MATURATION SEQUENCE

PRONORMOBLAST Basophilic normoblast (prorubricyte)

Nucleus. 
• Nucleus.  The chromatin begins to
condense, revealing clumps along the
The nucleus takes up much of the cell (N:C ratio of periphery of the nuclear membrane and a
8:1). The nucleus is round to oval, containing one or few in the interior. As the chromatin
two nucleoli. The purple red chromatin is open and condenses, the parachromatin areas
contains few, if any, fine clumps.
become larger and sharper, and the N:C
• Cytoplasm.  ratio decreases to about 6:1. The
chromatin stains deep purple-red. Nucleoli
The cytoplasm is dark blue because of the may be present early in the stage but
concentration of ribosomes. The Golgi complex may disappear later.
be visible next to the nucleus as a pale, unstained
area. Pronormoblasts may show small tufts of
irregular cytoplasm along the periphery of the Cytoplasm. 
membrane. When stained, the cytoplasm may be a
deeper, richer blue than in the
• Division.  pronormoblast—hence the
The pronormoblast undergoes mitosis and gives rise to name basophilic for this stage.
two daughter pronormoblasts. More than one division
is possible before maturation into basophilic Division. 
normoblasts. The basophilic normoblast undergoes
mitosis, giving rise to two daughter cells.
• Location. 
More than one division is possible before
The pronormoblast is present only in the bone marrow the daughter cells mature into
in healthy states. polychromatic normoblasts.

• Cellular activity.  Location. 

The pronormoblast begins to accumulate the
components necessary for hemoglobin production.
The proteins and enzymes necessary for iron uptake
and protoporphyrin synthesis are produced. Globin
production begins.3
• Length of time in this stage.  occurs,3 but the many cytoplasmic
This stage lasts slightly more than 24 hours.3
The basophilic normoblast is present only in
the bone marrow in healthy states.
Cellular activity. 
Detectable hemoglobin synthesis
organelles, including ribosomes and a
substantial amount of messenger
ribonucleic acid (RNA; chiefly for
hemoglobin production), completely mask
the minute amount of hemoglobin
pigmentation.
Length of time in this stage.  • Length of time in this stage. 
This stage lasts slightly more than 24
hours.3 This stage lasts approximately 30 hours.3

Polychromatic (polychromatophilic) normoblast Orthochromic normoblast (metarubricyte)

(rubricyte)
• Nucleus. 

The nucleus is completely condensed (i.e., pyknotic) or

• Nucleus.  nearly so. As a result, the N:C ratio is low or
approximately 1:2.
The chromatin pattern varies during this stage of
development, showing some openness early in the • Cytoplasm. 
stage but becoming condensed by the end. The
condensation of chromatin reduces the diameter of The increase in the salmon-pink color of the cytoplasm
the nucleus considerably, so the N:C ratio decreases reflects nearly complete hemoglobin production. The
from 4:1 to about 1:1 by the end of the stage. residual ribosomes react with the basic component of
Notably, no nucleoli are present. the stain and contribute a slightly bluish hue to the
cell, but that fades toward the end of the stage as the
• Cytoplasm.  organelles are degraded.

This is the first stage in which the pink color associated
with stained hemoglobin can be seen. The stained
color reflects the accumulation of hemoglobin
pigmentation over time and concurrent decreasing
amounts of RNA. The color produced is a mixture of
pink and blue, resulting in a murky gray-blue. The
stage’s name refers to this combination of multiple
colors, because polychromatophilic means “many
color loving.”
• Division. 
The orthochromic normoblast is not capable of
division due to the condensation of the chromatin.
• Location. 
The orthochromic normoblast is present only in the
bone marrow in healthy states.

• Division.  • Cellular activity. 

This is the last stage in which the cell is capable of
undergoing mitosis, although likely only early in the
stage. The polychromatic normoblast goes through
mitosis, producing daughter cells that mature and
develop into orthochromic normoblasts.
• Location.  holding organelles in proper location in the cytoplasm,
The polychromatic normoblast is present only in the
bone marrow in healthy states.
• Cellular activity.  the nucleus enveloped by cell membrane.4Nonmuscle
Hemoglobin synthesis increases, and the accumulation
begins to be visible in the color of the cytoplasm.
Cellular organelles are still present, particularly
ribosomes, which contribute a blue aspect to the
cytoplasm. The progressive condensation of the
nucleus and disappearance of nucleoli are evidence of
progressive decline in transcription of
deoxyribonucleic acid (DNA).
Hemoglobin production continues on the remaining
ribosomes using messenger RNA produced earlier.
Late in this stage, the nucleus is ejected from the cell.
The nucleus moves to the cell membrane and into a
pseudopod-like projection. As part of the maturation
program, loss of vimentin, a protein responsible for
is probably important in the movement of the nucleus
to the cell periphery.1 Ultimately, the nucleus-
containing projection separates from the cell by having
the membrane seal and pinch off the projection with
myosin of the membrane is important in this pinching
process.5 The enveloped extruded nucleus, called
a pyrenocyte,1 is then engulfed by bone
marrow macrophages. The macrophages recognize
phosphatidlyserine on the pyrenocyte surface as an
“eat me” flag.6 Other organelles are extruded and
ingested in similar fashion. Often, small fragments of
nucleus are left behind if the projection is pinched off
before the entire nucleus is enveloped. These
fragments are called Howell-Jolly bodies when seen in
peripheral blood cells (Table 19-3 and Figure 19-1) and
are typically removed from the cells by the splenic
macrophage pitting process once the cell enters the
circulation.
• Length of time in this stage.  film is made (). The residual ribosomes appear as a
This stage lasts approximately 48 hours.3
Polychromatic (polychromatophilic) erythrocyte or reticulocyte
• Nucleus.  immediately preceding the mature erythrocyte, even
Beginning at the polychromatic erythrocyte stage,
there is no nucleus. The polychromatic erythrocyte is a
good example of the prior statement that a cell may
not have all the classic features described but may be
staged by the preponderance of features. In particular,
when a cell loses its nucleus, regardless of cytoplasmic
appearance, it is a polychromatic erythrocyte.
• Cytoplasm.  erythrocyte because it lacks a nucleus and is no longer
The cytoplasm can be compared with that of the late
orthochromic normoblast in that the predominant
color is that of hemoglobin. By the end of the
polychromatic erythrocyte stage, the cell is the same
color as a mature RBC, salmon pink. It remains larger
than a mature cell, however. The shape of the cell is
not the mature biconcave disc but is irregular in
electron micrographs (Figure 8-8, B).
• Division.  the basophilia of polychromatic erythrocytes,
Lacking a nucleus, the polychromatic erythrocyte
cannot divide.
• Location.  A second functional change in polychromatic
The polychromatic erythrocyte resides in the bone
marrow for 1 day or longer and then moves into the
peripheral blood for about 1 day before reaching
maturity. During the first several days after exiting the
marrow, the polychromatic erythrocyte is retained in
the spleen for pitting of inclusions and membrane
polishing by splenic macrophages, which results in the
biconcave discoid mature RBC.7
• Cellular activity. 
The polychromatic erythrocyte completes production
of hemoglobin from residual messenger RNA using the
remaining ribosomes. The cytoplasmic protein
production machinery is simultaneously being
dismantled. Endoribonuclease, in particular, digests
the ribosomes. The acidic components that attract the
basophilic stain decline during this stage to the point
that the polychromatophilia is not readily evident in
the polychromatic erythrocytes on a normal
peripheral blood film stained with Wright stain. A
small amount of residual ribosomal RNA is present,
however, and can be visualized with a vital stain such
as new methylene blue, so called because the cells are
stained while alive in suspension (i.e., vital), before the
mesh of small blue strands, a reticulum, or, when
more fully digested, merely blue dots (Box 8-2Figure
8-9). When so stained, the polychromatic erythrocyte
is called a reticulocyte. However, the name
reticulocyte is often used to refer to the stage
when stained with Wright stain and without
demonstrating the reticulum.
• Cellular Basophilia: Diffuse and Punctate
The reticulum of a polychromatic erythrocyte
(reticulocyte) is not seen using Wright stain. The
residual RNA imparts the bluish tinge to the cytoplasm
seen in Figure 8-8, A. Based on the Wright-stained
appearance, the reticulocyte is called a polychromatic
an erythroblast but has a bluish tinge. When
polychromatic erythrocytes are prominent on a
peripheral blood film, the examiner uses the comment
polychromasia or polychromatophilia. Wright-stained
polychromatic erythrocytes are also called diffusely
basophilic erythrocytes for their regular bluish tinge.
This term distinguishes polychromatic erythrocytes
from red blood cells with punctate basophilia, in which
the blue appears in distinct dots throughout the
cytoplasm. More commonly known as basophilic
stippling (Table 19-3and Figure 19-1), punctate
basophilia is associated with some anemias. Similar to
punctate basophilia is due to residual ribosomal RNA,
but the RNA is degenerate and stains deeply with
Wright stain.
erythrocytes is the reduced production of receptors
for the adhesive molecules that hold developing RBCs
in the marrow (see details later). As these receptors
decline, cells are freed to leave the marrow.
• Length of time in this stage. 
The cell typically remains a polychromatic erythrocyte
for about 3 days,3 with the first 2 days spent in the
marrow and the third spent in the peripheral blood,
although possibly sequestered in the spleen.
ERYTHROCYTE

• Nucleus. 
No nucleus is present in mature RBCs.

• Cytoplasm. 
The mature circulating erythrocyte is a biconcave disc
measuring 7 to 8 μm in diameter, with a thickness of
about 1.5 to 2.5 μm. On a stained blood film, it
appears as a salmon pink-staining cell with a central
pale areathat corresponds to the concavity. The
central pallor is about one third the diameter of the
cell.

• Division. 
The erythrocyte cannot divide.

• Location and length of time in this stage. 

Mature RBCs remain active in the circulation for
approximately 120 days.11 Aging leads to their removal
by the spleen as described subsequently.

• Cellular activity. 
The mature erythrocyte delivers oxygen to tissues,
releases it, and returns to the lung to be
reoxygenated. The dynamics of this process are
discussed in detail in Chapter 10. The interior of the
erythrocyte contains mostly hemoglobin, the oxygen-
carrying component. It has a surface-to-volume ratio
and shape that enable optimal gas exchange to occur.
If the cell were to be spherical, it would have
hemoglobin at the center of the cell that would be
relatively distant from the membrane and would not
be readily oxygenated and deoxygenated. With the
biconcave shape, even hemoglobin molecules that are
toward the center of the cell are not distant from the
membrane and are able to exchange oxygen.
The cell’s main function of oxygen delivery throughout
the body requires a membrane that is flexible and
deformable—that is, able to flex but return to its
original shape. The interaction of various membrane
components described in Chapter 9 creates these
properties. RBCs must squeeze through small spaces
such as the basement membrane of the bone marrow
venous sinus. Similarly, when a cell enters the red pulp
of the spleen, it must squeeze between epithelial cells
to move into the venous outflow. Deformability is
crucial for RBCs to enter and subsequently remain in
the circulation.