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HAEMOPOIESIS AND ITS REGULATION/ERYTHROID DEVELOPMENT AND MATURATION


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INTRODUCTION

  • The process of blood cell formation is called haemopoiesis
  • The process of formation of each cellular components of the blood are called;
    • Erythropoiesis – red cell formation
    • Leukopoiesis – white blood cell formation
    • Granulopoiesis – granulocyte formation
    • Megakaryopoiesis – platelet formation Need for Haemopoiesis
  • Haemopoiesis involves a very high level of cell turnover, which is demanded by the need to replace mature circulating blood cells at a rapid rate
  • This is necessitated by the limited lifespan of the mature cells
  • Granulocytes survive for only a few hours (6-8 hours) and erythrocytes for a few months (90-120 days)
  • Due to the short half-life of these cells, about 1013 new cells must be produced each day to maintain steady-state blood counts
  • Higher production may however be required based on body demand especially during blood loss or during acute infection
  • During blood loss, the cell production will favour erythropoiesis and during infection, production is directed towards white cells

SITES OF HAEMOPOIESIS

Fetal Sites

  • In the few weeks of gestation, the embryonic haemopoietic stem cells develop from mesenchymal cells in the yolk sac which serves as a transient site for haemopoiesis
  • However, definitive haemopoiesis derives from a population of stem cells which was first observed on the aorta-gonads-mesonephrons (AGM) region
  • These common precursors of endothelial and haemopoietic cells (haemangioblasts) are believed to seed the liver, spleen and bone marrow
  • Placenta also contribute to haemopoiesis
  • From 6 weeks until 6-7 months of fetal life, the liver and spleen are the major haemopoietic organs and continue to produce blood cells until about 2 weeks after birth
  • Megaloblasts are the first recognizable cells followed by normoblast
  • The spleen is also active during this period particularly in the production of lymphoid cells
  • Foetal thymus is a transient site for some lymphocytes

Sites of Haemopoiesis

 

Birth, Infancy and Adult Sites

  • From 20 week, bone marrow becomes increasingly important and by birth, the main haemopoietic organ
  • In infancy, all the bone marrow is haemopoietic
  • But during childhood, there is progressive fatty replacement of marrow throughout the

long bones” so that in adult life, active marrow are confined to;

  • The central skeleton (skull, vertebra, sternum, ribs, scapula, pelvic bones)
  • Proximal and distal ends of the femurs and humeri
  • In the first 2-3 years after birth, active marrow is found in all bones both long and flat bones

 

Bone Marrow

  • The bone marrow, which is the major site of haemopoiesis in adults, contains cells at different stages of maturation
  • There are primitive cells which serves as the precursors for the mature cell lineages  The recognizable cells in the marrow include;
    • Myeloblast
    • Myelocytes
    • Metamyelocytes
    • Erythroblast
    • Normoblast
    • Segmented granulocytes – Neutrophils, Eosinophil, Basophils
    • Erythrocytes
    • Monocytes
    • Megakaryocytes
    • T and B Lymphocytes
  • Earlier stages of development in the marrow are progressively less morphologically distinct in their lineage affiliation and fewer in number
  • The least frequent cells, which cannot be discriminated morphologically, are the committed progenitor cell populations and the stem cells

Normal Bone Marrow

 

Red Marrow

  • Red marrow forms all types of blood cells i.e. red blood cells, white blood cells, platelets and is also active in the destruction of red blood cells
  • Red marrow is therefore one of the largest and most active organ of the human body approaching the size of liver in overall mass although distributed in various parts of the body
  • About 2/3rd of its mass function in white cell production and 1/3rd in red cell production
  • This is due to the shorter life span and greater turnover of the white blood cells in comparison with red blood cells that have lifespan 90-120 days

Stem Cell

  • Haemopoiesis starts with a common pluripotential stem cell
  • The exact phenotype of the human stem cell is unknown, but on immunological testing, it is CD34+, CD38- and has the appearance of a small or medium sized lymphocyte
  • The stem cells are the most important cells in haemopoietic cell production
  • They are ultimately responsible for regenerating haemopoiesis following damage to the haemopoietic system by myelotoxic chemotherapy or after stem cell transplantation
  • This is accomplished by stem cell division, producing new stem cells to maintain the stem cell pool (stem cell renewal) and differentiating cells that are the progenitor cells of each of the blood cell lineage
  • Estimates of stem cell frequency in human bone marrow are about one stem cell per 20 million nucleated cells

Stem Cell Properties, Phenotype and Purification

  • Morphologically, haemopoietic stem cells are undifferentiated and resemble small lymphocytes
  • Normally, a large fraction is in quiescent or G0 phase of the cell cycle
  • This protects them from the action of cell cycle-dependent drugs such as 5’-flourouracil  And S-phase-specific agents such as cytosine arabinoside and Hydroxyurea
  • The quiescent state of stem cells is maintained by transforming growth factor β (TGF-β) at very low concentrations

Stages in Haemopoietic Cell Development

 

PHENOTYPIC MARKERS OF PRIMITIVE CELLS

STEM CELLS                                                      PROGENITOR CELLS

CD34 positive

CD34 positive

CD38 negative

 

CD33 negative

CD33 negative

Lin negative

Lin +

HLA-DR negative OR weakly positive

HLA-DR positive

 

Hierarchical Organization of Haemopoiesis

 

REGULATION OF CELL PRODUCTION

  • The maintenance of blood cell production between the normal narrow limit requires that the regulatory mechanisms operate in a controlled and coordinated way and are able to elicit a concerted response in the different areas of the active marrow
  • The varied functions of the blood cells also imply that cells in different tissues of the body can signal the need of different cells production e.g. anoxia condition leads to red cell production
  • The response to stimulation also implies that there must be inhibitory signals which play a role in avoiding overproduction of cells thereby preventing sluggishness of blood flow due to hyperviscosity syndrome
  • The concerted effort of cytokines, extracellular matrix and adhesion molecules have led to homing of cells into an environment where they can respond to a host of regulatory molecules

Stages in the Homing and Mobilization of Stem Cells

 

HAEMOPOIETIC GROWTH FACTORS

General Characteristics of Myeloid and Lymphoid Growth Factors

  • They are glycoproteins that act at very low concentration
  • They act hierarchically
  • They are usually produced by many cell types
  • They usually affect more than one lineage
  • They are usually active on stem/progenitor cells and on functional end cells
  • They usually show synergistic or additive interactions with other growth factors
  • They act on the neoplastic equivalent of a normal cell
  • They have multiple actions
    • Proliferation
    • Differentiation
    • Maturation
    • Prevention of apoptosis
  • They mediate their actions through specific receptors
  • They can act by cell-to-cell contact or circulate in the plasma

Examples of Myeloid and Lymphoid GFs

  • Act on pluripotential stem cells
    • Stem cell factor
    • Flt ligand
  • Act on multipotential progenitor cells
    • IL-3
    • GM-CSF
    • IL-6
    • G-CSF
    • Thrombopoietin
  • Act on committed progenitor cells
    • G-CSF
    • M-CSF
    • IL-5 (eosinophil CSF)
    • Erythropoietin
    • Thrombopoietin
  • Inhibitors of Cell Proliferation
    • Macrophage inflammatory protein (MIP-1α)
    • Transforming growth factor β (TGF-β)
    • Tumor necrosis factor (TNF)

STROMAL CELL-MEDIATED HAEMOPOIESIS

  • In the marrow, haemopoiesis occurs in association with stromal cells
  • The stromal cells produce a variety of cytokines either constitutively or following stimulation
  • IL-1, IL-11 and SCF[1] can be detected in marrow stromal cells
  • In addition, M-CSF, GM-CSF, G-CSF and TGF-β can also be produced by stromal cells either constitutively or following induction by a variety of stimuli
  • The association of some of these factors with the different cell types constitute the bone marrow microenvironment
  • The marrow stromal cells are heterogeneous, comprising;
    • Fibroblasts
    • Reticular cells
    • Adipocytes; and
    • Endothelial cells
  • They are responsible for the production of most growth factors
  • Macrophages also have roles in cytokines production and are also a functional part of the environment

ERYTHROID DEVELOPMENT AND MATURATION

  • Erythropoiesis – this is a process of red cell production
  • This process passes through many stages of mitotic cell division
  • A single stem cell can produce between 8-16 red cells
  • Erythropoiesis normally maintains the steady state of an individual’s red cell mess, producing 1011-1012 new red cells every day to replace those that are lost through senescence or premature destruction
  • Primitive erythropoiesis is first evident at around 3 weeks of gestation in the yolk sac
  • So many transcription factors are involved in the red cell production, such factors include;
    • GATA1
    • GATA2
    • FOG1
    • NFE2
    • TALI
    • RUNXI
  • All these factors work together to aid differentiation and maturation of red cell
  • During normal erythroid development, GATA2 probably initiates the erythroid programme and plays an important role in the expansion and maintenance of haemopoietic progenitors
  • Its replaced during terminal erythroid maturation by GATA-1, which is essential for the terminal differentiation and maturation of both megakaryocyte and erythroid cells

STAGES OF DEVELOPMENT

Haemopoietic stem cell

CFU-GEMM

Early BFU-E

Late BFU-E

CFU-E

Pronormoblast

Early normoblast

Intermediate normoblast

Late normoblast

Reticulocytes

Mature red blood cells

  • This involves;
    • A gradual appearance of haemoglobin in the cell
    • A gradual disappearance of RNA from the cell
    • Progressive degeneration of the cell’s nucleus which is eventually extruded from the cell
    • The gradual loss of cytoplasmic organelles e.g. mitochondrion
    • A gradual reduction in cell size
  • CFUGEMM, BFUE, and CFUE are called progenitor cells; while
  • Pronormoblasts, early, intermediate and late normoblasts and reticulocytes are called precursor cells

 

REQUIREMENTS FOR NORMAL ERYTHROPOIESIS

The marrow required many other requirements for effective erythropoiesis. These include;

  • Proteins – red cell protein and globin part of Hb
  • Vitamins – DNA, folate metabolism (Vit C), Vit E, Vit B6
  • Folic acid – DNA
  • Iron – required for haem part of Hb
  • Copper
  • Cobalt
  • Thyroid hormone (TSH)  Adrenocortical hormone
  • Human growth hormone
  • Androgen – stimulates erythropoietin leading red cell production
  • Oestrogen depresses erythropoiesis

HAEMOGLOBIN

  • Red cells contain approximately 640 million haemoglobin molecules
  • Adult Hb A consists of 4 polypeptide chains, α2β2, each with its own haem group  MW of HbA is 68 000
  • Normal adults also contain small quantity of 2 other haemoglobins HbF and HbA2
  • These Hbs contain α chain, but γ and δ chains instead of β
  • Major switch from fetal to adult Hb occurs 3-6 months after birth

HAEMOGLOBIN SYNTHESIS

  • This occurs largely in the mitochondria by a series of biochemical reactions commencing with the condensation of glycine and succinyl Coenzyme A under the action of a rate limiting enzyme – δ aminolaevulinic acid synthase
  • Pyridoxal phosphate is a co-enzyme for this reaction
  • Ultimately, protoporphyrin combines with iron in the ferrous state to form haem
  • Each molecule of which combines with a globin chain on the polyribosomes to form Haemoblobin

 

 

[1] SCF – Stem Cell Factor