-Hemor
rhage from the umbilical cord may be the result of
intrinsic vascular abnormalities, inflammation of the
cord, velamentous insertion of the cord, coagulation
defects, or an unusually short cord. A normal cord can
rupture during a precipitous or assisted delivery or if it
becomes tangled around the infant. Accidental incision
of the placenta during delivery also can result in
bleeding.
Fetal-Maternal Hemorrhage
Hemorrhage. Fetal cells may be found
in the maternal circulation in about half of all pregnan
pregnan-
cies. 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 pregnan
pregnan-
cies.„ú
35
fkzoqs¥ozkjoq
Fetal-maternal {k¥~jj{ohk
hemorrhage g} is o}}~|gozk€ gz{ pro
associated with ‚j~-
pro-
cedures 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 incompatibil
incompatibil-
ity, 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 hemo-
lytic disease of the newborn. Vaginal or peripheral blood
samples from women with vaginal bleeding late in preg-
nancy 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.
1303
Maternal red blood cells are incubated with anti-D and
then washed. Indicator D-cells are added, and in the pres-
ence of fetal D-positivee 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 condi-
tion, including manyy 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. Although it offers improved
precision and accuracy, flow cytometry is expensive
and requires specially trained technicians. Most centers
cannot offer this testing 24 hours a day. The wider use of
hematology analyzers with multiparameter flow cytom-
etry technology may allow more laboratories to offer this
testing in the future.„ú
ûüýþÿûüýþ Transfusion Syndrome.
Syndrome See Chapters 22
and 27. In monochorionic diamniotic multiple gesta gesta-
tions, twin-twin transfusion syndrome (TTTS) can be diagdiag-
nosed by ultrasound demonstrating oligohydramnios in
the donor sac and polyhydramnios in the recipient sac.
Twin-twin transfusion syndrome occurs in 8% to 10% of
twin pregnancies with monochorionic diamniotic pla pla-
centation and accounts for about half of the perinatal
ˆ6 nTwin
deaths.61 g ok¥go
anemia ‚~q¡|¡z{k¥go
polycythemia }k ik|k
(TAPS)
sequence n­l g}
is
defined as the presence of anemia in the donor and polypoly-
cythemia 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 interfe
interfe-
tal 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, neuro-
logic, and dev
developmental disorders have been associated
with TTTS.ˆ616
]Hytic Anemia: Accelerated Red
Blood Cell Destruction
Accelerated destruction of RBCs is the end point of a
number of intrinsic, extrinsic, congenital, and acquired
RBC abnormalities. Because the RBCs of premature
infants and newborns have a shorter life span, hemolysis
is defined as a process that shortens the survival of the
RBCs relativee to the expected life span for the infant’s
gestational and postnatal age. In contrast to anemia
caused by blood loss, most infants with hemolysis have
some evidence of indirect hyperbilirubinemia and ele- ele
vated lactate dehydrogenase for age. Reticulocytosis
should accompany the hemolysis, although in conditions
complicated by bone marrow suppression (congenital
infections, chronic illness, or nutritional deficiency) or