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Chapter 39 Megaloblastic Anemias 523

compensate for reduced thymidine). This results in a high dUMP/ Morphology in Megaloblastosis from Cobalamin and Folate Deficiency
dTMP ratio and an increase in deoxyuridine triphosphate (dUTP), Is the Same
which can get misincorporated into DNA. At this juncture, an
editorial enzyme recognizes this faulty misincorporation and excises Peripheral Smear
dUTP. However, with a continued inadequate supply of deoxythy- • Increased mean corpuscular volume (MCV) with macro-
midine triphosphate (dTTP), there is a continued cycle of uracil ovalocytes (up to 14 µm), which is variously associated with
misincorporation into DNA in folate deficiency, 90–92 its removal by anisocytosis and poikilocytosis
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uracil-DNA glycosylase, and refilling of the missing base by DNA • Nuclear hypersegmentation of polymorphonuclear neutrophils
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polymerase β. However, with repetition over several cycles, multiple (PMNs) (one PMN with six lobes or 5% with five lobes)
single-strand nicks are introduced into DNA; this predisposes to • Thrombocytopenia (mild to moderate)
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chromosome breaks that can contribute to an increased risk of cancer • Leukoerythroblastic morphology (from extramedullary
associated with folate deficiency. 91,95,96 In addition, folate deficiency hematopoiesis)
can also lead to double-strand breaks in DNA, which are difficult to Bone Marrow Aspirate
repair when the two nicks are close to one another (within 12 bp of • General increase in cellularity of all three major hematopoietic
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each other) on opposite strands. Collectively, such double-strand elements
DNA breaks in folate-deficient cells predispose to the development • Abnormal erythropoiesis—orthochromatic megaloblasts
of acentric chromosomes, DNA fragments, and micronuclei. 94,98 • Abnormal leukopoiesis—giant metamyelocytes and “band” forms
This can even render folate-deficient tissues more permissive to (pathognomonic), hypersegmented PMNs
the integration of HPV16 DNA, and trigger (experimental) • Abnormal megakaryocytopoiesis—pseudo hyperdiploidy
carcinogenesis. 99

Chromosome and Cell Cycle Defects proerythroblasts that develop into later megaloblastic forms, 80% to
90% die in the bone marrow. Marrow macrophages effectively
Defective DNA synthesis caused by folate deficiency is reflected by scavenge dead or partially disintegrated megaloblasts. This is the basis
numerous chromosomal abnormalities, including abnormalities in for ineffective erythropoiesis (intramedullary hemolysis).
telomeres, which correlate with biomarkers of chromosomal instabil- Leukopoiesis is also abnormal. There is an absolute increase in
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ity and mitotic dysfunction. There is excessive chromosomal these cells, which are large and have similar sieve-like chromatin.
elongation with despiralization associated with random breaks and Spectacular giant (20 to 30 µm) metamyelocytes and “band” forms
exaggerated centromere constriction, expression of folate-sensitive that are often seen are pathognomonic for megaloblastosis (see Fig.
fragile sites in hematopoietic cells, and reduced biosynthesis, acetyla- 39.7). There may be bizarre nucleoli with small cytoplasmic vacuoles.
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tion, and methylation of arginine-rich histone. All this leads to It is probable that giant metamyelocytes cannot easily traverse marrow
perturbation of the cell cycle with an increased proportion of cells in sinuses, and their maturation into circulating hypersegmented poly-
prophase of the mitotic cycle and G 2 that leads to apoptosis of ery- morphonuclear neutrophils (PMNs) is unlikely. Granulation of the
throid precursors and anemia. 22 cytoplasm remains unaffected.
Megakaryocytes may be normal or increased in numbers and may
exhibit additional complexities in megaloblastic expression (see Fig.
MORPHOLOGIC EXPRESSION OF MEGALOBLASTOSIS 39.7). Complex hypersegmentation (i.e., pseudohyperdiploidy) is
associated with liberation of fragments of cytoplasm and giant
There is widening disparity in nuclear-cytoplasmic asynchrony as a platelets into the circulation. The net output of platelets is decreased
cobalamin- or folate-deficient cell divides, until the more mature in severe megaloblastosis, and abnormal but reversible platelet dys-
generations of daughter cells die in the marrow or are arrested (as function has been documented. 22
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megaloblastic cells) at various stages of the cell cycle. The plethora In early cobalamin or folate deficiency, normoblasts may dominate
of bone marrow morphologic changes can lead an untrained observer the marrow with only a few megaloblasts seen. Complete transforma-
to the diagnosis of erythroleukemia. All proliferating cells exhibit tion to megaloblastic hematopoiesis is observed in florid cases and is
megaloblastosis, including the luminal epithelial mucosal cells of the reflected by various degrees of pancytopenia.
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entire gastrointestinal tract, cervix, vagina, and uterus. However, The earliest manifestation of megaloblastosis is an increase in
megaloblastic changes are most striking in the blood and bone mean corpuscular volume (MCV) with macro-ovalocytes (up to
marrow. Ineffective hematopoiesis extends into long bones, and the 14 µm) (see Fig. 39.7). Because these cells have adequate hemoglobin,
bone marrow aspirate (which is superior to the biopsy for observing the central pallor, which normally occupies about one-third of the
megaloblastosis) exhibits trilineal hypercellularity, especially of the cell, is decreased. By contrast, thin macrocytes have larger than
erythroid series. The appearance of exuberant cell proliferation with normal central pallor (Table 39.1). In severe anemia, poikilocytosis
numerous mitotic figures is misleading because these cells are actually and anisocytosis are evident. Cells containing remnants of DNA (i.e.,
proliferating very slowly (see box on Morphology in Megaloblastosis Howell-Jolly bodies), arginine-rich histone, and nonhemoglobin iron
from Cobalamin and Folate Deficiency Is the Same). (i.e., Cabot rings) may be observed. Extramedullary megaloblastic
Erythroid hyperplasia reduces the myeloid-to-erythroid ratio from hematopoiesis may also result in a leukoerythroblastic picture.
3 : 1 to 1:1. Proerythroblasts are not as obviously abnormal as later Nuclear hypersegmentation of DNA in PMNs strongly suggests
forms; they may simply be larger (promegaloblasts). Megaloblastic megaloblastosis when associated with macro-ovalocytosis (see Fig.
changes are most strikingly displayed in intermediate and orthochro- 39.7). Normally fewer than 5% of PMNs have more than five lobes,
matic stages, which are larger than their normoblastic counterparts. and no cells have more than six lobes in the peripheral blood. If
In contrast to the normally dense chromatin of comparable normo- megaloblastosis is suspected (greater than 5% PMNs with more than
blasts, megaloblastic erythroid precursors have an open, finely stippled, five lobes or a single PMN with more than six lobes), a formal lobe
reticular, sieve-like pattern (Fig. 39.7). The orthochromatic megalo- count/PMN (i.e., lobe index) above 3.5 may be obtained.
blast, with its hemoglobinized cytoplasm, continues to retain its large Ineffective use of iron results in an increased percentage of satura-
sieve-like immature nucleus, in sharp contrast to the clumped chro- tion of transferrin and increased iron stores. If there is associated iron
matin of orthochromatic normoblasts. The nucleus is often eccentri- deficiency, the MCV may be normal, and only iron therapy can
cally placed in these large oval or oblong cells, and lobulation or unmask the megaloblastic manifestations in the peripheral blood. In
indentation of nuclei with bizarre karyorrhexis is often seen. In cells thalassemia, the entire erythrocyte morphology normally expected in
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destined for the circulation as macro-ovalocytes, the nucleus may megaloblastosis is masked ; however, megaloblastic leukopoiesis is
occasionally not be completely extruded. Of the potential progeny of still observed. Significant intramedullary hemolysis (ineffective