Production of Specific Cell lineages

1. Erythrocyte Production (Erythropoiesis

Objectives

At the end of this lesson, you are expected to:

 Define erythropoiesis

 Discuss the process of erythropoiesis

 Discuss maturation stages and characteristics of each stages of

erythrocytes

 Explain how erythropiesis is regulated and list the effects of the

hormone erythropoietin on erythropoiesis

 Discuss ineffective erythropoiesis

 Explain megaloblastic erythropoiesis

 Erythrocyte Production (Erythropoiesis)
 Erythropoiesis
• Begins with the development of primitive erythrocytes
• Myeloid lineage cells

 Basic substances
• Amino acids (proteins), iron, Vit B12, Vit B6, folic acid and the trace
minerals cobalt and nickel

 Regulated by erythropoietin

 Androgen and thyroid hormones can also stimulate erythropoiesis

 Erythroid Progenitor cells (BFU-E & CFU-E)
 Derived from CFU-GEMM)
• Burst-Forming Unit-Erythroid (BFU-E)

 BFU-E produces a large multiclustered colony

Regulator: Burst
• Unipotent
Promoting Activity
• 16 or more clusters (BPA)
• Few receptors for erythropoietin (EPO)
• Takes one week to mature to the CFU-E

 CFU-E
• Has many receptor sites for EPO, highly sensitive
• Proliferate into pronormoblasts (Proerythroblast): Takes 1 week
Erythropoiesis

•Source: Rodak’s Hematology: clinical principles and application, 5th ed page:97

Erythropoiesis
 Terminologies of Erythrocyte precursors
Normoblastic Erythroblstic Rubriblastic
(US) (Europe) (Similar to WBC)
Pronormoblast Proerythroblast Rubriblast
Basophilic normoblast Basophilic erythroblast Prorubricyte
Polychromatic Polychromic Rubricyte
(polychromatophilic) (polychromatophilic)
Normoblsat erythroblast
Orthochromic normoblast Orthochromic erythroblast Metarubricyte

Polychromatic Polychromatic Polychromatic

(polychromatophilic) (polychromatophilic) (polychromatophilic)
erythrocyte erythrocyte Erythrocyte
Erythrocytes Erythrocytes Erythrocytes
 Proerythroblast/ Pronormoblast/ Rubriblast
 Earliest morphologically recognizable cell
• 1% of bone marrow cells
 Homogeneous nucleus with nucleoli and fine
chromatin pattern
 12 - 20m in diameter

 Nucleus stains reddish-blue (dark appearing) and

has reticular appearance
 Nucleus large round to oval, and not condensed
 1-2 nucleoli
• Mitosis present

• Cytoplasm: Scanty , only a rim around the

nucleus; deep basophilic (blue)
• No hemoglobin (globin production begins)
Proerythroblast
 Basophilic erythroblast/ Early normoblast / prorubricyte
Slightly smaller than Proerythroblast
 Nuclear material has begun to coarsen
• Nuclear chromatin shows sharp
contrast between light & dark areas
(Clumped)
• Nucleoli present in early stage but
disappeared latter
 Cytoplasm: wider ring of deep blue than
previous stage (Basophilic cytoplasm)
 Detectable hemoglobin synthesis occurs
but pigmentation is masked
 N:C = 6:1
 Active mitosis
 Slight reduction in size 10 -15µm
 1-4% of BM cells
Basophilic erythroblast (Right)
 Polychromatophilic erythroblast/ Intermediate Normoblast/
Rubricyte
 10-12m in diameter
 Nuclear chromatin is thickened and
irregularly condensed (Clumped/coarse)
• Pyknotic nucleus
• Nucleoli no longer visible
• Nucleus is eccentric
 Decreased N:C ratio from 4:1 to 1:1

 Cytoplasm: Pink coloration mixed with

basophilia (lilac/polychromatic)
• Light gray blue appearance
• Due to increased Hgb appearance

 Reduced mitoses (last stage for division)

 10-20% BM cells
Polychromatophilic erythroblast (Right)
Orthochromatic erythroblast/ Late normoblast/ Metarubricyte
 Size: 8-10m in diameter.
 Incapable of further DNA synthesis (No mitosis)

 Nucleus: N:C: 1:2 or 1:3

• Nucleus is completely condensed (pyknotic)
• Chromatin structureless
• Central or eccentric
• At later stage nucleus will be extruded

 Acidophilic erythroblast (reddish -pink-orange)

• Presence of large quantities of Hgb

 Not capable of division due to the condensation of

the chromatin.
 Howell Jolly bodies
Orthochromatic erythroblast (Right)
 Polychromatic Erythrocyte/Reticulocyte
 Size: 8-10m in diameter
 Young erythrocytes with a network of clumped ribosomes and Rough
endoplasmic reticulum.
 Has no organelles

 Is not concave but has irregular shape

 Retained in spleen for pitting and polishing by splenic macrophages = biconcave

 Part of maturation phase occurs in BM and the later part takes in circulating blood

 Cytoplasm is diffusely basophilic (Blue)

• Polychromatophilic (due to RNA)
• Contains 2/3 of total hemoglobin
 Have Transferrin receptors
 30% of total hemoglobin produced
 Polychromatic Erythrocyte - Reticulocyte
 Supra Vital staining required to make
visible.

 When stained with New Methylene Blue are

called Reticulocytes
• Ribosomes containing residual RNA

 Makes up 1-2% of all erythrocytes in PB

 Lose RNA and matures into RBC in 1-2 days

 Provide an index of rate of RBC formation

Polychromatic Erythrocyte (Right)
 Mature erythrocyte
 Size: 6-8m in diameter
 Reddish, circular, biconcave cells filled with hemoglobin
 One third of the cell area (central pallor) due to biconcave shape
 No visible internal structure.
 Does not synthesize protein
 Transports oxygen to tissue
ERYTHROPOIESIS Summary

12-20µm- basophilic cytoplasm,

nucleus with nucleoli (0-2), NC: 8:1

10-15µm-mitosis, basophilic
cytoplasm, nucleoli disappears.
NC: 6:1

10-12µm-’POLYCHROMASIA’ NC: 4:1

Hgb appears, nucleus condenses.

8-10µm- PYKNOTIC Nucleus.

Extrusion, Hgb is maximum. NC: 1:2

8 -10µm- Reticulum of basophilic

material in the cytoplasm.

6 -8µm- Mature red cell with Hb.

 Duration of erythropoiesis
 HSC to RBC
• 21 days

 Differentiation phase: from Pronormoblast to

reticulocyte phase
• 5 days

 Maturation phase: from reticulocyte to RBC-

• 2 days
Erythropoiesis Summary
 Erythropoiesis Summary…..
Polychromatophilic (or
Proerythroblast Reticulocyte intermediate)
or (brilliant cresyl Erythroblast or
pronormoblast blue dye) Reticulocyte
Normoblast

A B C D

Orthochromatic
Basophilic erythroblast Orthochromatic (Acidophilic) erythroblast
Dividing Polychromatophilic Or
or erythroblast
Erythroblast or Late Erythroblast
Early Normoblast Extruding Nucleus
Normoblast

E F G H
Name the Erythrocyte precursors indicated by the
arrow
Polychromatophilic Erythroblast
Basophilic erythroblast

Orthochromatophic Erythroblast

Proerythroblast
Regulation of Erythropoiesis
 Factors regulate Erythropoiesis
 General factors  For Hgb Formation
-Erythropoietin - First class protein & amino
-Thyroxine acids
-Hemopoietic growth factors - Iron, Copper
-Vitamins - Cobalt & Nickel

 Maturation factors  Vitamins

- Cobolamin (Vit.B12) -Vit-C
-Intrinsic factor -Riboflavin (vit B2)
-Folic acid (Vit B9) -Nicotinic acid (Vit B3)
-Pyridoxin (Vit B6)
 Regulation of Erythropoiesis
 Erythropoietic activity is regulated by the hormone erythropoietin

 Erythropoietin:
• a heavily glycosylated hormone (40% carbohydrate) with a
polypeptide of 165 amino acids
• There are no pre-formed stores of erythropoietin
• The stimulus to the production of the hormone is the oxygen
tension in the tissues (including the kidneys)
• Its action is in the bone marrow
• 85 – 90% produced by kidney
• 10 – 15% produced in liver
Role of Erythropoietin
Erythropoietin production
increases when there is tissue
hypoxia due to:
• Low blood hemoglobin
levels (e.g., anemia)

• Impaired oxygen release

from hemoglobin (e.g.,
hemoglobinopathies)

• Low atmospheric oxygen

(e.g., high altitude)

• Pulmonary disease and

Circulatory defects
 Role of Erythropoietin ……
 Erythropoietin accelerates nearly every stage of red cell production:

 It increases committed stem cells division and differentiation

 It increases the rate of cell division

 It speeds up the incorporation of iron into the developing red cells

 It shortens the time cell maturation, and hastens the entry of

reticulocytes into the peripheral circulation

In what conditions erythropoietin production reduced?

Hormonal factors affecting Erythropoiesis
1. Androgens: increase erythropoiesis by stimulating the production of
erythropoietin from kidney but inhibited by Estrogen

2. Thyroid hormones: Stimulate the metabolism of all body cells

including the bone marrow cells, thus, increasing erythropoiesis.
• Hypothyroidism is associated with anemia while hyperthyroidism
is associated with polycythemia.

3. Glucocorticoids: Stimulate the general metabolism and also stimulate

the bone marrow to produce more RBCs.
• Progenitor cells have both erythropoietin as well as glucocorticoid receptors
• In Addison’s disease (hypofunction of adrenal cortex) anemia
• Cushing’s disease (hyperfunction of adrenal cortex) polycythemia
 Hormonal factors ……
4. Pituitary gland: Affects erythropoiesis both directly and indirectly
through the action of several hormones such as Erythropoietin

5. Haematopoietic growth factors: Are secreted by lymphocytes,

monocytes & macrophages to regulate the proliferation and
differentiation of progenitor stem cells to produce blood cells.
• Interleukin-3
• Interleukin-5
• Interleukin-6
• Interleukin-11
 Other factors….
State of liver & bone marrow:
1. Liver: Healthy liver is essential for normal erythropoiesis because
the liver is the main site for storage of vitamin B12 , folic acid, iron
& copper. In chronic liver disease anemia occurs.

2. Bone marrow: When bone marrow is destroyed by ionizing

irradiation or drugs, aplastic anemia occurs.
 Ineffective erythropoiesis/Intramedullary hemolysis
 Erythropoiesis is not entirely efficient since 10-15% of
eryhtropoiesis in a normal bone marrow is ineffective,

 Developing erythroblasts die within the marrow without producing

mature cells
• Ingested by macrophages

 This process is substantially increased in a number of anemias.

 Megaloblastic Erythropoiesis
 Megaloblasts are pathologic cells that are not present in the normal
adult bone marrow

 Deficiency in vitamin B12 or folic acid or both

• Defective DNA synthesis

 In megaloblastic erythropoiesis, the nucleus and cytoplasm do not

mature at the same rate

 Thus nuclear maturation lags behind cytoplasmic hemoglobinization

(RNA and protein synthesis)
• Megalocytes produced (9-12m in diameter).
• Megaloblasts in bone marrow
 Assignments to be discussed
1. Mature RBCs have no organelles. What is the functional advantage?
2. Why newborns has high number of erythrocytes than adults?
3. What are the mechanisms to increase erythrocytes in to the
circulation?
4. Discus how reticulocytes early released in to the circulation from
bone marrow?
5. Erythrocyte maturation starting from BFU-E takes about 18 – 21
days. If so, how the body compensates emergency need of red blood
cells during blood loos such as hemorrhage?
6. How microcytic and macrocytic red blood cells produced?
7. Is RBCs live or alive? Justify