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Chapter 26 Biology of Erythropoiesis, Erythroid Differentiation, and Maturation 301

Fig. 26.2 ERYTHROID MATURATION SEQUENCE. As proliferation parameters (i.e., rates of deoxyri-
bonucleic acid [DNA] and ribonucleic acid [RNA] synthesis) and cell size decrease, accumulation of erythroid-
specific proteins (i.e., heme and globin) increases, and the cells adapt their morphologic characteristics.
(Modified from Granick S, Levere R: Heme synthesis in erythroid cells. In Moore CV, Brown EB, editors: Progress in
hematology, Vol 4, Orlando, FL, 1964, Grune & Stratton, p 1.)

enucleated, the reticulocyte. During this last mitosis, the inactive recruited as part of the stress response. 82,83,105,106 Genetic heterogeneity
dense nucleus of the orthochromatic erythroblast moves to one side in the control of gene expression of these receptors may add another
of the cell and is extruded, encased by a thin cytoplasmic layer, the layer of variability in recovery from anemia in humans.
pyrenocyte that is ingested by marrow macrophages. By contrast, the The importance of GR in the control of stress erythropoiesis was
majority of the cytoplasm and plasma proteins form the reticulocyte established by studies in transgenic mice harboring a dimerization-
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that is released in the blood stream to further mature into red blood defective GR (GR dim/dim mice). These mice have normal steady-state
cell. Although all mammals have enucleated cells in their circulation, erythropoiesis but were unable to increase red blood cell production
the evolutionary advantage of enucleation is not readily apparent. It in response to hypoxia. Gene deletion studies established that GR
may allow for more red cell deformability when traveling through the facilitates stress erythropoiesis in mice by blocking maturation of
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small vasculature, or it may minimize cardiac workload. erythroid precursors and inducing a limited self-renewal state.
Maturation from proerythroblast to reticulocyte likely does not Although clinical observations indicating that the GR ligand such
always adhere to a rigid sequence in which each division is associated as dexamethasone stimulates erythropoiesis have been available since
with the production of two more differentiated and morphologically 1961, 109,110 the precise role of GR in human erythropoiesis is still
distinct daughter cells (i.e., basophilic erythroblast gives rise to two unclear. Murine GR is not polymorphic, whereas human GR (GR/
polychromatophilic ones). Rather, significant flexibility, both in the NR3C1 located in the 5q31-32 region of chromosome 5 and deleted
number and rate of divisions and in the rate of enucleation, may be in 5q-syndrome) contains several single-nucleotide polymorphisms
allowed. Such deviations from the normal orderly maturation sequence (SNPs). 111–114 Because of this genetic diversity, human cells may
may be dictated by the level of EPO or “stress” conditions. Thus in express more than 260 isoforms with slightly or greatly different
cases of acute demand for red cell production (because of blood loss biologic activities. The most studied isoform is GRα, an isoform
or hemolysis), the kinetics of formation of new reticulocytes are similar to the murine GR. Alternative splicing between exon 3-4
significantly more rapid. Resulting red cells may be larger (i.e., with generates GRγ, an isoform containing an additional arginine in the
increased mean corpuscular volume). This has led to the concept of DNA-binding domain that reduces the transactivation potential by
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“skipped” divisions. The orderly unilineage differentiation pathway half. An alternative splicing of exon 9 generates messenger RNA
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shown in Fig. 26.1 is likely restricted to conditions of steady-state (mRNA) encoding the dominant-negative GRβ isoform. It is
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hematopoiesis. Similar to occurrences in the lymphoid system, alter- debatable whether an isoform with dominant-negative action similar
native routes are taken under conditions of “stress.” Murine models to GRβ exists in mice. 112,115 It is generally accepted that responses to
have been developed to address phenotype-function cell relationships GR ligands depend on the signal transduction potential of the GR
during recovery from acute and chronic erythroid stress. A model isoforms expressed by different cells and tissues. Studies in human
for acute stress is represented by the hemolytic anemia induced by nonerythroid cell types have identified that GR isoform expression
phenylhydrazine treatment. Recovery from this acute anemia involves predicts the variegation of cellular response to dexamethasone in
recruitment of the spleen as an additional erythropoietic site and vitro. 116–118 Recently clinicians have established important correla-
3
is dependent on EPO. The amount of H-thymidine incorporated tions between GR haplotype and variability in patients’ responses to
by splenic erythroblasts produced in response to this stress initially glucocorticoids and in the development of glucocorticoid resistance
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represented the biologic assay for EPO. Genetic evidence indicates in several disorders. GR polymorphism and/or epigenetic changes
that recovery from this hemolytic anemia is controlled by a receptor are emerging as the leading cause for dexamethasone unresponsive-
complex formed between the EPOR and a truncated version of the ness or for development of dexamethasone resistance in patients
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s
Stk receptor encoded by Fv2 , a locus that also determines strain with inflammatory and autoimmune diseases (i.e., Crohn disease,
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susceptibility to Friend virus infection. An additional control systemic lupus) and in chronic depression. 119,121–123
on the response to acute erythroid stress in mice is exerted by the Similarly, several in vitro and in vivo studies suggest that
glucocorticoid receptor (GR), because mice in which this receptor is variegation of GR isoform expression may also have biologic and
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targeted recover poorly from phenylhydrazine treatment. On the clinical effects on terminal erythroid maturation. The numbers of
other hand, experimentally induced mutations in genes involved in erythroblasts generated by murine erythroid progenitors in response
the regulation of erythroid differentiation, such as signal transducer to dexamethasone is fairly consistent, whereas the number generated
and activator of transcription 5 (STAT5 null100 ) and GATA1 LOW101 , by human erythroid progenitors from different individuals may vary
or inability of response to reactive oxygen species (ROS) challenge by 1 to 2 logs, 45,49 likely reflecting the genetic background of human
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(i.e., Foxo3 deficiency) increase the rate of erythroblast apoptosis. GR. In addition, the frequency of the rs6198 SNP is greater than
The spleen is also recruited as a hemopoietic site in response to normal in patients with the Philadelphia-negative myeloproliferative
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chronic erythroid stress. 103,104 Several studies in aggregate suggest neoplasm polycythemia vera (PV) (55%, p = .0028) and primary
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that the erythron does not respond to stress only by amplifying the myelofibrosis and with Diamond-Blackfan anemia (DBA) (43%,
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normal erythroid progenitor cell compartments (i.e., CMP, MEP, p = .03), suggesting that genetic conditions favoring GRβ expres-
and CFU-E), but by generating alternative routes of differentiation, sion may represent host genetic modifiers in diseases with altered
possibly through cooperation between EPOR and other receptors terminal erythroid differentiation. A retrospective analysis of 499
(e.g., Stk, GR, soluble KL, BMP4/Hedgehog pathway) specifically patients with primary myelofibrosis (PMF) indicated that the rs6198