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1) The cell size diminishes.
2) Ripening of the cytoplasm. the staining reaction of
the cytoplasm changes, the RNA starts diminishing)
These two things take place simultaneously.
a) So, colour changes to pink from blue due to
reduction of RNA matter such that in
Romanowsky stains,
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there is a change in colour from deep blue to pink
due to progressive formation of acidophilic staining
haemoglobin and the simultaneous reduction of
RNA which is responsible for basophilic of
cytoplasm.
b) Haemoglobin formation.
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3) Ripening (maturing) of the nucleus having the large
reddish, purple, open-network of the nucleus of a
polychromatic normoblast converted to small deeply
staining blue black structureless nucleus of the
orthochromatic normoblast before it is eventually lost.
i.e. the nucleus of the pronormoblast is larger than that
of the orthochromatic normoblast.
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The former (ponormoblast) has structures and stains
reddish purple, while the later (orthochromatic) stains
blue-black and is structureless.
Mitotic division of the developing cells in this series
occurs up to the stage of the polychromatic
normoblast and therefore the orthochromatic
normoblast is not considered capable of mitotic
division.
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Characteristics of developing normoblast
in romanowsky stained film.
1. Pronormoblast
Pronormoblast is a large oval cell. It comes about as
a result of activation by erythropoeitin (hormone)
which is produced in the kidney.
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Cont:
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2. Basophillic normoblast : (Early)
The size varies from 10-16 micrometer.
The nucleus is relatively large but smaller than
pronormoblast stage.
The cytoplasm is smaller to that of pronormoblastic
but slightly stage (plentiful).
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The chromatic strands are more thicker and more
deeply staining living a coarser appearance due to
RNA.
At this stage the nucleus have disappeared.
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3. Polychromatic normoblast (intermediate)
They vary from 8/14 micrometer in diameter.
The nucleus is smaller and occupies a smaller part of
the cell.
It is reddish purple in colour.
The cytoplasm is large and begins to acquire
haemoglobin, thus taking acidophillic – pink or pale
11 pink colour.
4. Orthochromatic normoblast (late)
The size ranges from 8-10 micrometer in diameter.
The nucleus is small with condensed homogenous
structereless chromatin.
It stains blue-black colour. The nucleus is eccentric
and sometimes lobulated.
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The cytoplasm is typically acidophillic (pink) because
haemoglobinization has started taking place which
gives it acidophillic staining.
The nucleus is lost at this stage.
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5. Reticulocytes
It is a flat disc shaped and non-nucleated cell larger
than a mature red blood cell.
When stained with romanowsky stains shows the
diffuse oale basophillia.
The cell is called polymetachromatophillic cell, while
with supra vital stains, the basophillic material
appears in the form of reticulum and chromatic
14 strands, which stains dark red.
Cont:
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As the cell matures it becomes smaller in size and the
nucleus diminishes and eventually lost i.e. the
chromatin becomes less dense and the nucleus
diminishes and eventually lost.
Nucleoli disappear before the nucleus is lost.
At this stage the cytoplasm losses the protein
synthesis property and acquires haemoglobin.
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6. Erythrocyte
The nucleus is not present.
The biconcave orange cytoplasm has the paler center
occupying a third of the cell area.
The size is about 6.7-7.7 micrometer in diameter and
2.5 micrometer thick.
The outline is regular.
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Characteristics of erythrocytes
When unstained, they appear as non-nucleated pale
greenish yellow biconcave discs.
When stained by the romanowsky stain, they have
affinity for eosin and therefore they stain pinkish
under a microscopic exam.
Their normal range,
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Women – 3.9-5.9m/mm3
Men - 4.5-6.5 m/mm3
Children – 4-6 m/mm3
Their life span is 110-120 days after which they are
removed by the R.E.S where they are broken down
to some of their constituents, some of which are
reutilized e.g. Fe and protein for haemopoiesis.
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When placed in hypotonic saline, they swell and loose
the biconcavity and assume a different shape
(spherical).
They contain the red pigment called haemoglobin.
They have no granules. They are living cells. They
contain enzymes of both anaerobic glycolytic pathway
of (pentose, phosphate pathway).
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60% of RBC is H2O and 40% is solid dry substance of
which 90% is haemoglobin and 10% is stroma.
RBC are flexible thus they can readily be destroyed or
distorted in shape e.g. when passing through small
capillaries but quickly resume their normal shape after
passing.
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Cont:
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Cont:
Types of erythropoiesis
1. Microcytic erythropoiesis
This is where cells are produced being smaller in
size. It happens because of disease conditions e.g.
iron deficiency anaemia, thalassaemia etc.
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2. Macrocytic erythropoiesis
Cells are produced being larger in size
due to abnormal conditions e.g. vitamin B12 / Folic
acid deficiency. Also in haemolytic anaemia.
3. Normocytic / normoblastic erythropoiesis.
It gives rise to normal red blood cells.
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Requirement of erythropoiesis
This can be divided into 2 categories:
a) nutritional requirement.
b) non nutritional requirement.
Nutritional requirements deal with diet e.g. iron,
vitamin, folates, trace metal, amino acids.
Non nutritional requirements include hormones e.g.
26 adrenaline, erythropoietin steroids.