ERYTHROKINETICS their cells does not unload oxygen to the tissues
readily, so newborns are slightly hypoxic compared
• Erythrokinetics is the term describing the dynamics
of RBC production and destruction.
• Erythron is the name given to the collection of all
stages of erythrocytes throughout the body: the
developing precursors in the bone marrow and the
circulating erythrocytes in the peripheral blood and
the vascular spaces within specific organs such as
the spleen. When the term erythron is used, it
conveys the concept of a unified functional tissue.
*The erythron is distinguished from the RBC mass. The
erythron is the entirety of erythroid cells in the body,
whereas the RBC mass refers only to the cells in
circulation.
Hypoxia—the stimulus to red blood cell production
• As mentioned previously, the role of RBCs is to carry
oxygen. To regulate the production of RBCs for that
purpose, the body requires a mechanism for sensing
whether there is adequate oxygen being carried to
the tissues. If not, RBC production and the functional
efficiency of existing cells must be enhanced. Thus a
second feature of the oxygen-sensing system must
be a mechanism for influencing the production of
RBCs.
• The primary oxygen-sensing system of the body is
located in peritubular fibroblasts of the
kidney. Hypoxia, too little tissue oxygen, is detected
by the peritubular cells, which
produce erythropoietin (EPO), the major stimulatory
cytokine for RBCs. Under normal circumstances, the
amount of EPO produced fluctuates very little,
maintaining a level of RBC production that is
sufficient to replace the approximately 1% of RBCs
that normally die each day. When there is
hemorrhage, increased RBC destruction, or other
factors that diminish the oxygen-carrying capacity of
the blood, the production of EPO is increased.
Hypoxia and Red Blood Cell Production
• Decreased RBC number, however, is only one
cause of hypoxia. Another cause is the failure of
each RBC to carry as much oxygen as it should.
This can occur because the hemoglobin is defective
or because there is not enough hemoglobin in each
cell. The hypoxia may be unrelated to the RBCs in
any way; poor lung function resulting in diminished
oxygenation of existing RBCs is an example.
• The kidney’s hypoxia sensor cannot know why there
is hypoxia, but it does not matter. Even when there
are plenty of RBCs compared with the reference
interval, if there is still hypoxia, stimulation of RBC
production is warranted because the numbers
present are not meeting the oxygen need. An
elevation of RBC numbers above the reference
interval, erythrocytosis, is seen in conditions such as
lung disease and cardiac disease in which the blood
is not being well oxygenated. Newborns have higher
numbers of RBCs because the fetal hemoglobin in
with adults. To compensate, they make more RBCs.
•
• Hypoxia increases EPO production in peritubular
cells mainly by transcriptional regulation. The EPO
gene has a hypoxia-sensitive region (enhancer) in
its 3′ regulatory component.15 When oxygen tension
in the cell decreases, hypoxia-inducible factor-1, a
transcription factor, is assembled in the
cytoplasm,16 migrates to the nucleus, and interacts
with the 3′ enhancer of the gene. This results in
transcription of more EPO messenger RNA
molecules, and production of more EPO.
Erythropoietin
Structure. 
EPO is a thermostable, nondialyzable, glycoprotein
hormone with a molecular weight of 34 kD.17 It consists of
a carbohydrate unit that reacts specifically with RBC
receptors and a terminal sialic acid unit, which is
necessary for biological activity in vivo.18 On desialation,
EPO activity ceases.1
Action. 
EPO is a true hormone, being produced at one location
(the kidney) and acting at a distant location (the bone
marrow). It is a growth factor (or cytokine) that initiates an
intracellular message to the developing RBCs; this
process is called signal transduction. EPO must bind to
its receptor on the surface of cells to initiate the signal or
message. The receptor is a transmembrane homodimer
consisting of two identical polypeptide chains.20 EPO-
responsive cells vary in their sensitivity to EPO.21 Some
are able to respond to low levels of EPO,22 whereas
others require higher levels. In healthy circumstances
when RBC production needs to proceed at a modest but
regular rate, the cells requiring only low levels of EPO
respond. If EPO levels rise secondary to hypoxia,
however, a larger population of EPO-sensitive cells is
able to respond.
• The binding of EPO, the ligand, to its receptor on
erythrocyte progenitors initiates a cascade of
intracellular events (“the program”) that ultimately
leads to cell division, maturation, and more red
blood cells entering the circulation. EPO’s effects are
mediated by Janus-activated tyrosine kinase 2
(JAK2) signal transducers that are associated with
the cytoplasmic domain of the EPO receptor and
ultimately affect gene expression in the RBC
nucleus. 23 EPO has three major effects: allowing
early release of reticulocytes from the bone marrow,
preventing apoptotic cell death, and reducing the
 time needed for cells to mature in the bone marrow.
The essence is that EPO puts more RBCs into the
circulation at a faster rate than occurs without its
stimulation.
Early release of reticulocytes.  among other things, the degradation of chromatin
• EPO promotes early release of developing erythroid
precursors from the marrow by two mechanisms.
EPO induces changes in the adventitial cell layer of
the marrow/sinus barrier that increase the width of
the spaces for RBC egress into the sinus.24 This
mechanism alone, however, is insufficient for cells to
leave the marrow. RBCs are held in the marrow
because they express surface membrane receptors
for adhesive molecules located on the bone marrow
stroma. EPO downregulates the expression of these
receptors so that cells can exit the marrow earlier
than they normally would. The result is the presence
in the circulation of reticulocytes that are still very
basophilic because they have not spent as much
time degrading their ribosomes or making
hemoglobin as they normally would before entering
the bloodstream. These are called shift
reticulocytes because they have been shifted from
the bone marrow early (Figure 8-8, A). Their bluish
cytoplasm with Wright stain is evident, so the overall
blood picture is said to have polychromasia. Even
nucleated RBCs (i.e., normoblasts) can be released
early in cases of extreme anemia when the demand
for RBCs in the peripheral circulation is great.
Releasing cells from the marrow early is a quick fix,
so to speak; it is limited in effectiveness because the
available precursors in the marrow are depleted
within several days and still may not be enough to
meet the need in the peripheral blood for more cells.
A more sustained response is required in times of
increased need for RBCs in the circulation.
Inhibition of apoptosis.
• A second, and probably more important, mechanism
by which EPO increases the number of circulating
RBCs is by increasing the number of cells that will
be able to mature into circulating erythrocytes. It
does this by decreasing apoptosis, the programmed
death of RBC progenitors.
Apoptosis: Programmed cell death.  signal. Apoptosis of RBCs is a cellular process that
• As noted previously, it takes about 18 to 21 days to
produce an RBC from stimulation of the earliest
erythroid progenitor (BFU-E) to release from the
bone marrow. In times of increased need for RBCs,
such as when there is loss from the circulation
during hemorrhage, this time lag would be a
significant problem. One way to prepare for such a
need would be to maintain a store of mature RBCs
in the body for emergencies. RBCs cannot be stored
in the body for this sort of eventuality, however,
because they have a limited life span. Therefore,
instead of storing mature cells for emergencies, the
body produces more CFU-Es than needed at all
times. When there is a basal or steady-state
demand for RBCs, the extra progenitors are allowed
to die. When there is an increased demand for
RBCs, however, the RBC progenitors have about an
8- to 10-day head start in the production process.
This process of intentional wastage of cells occurs
by apoptosis, and it is part of the cell’s genetic
program.
Process of apoptosis.
• Apoptosis is a sequential process characterized by,
into fragments of varying size that are multiples of
180 to 185 base pairs long; protein clustering; and
activation of transglutamase. This is in contrast to
necrosis, in which cell injury causes swelling and
lysing with release of cytoplasmic contents that
stimulate an inflammatory response (Chapter 6).
Apoptosis is not associated with inflammation.
• During the sequential process of apoptosis, the
following morphologic changes can be seen:
condensation of the nucleus, causing increased
basophilic staining of the chromatin; nucleolar
disintegration; and shrinkage of cell volume with
concomitant increase in cell density and compaction
of cytoplasmic organelles, while mitochondria remain
normal.28 This is followed by a partition of cytoplasm
and nucleus into membrane-bound apoptotic bodies
that contain varying amounts of ribosomes,
organelles, and nuclear material. The last stage of
degradation produces nuclear DNA fragments
consisting of multimers of 180 to 185 base pair
segments. Characteristic blebbing of the plasma
membrane is observed. The apoptotic cell contents
remain membrane-bound and are ingested by
macrophages, which prevents an inflammatory
reaction. The membrane-bound vesicles display so-
called “eat me” signals on the membrane surface
(discussed later) that promote macrophage
ingestion.29
Evasion of apoptosis by erythroid progenitors and
precursors.
• One effect of EPO is an indirect avoidance of
apoptosis by removing an apoptosis induction
depends on a signal from either the inside or outside
of the cell. Among the crucial molecules in the
external messaging system is the death receptor
Fas on the membrane of the earliest RBC
precursors, while its ligand, FasL, is expressed by
more mature RBCs. When EPO levels are low, cell
production should be at a low rate because hypoxia
is not present. The excess early erythroid precursors
should undergo apoptosis. This occurs when the
older FasL-bearing erythroid precursors, such as
polychromatic normoblasts, cross-link with Fas-
marked immature erythroid precursors, such as
pronormoblasts and basophilic normoblasts, which
are then stimulated to undergo apoptosis.28 As long
as the more mature cells with FasL are present in
the marrow, erythropoiesis is subdued. If the FasL-
bearing cells are depleted, as when EPO stimulates
early marrow release, the younger Fas-positive
precursors are allowed to develop, which increases
the overall output of RBCs from the marrow. Thus
 early release of older cells in response to EPO
indirectly allows more of the younger cells to mature.
• A second mechanism for escaping apoptosis exists
for RBC progenitors: direct EPO rescue from
apoptosis. This is the major way in which EPO is
able to increase RBC production. When EPO binds
to its receptor on the CFU-E, one of the effects is to
reduce production of Fas ligand.31 Thus the younger
cells avoid the apoptotic signal from the older cells.
Additionally, EPO is able to stimulate production of
various anti-apoptotic molecules, which allows the
cell to survive and mature.31, 32 The cell that has the
most EPO receptors and is most sensitive to EPO
rescue is the CFU-E, although the late BFU-E and
early pronormoblast have some receptors.33 Without
EPO, the CFU-E does not survive.34
Reduced marrow transit time.
• Apoptosis rescue is the major way in which EPO increases
RBC mass—by increasing the number of erythroid cells that
survive and mature to enter the circulation. Another effect of
EPO is to increase the rate at which the surviving precursors
can enter the circulation. This is accomplished by two
means: increased rate of cellular processes and decreased
cell cycle times.
•  Among the processes that are accelerated is
hemoglobin production. As mentioned earlier,
another accelerated process is bone marrow egress
with the loss of adhesive receptors and the
acquisition of egress-promoting surface molecules.
The other process that is accelerated is the
cessation of division. Cell division takes time and
would delay entry of cells to the circulation, so cells
enter cell cycle arrest sooner. As a result, the cells
spend less time maturing in the marrow.  enzyme function to live 120 days. Because RBCs
• EPO also can reduce the time it takes for cells to
mature in the marrow by reducing individual cell
cycle time, specifically the length of time that cells
spend between mitoses.
• With the decreased cell cycle time and fewer mitotic
divisions, the time it takes from pronormoblast to
reticulocyte can be shortened by about 2 days total.
If the reticulocyte leaves the marrow early, another
day can be saved, and the typical 6-day transit time
is reduced to fewer than 4 days under the influence
of increased EPO.
Measurement of erythropoietin.
• Quantitative measurements of EPO are performed
on plasma and other body fluids. EPO can be
measured by chemiluminescence. Although the
reference interval for each laboratory varies, an
example reference interval is 4 to 27 mU/
L.40 Increased amounts of EPO in the urine are
expected in most patients with anemia, with the
exception of patients with anemia caused by renal
disease.
Therapeutic uses of erythropoietin. 
• Recombinant erythropoietin is used as therapy in
certain anemias such as those associated with
chronic kidney disease and chemotherapy. It is also
used to stimulate RBC production prior to
autologous blood donation and after bone marrow
transplantation. 
• Unfortunately, some athletes illicitly use EPO
injections to increase the oxygen-carrying capacity
of their blood to enhance endurance and stamina,
especially in long-distance running and cycling. The
use of EPO is one of the methods of blood doping,
and aside from being banned in organized sports
events, it increases the RBC count and blood
viscosity to dangerously high levels and can lead to
fatal arterial and venous thrombosis
Other stimuli to erythropoiesis
• In addition to tissue hypoxia, other factors influence
RBC production to a modest extent. It is well
documented that testosterone directly stimulates
erythropoiesis, which partially explains the higher
hemoglobin concentration in men than in women.
Also, pituitary42 and thyroid43 hormones have been
shown to affect the production of EPO and so have
indirect effects on erythropoiesis.
Erythrocyte destruction
• All cells experience the deterioration of their
enzymes over time due to natural catabolism. Most
cells are able to replenish needed enzymes and
continue their cellular processes. As a nonnucleated
cell, however, the mature erythrocyte is unable to
generate new proteins, such as enzymes, so as its
cellular functions decline, the cell ultimately
approaches death. The average RBC has sufficient
lack mitochondria, they rely on glycolysis for
production of adenosine triphosphate (ATP). The
loss of glycolytic enzymes is central to this process
of cellular aging, called senescence, which
culminates in phagocytosis by macrophages. This is
the major way in which RBCs die normally.
Macrophage-mediated hemolysis (extravascular
hemolysis)
• At any given time, a substantial volume of blood is in
the spleen, which generates an environment that is
inherently stressful on cells. Movement through the
red pulp is sluggish. The available glucose in the
surrounding plasma is depleted quickly as the cell
flow stagnates, so glycolysis slows. The pH is low,
which promotes iron oxidation. Maintaining reduced
iron is an energy-dependent process, so factors that
promote iron oxidation cause the RBC to expend
 more energy and accelerate the catabolism of
enzymes.
• In this hostile environment, aged RBCs succumb to
the various stresses. Their deteriorating glycolytic
processes lead to reduced ATP production, which is
• Although most natural RBC deaths occur in the
complicated further by diminished amounts of
available glucose. The membrane systems that rely
on ATP begin to fail. Among these are enzymes that
maintain the location and reduction of phospholipids
of the membrane. Lack of ATP leads to oxidation of
membrane lipids and proteins. Other ATP-dependent
enzymes are responsible for maintaining the high
level of intracellular potassium while pumping
sodium out of the cells. As this system fails,
intracellular sodium increases and potassium
decreases. The effect is that the selective
permeability of the membrane is lost and water
enters the cell. The discoid shape is lost and the cell
becomes a sphere
Mechanical hemolysis (fragmentation or intravascular
hemolysis)
spleen, a small portion of RBCs
rupture intravascularly (within the lumen of blood
vessels). The vascular system can be traumatic to
RBCs, with turbulence occurring in the chambers of
the heart or at points of bifurcation of vessels. Small
breaks in blood vessels and resulting clots can also
trap and rupture cells. The intravascular rupture of
RBCs from purely mechanical or traumatic stress
results in fragmentation and release of the cell
contents into the plasma; this is
called fragmentation or intravascular hemolysis.

• RBCs must remain highly flexible to exit the spleen • When the membrane of the RBC has been
breached, regardless of where the cell is located
by squeezing through the so-called splenic
when it happens, the cell contents enter the
sieve formed by the endothelial cells lining the
surrounding plasma. Although mechanical lysis is a
venous sinuses and the basement membrane.
relatively small contributor to RBC demise under
Spherical RBCs are rigid and are not able to
normal circumstances, the body still has a system of
squeeze through the narrow spaces; they become
plasma proteins, including haptoglobin and
trapped against the endothelial cells and basement
hemopexin, to salvage the released hemoglobin so
membrane. In this situation, they are readily
that its iron is not lost in the urine.
ingested by macrophages that patrol along the
sinusoidal lining

• Some researchers view erythrocyte death as a

nonnucleated cell version of apoptosis,
termed eryptosis,49 that is precipitated by oxidative
stress, energy depletion, and other mechanisms that
create membrane signals that stimulate
phagocytosis. It is highly likely that there is no single
signal but rather that macrophages recognize
several.

• When an RBC lyses within a macrophage, the major

components are catabolized. The iron is removed
from the heme. It can be stored in the macrophage
as ferritin until transported out. The globin of
hemoglobin is degraded and returned to the
metabolic amino acid pool. The protoporphyrin
component of heme is degraded through several
intermediaries to bilirubin, which is released into the
plasma and ultimately excreted by the liver in bile.