erytHroCyte produCtion

The erythrocyte is a vehicle for the transport of hemoglobin, which is

produced in precursor cells of the erythrocytes, the normoblasts. The
function of hemoglobin is the transport of oxygen and carbon dioxide.

NORMOBLASTIC MATURATION
The earliest recognizable erythroid precursor is the pronormoblast
The nucleus has a fine, uniform chromatin pattern that is somewhat
More distinct and More intensely stained than that of the myeloblast.

The pronormoblast
undergoes mitosis and forms two basophilic normoblasts.
The basophilic normoblast is somewhat smaller and has
slightly coarser chromatin that stains intensely;

After mitosis of the basophilic normoblast, evidence of continuing

hemoglobin production becomes visible in the cytoplasm of the two
daughter cells as polychromasia, that is, mixtures of the red-staining of
hemoglobin with the blue of RNA in varying shades of gray. This cell is
the polychromatophilic normoblast
The polychromatophilic normoblast Undergoes one Or two mitotic divisions.
After the last mitosis, the nucleus becomes small and dense (pyknotic),
and the orthochromatic normoblast stage is reached. Mitosis
is no longer possible.

Finally, accompanied by cytoplasmic contractions and undulations, the

nucleus and a small rim of cytoplasm are ejected from the orthochromatic
normoblast, forming the reticulocyte

In the marrow, developing erythroid cells are usually in contact with

macrophages in what are termed erythroblastic islands.
During proliferation and maturation, iron is transferred from plasma
transferrin into the cells in the normoblastic series. The pronormoblast
and the basophilic normoblast have the highest content of RNA, which
begins to decline in the polychromatophilic normoblasts as hemoglobin
increases in amount. Synthesis of RNA gradually decreases in each stage
through the orthochromatic normoblasts. When the nucleus is no longer
present (in the reticulocyte), RNA synthesis ceases, yet the RNA already
present remains for a few days, and protein and heme synthesis continue
in the reticulocyte until the cell loses its RNA and mitochondria.

During this maturation process, three or four mitotic divisions occur

in a period of 3 days, resulting in the potential production of 16 reticulocytes
from each pronormoblast. The Reticulocytes are larger than mature red cells
and remain in the marrow stroma for 1–2 days before being released into the blood.

In the marrow, the reticulocytes are about equal in number to the

nucleated erythrocytes and slightly greater in number than the reticulocytes
in the circulating blood. If Sufficiently severe hypoxia is present, this
marrow pool of reticulocytes can be released. This approximately doubles
the number of circulating reticulocytes.

Normally, reticulocytes remain as such, slowly synthesizing hemoglo-

bin, for 2–3 days in the marrow and for 1 day in the blood. Residual
ribosomes, mitochondria, and other organelles are then removed, and the
mature erythrocytes circulate for about 120 days.

MEGALOBLASTIC MATURATION
Abnormal maturation of erythroid precursors that occurs in vitamin B
deficiency or folic acid deficiency is known as megaloblastic maturation,
and the abnormal erythroid cells are called megaloblasts. Because of
impaired ability of the cells to synthesize DNA, the intermitotic and
mitotic phases are prolonged. This results in enlarged cells, with nuclear
maturation lagging behind cytoplasmic maturation (nuclear–cytoplasmic
dissociation
Karyorrhexis, or breaking up of the nucleus, and Howell-Jolly bodies are frequently noted.
Megaloblastic development parallels normoblastic maturation

REGULATION OF ERYTHROCYTE
PRODUCTION
The number of erythrocytes in the blood may be regulated by changing
the rate of production. The rate of erythrocyte destruction does not vary
appreciably in normal individuals. Increased production of erythrocytes
occurs when oxygen transport to the tissues is impaired, as in anemia, in
cardiac or pulmonary disorders, and in the low oxygen tension of high
altitudes. Erythrocyte production decreases when an individual is hypertransfused
or exposed to high oxygen tension.

Oxygen affinity of hemoglobin is modulated by the concentration of

phosphates, in particular 2,3-diphosphoglycerate (2,3-DPG) in the red
cell. These phosphates combine with the -chains of reduced hemoglobin
and diminish its affinity for oxygen (Fig. 31-8). In areas of tissue hypoxia,
as oxygen moves from hemoglobin into the tissues, the amount of reduced
hemoglobin in the red cells increases, binding more 2,3-DPG, further
reducing its oxygen affinity so that more oxygen can be delivered to the
tissues. If hypoxia persists, depletion of free 2,3-DPG leads to increased
glycolysis, production of more 2,3-DPG, and a persistently lower oxygen
affinity of the hemoglobin.

Tissue hypoxia induces formation of EPO via production of the transcriptional

factor, hypoxia-inducible factor-1(HIF-1).

EPO effects the production of more red cells in the bone marrow. It acts
by inducing committed progenitor cells (CFU-E and BFU-E) in the
marrow to proliferate and differentiate into pronormoblasts by shortening
the generation time of normoblasts, and by promoting early release of
reticulocytes into the blood. The result is increased numbers of marrow
normoblasts in a normal ratio of cell types, a condition known as normoblastic
hyperplasia. Increased Cellular expression of HIF-1
can result from a 598CT mutation in the von Hippel–Lindau gene (the von Hippel–
Lindau protein is involved in HIF-1 degradation). This results in elevation

of EPO levels and an autosomal recessive form of hereditary

polycythemia known as Chuvash polycythemia

Elevated levels are detected in patients with secondary polycythemia and in those
with aplastic anemia. Decreased levels below the normal range are found
in normal individuals after transfusion and in those with primary polycythemia
(polycythemia vera). However, considerable overlap exists, and
normal EPO levels may be found in both primary and secondary polycythemia
Anti-EPO Antibodies have been described in pure red cell aplasia and systemic lupus erythematosus

SYNTHESIS OF HEMOGLOBIN
Heme Synthesis
Heme synthesis occurs in most cells of the body, except the mature erythrocytes,
but most abundantly in the erythroid precursors. Succinylcoenzyme
A condenses with glycine to form the unstable intermediate -amino -ketoadipic acid, which
Is readily decarboxylated to -aminolevulinic acid (ALA)
Globin Synthesis
Globin synthesis occurs in the cytoplasm of the normoblast and reticulocyte.

Control of hemoglobin synthesis is exerted primarily through the

action of heme. Increased heme inhibits further heme synthesis by inhibiting
the activity and synthesis of ALA synthase. Heme Also promotes globin synthesis,
mainly at the site of chain initiation, and the interaction of
ribosomes with mRNA.

STRUCTURE AND FUNCTION OF

HEMOGLOBIN
Normal adult HbA consists of four heme groups and four polypeptide chains (two
-chains and two -chains), which form a roughly globular hemoglobin
molecule (Fig. 31-11).

The sigmoid-shaped oxygen dissociation curve of hemoglobin reflects

this increasing affinity for oxygen with increasing partial pressure of
oxygen in the lungs (see Fig. 31-8). In the tissues, conversion of oxygenated
Hb to Hb, decreasing pH, and increasing temperature produced by metabolic
processes, as well as the binding of more 2,3-DPG To Hb, Result in a shift of the
Hb-oxygen dissociation
curve to the right, favoring the release of oxygen from hemoglobin.

erytHroCyte destruction
The erythrocyte gradually undergoes metabolic changes over the course
of its 120-day life span, at which time the less viable senescent cell is
removed from the circulation.

Older red cells have a smaller surface area and

an increased mean cell hemoglobin concentration (MCHC) compared
with younger cells. Furthermore, aged red cells lose sialic acid from their
membranes, exposing an asialoglycophorin. This senescent antigen is recognized
and an autoantibody is synthesized by the host. After binding
of the autoantibody, the senescent cell is removed from the circulation by the mononuclear
phagocyte system, primarily within the spleen. About 3 million cells are normally removed from the blood per second with no
demonstrable histologic evidence of erythrophagocytosis.

DEGRADATION OF HEMOGLOBIN
After removal of the red cell from the circulation, hemoglobin is broken
down within the macrophages of the mononuclear phagocyte system into
its three constituents: iron, protoporphyrin, and globin. The iron goes into
storage and may be completely reutilized. The globin may be degraded
and returned to the amino acid pool of the body. In contrast, the protoporphyrin
ring is split, converted to bilirubin, and excreted from the body.
In the macrophage, the protoporphyrin ring is cleaved by a heme
oxidase enzyme at the 
-methene bridge, yielding 1 mol of carbon monoxide

(CO) and 1 mol of biliverdin. The CO appears in the blood as

HbCO and is eventually exhaled. Biliverdin is reduced to bilirubin in the
macrophage, and bilirubin is transported to the liver by plasma albumin.
It is removed from the plasma by the liver cell, conjugated mainly with
glucuronide, and excreted in the bile. In the intestine, reduction by bac-
teria occurs, and bilirubin is transformed into urobilinogen, mesobiliru-
binogen, and stercobilinogen, compounds that are collectively designated
urobilinogens.

ErytHrokinetiCS
The balance between delivery of erythrocytes to the blood and removal of
erythrocytes from the blood results in a relatively constant hemoglobin
mass in the circulation. Anemia occurs when removal of erythrocytes from
the blood is increased and cannot be compensated for by increased production,
or.when delivery of erythrocytes to the blood is decreased, or when both processes exist together.
When anemia develops, tissue hypoxia leads to elevated levels of erythropoietin in the plasma. Resultant
normoblastic hyperplasia produces
more erythrocytes for delivery to the circulation. The marrow in a normal
individual is capable of six to eight times the normal output of erythrocytes
with extreme stimulation. This capacity must be compared with the output
actually attained when one is evaluating the marrow response of a given
patient.

MEASUREMENTS OF TOTAL PRODUCTION

OF ERYTHROCYTES OR HEMOGLOBIN
The total mass of erythropoietic cells in the body cannot be easily
measured. An estimate is made by examining a sample of bone marrow
from a normally active site and determining the cellularity and the percentage
of total nucleated cells that are erythropoietic . When marrow activity increases, usually the
additional hematopoietic cells replace the fat in the red marrow sites before
extension occurs into the yellow marrow of the long bones. One assumes
that the sample is representative of the marrow as a whole—an assumption
that usually is valid.

The plasma iron turnover is calculated from the serum iron level and
the rate of removal of injected radioactive iron from the plasma.

MEASUREMENTS OF TOTAL DESTRUCTION

OF ERYTHROCYTES OR HEMOGLOBIN
Determination of fecal urobilinogen is an estimate of the total excretion
of bile pigments—the breakdown products of heme. This measurement
includes pigment derived from hemoglobin formed and destroyed in the
marrow without ever reaching the circulating erythrocytes.

Increased urinary urobilinogen is seen with severe liver disease, as well as

with conditions associated with increased heme catabolism that overwhelm
the metabolic capacity of the liver.

MEASUREMENTS OF EFFECTIVE
PRODUCTION OF ERYTHROCYTES

Reticulocyte Count
Because the RNA of the reticulocyte disappears about a day after its entry
into the blood, enumeration of reticulocytes will be a measure of the
number of cells being delivered by the marrow to the blood each day, that
is, a measure of effective erythropoiesis. The absolute reticulocyte count
is calculated by multiplying the reticulocyte percentage by the erythrocyte
count.

second consideration is an adjustment for increased maturation time

of reticulocytes in the blood as a result of accelerated release from the
marrow, an effect of erythropoietin. The need for this is recognized by the
presence of large, polychromatic cells or nucleated red cells in the blood
film, indicating a shift of excessively immature reticulocytes from the
marrow into the blood. To avoid an overestimate of daily erythrocyte
production, a correction factor is used that is based on estimated maturation
time of reticulocytes in the blood.
The erythrocyte utilization of iron is a measure of the amount of an
injected dose of iron that appears in the hemoglobin of circulating erythrocytes.
This, too, is A measure of Effective erythropoiesis.

MEASUREMENTS OF EFFECTIVE SURVIVAL

OF ERYTHROCYTES IN BLOOD
The erythrocyte survival can be determined by removing a sample of
blood, labeling the erythrocytes with chromium-51 (excess 51Cr), inactivating the Cr remaining in the plasma,
and reinjecting the labeled erythrocytes into the patient. The results are usually expressed as the
Cr half-survival time. The normal range is 28–38 days.
the erythrocyte survival is also a measure of effective production of erythrocytes.