Red Blood Cells

Question 1

What is haemopoiesis?

What are erythrocytes, leukocytes and platelets derived from? Where are they produced?

Answer 1

Haemopoiesis = The formation and development/differentiation of blood cells (Greek Haem - blood poeisis - making)

RBC (erythrocytes), WBC (leukocytes) and platelets are derived from pluripotent/multipotent haematopoietic stem cells (HSCs) and produced throughout life in the bone marrow.

Question 2

What is the distribution of HSCs and progenitor cells in the bone marrow?

What can HSCs differentiate into?

Answer 2

HSCs and progenitor cells are distributed in an ordered manner within the bone marrow amongst mesenchymal cells, endothelial cells and the vascular urge with with the HSCs interact.

HSCs give rise to lymphoid stem cells and myeloid stem cells, from which RBCs, granulocytes, monocytes and platelets are derived.

Question 3

What is haemopoiesis regulated by?

What happens if the regulation of haemopoiesis gets disrupted?

What do HSCs have the ability to do?

Answer 3

Haemopoiesis is regulated by various different genes and transcription factors, growth factors, eg: erythropoietin and the micro environment. Disruption of this regulation can disturb the balance between proliferation and differentiation, which may lead to leukaemia or bone marrow failure.

HSC have the ability to: self renew (some daughter cells remain as HSCs, so that the pool of HSCs are not depleted).

Question 4

In the fetus p, where are HSCs derived from?

Describe the journey of HSC differentiation and the sites of HSC production in the body from the first three weeks of a foetus’ life to birth

Mention: RBCs, platelet precursors and macrophages

Answer 4

HSCs are derived from the mesodern (in the fetus), primitive red blood cells, platelet precursors and macrophages are initially formed in the vasculature of the embryonic yolk sac in the first 3 weeks before the liver takes over between 6 and 8 weeks of gestation as the main site of haemapoiesis

The liver continues to be the principle source of blood in the foetus until just before birth. The bone marrow however starts developing haemopoietic activity from as soon as 10 weeks gestation. After birth, the bone marrow is the sole site of haemopoisis in healthy individuals.

Question 5

Throughout a persons lifetime, in which bones does haemopoiesis take place in?

Answer 5

As children haemopoisis takes place in nearly all bones but by adulthood it is restricted to the bone marrow of the pelvis, the vertebrae and the sternum, along with the proximal ends of the long bones of the thigh and arm, femur and humerus.

Question 6

What situation would result in an increased haemopoietic drive?

What happens when there is an increased haemopoietic drive?

Answer 6

Meyler proliferative disorder, polyscithemia (diseases that cause an increased haemopoietic drive), or where there increased red blood cell destruction - haemolosis

In these situations, where there’s an increased haemopoietic drive, haemopoietic tissue may expand into other marrow cavities. Expanded haemopoiesis may lead to development of haemopoietic foci in the adult liver and spleen = extra medullary haemopoeisis (haemopoiesis occurring outside of the bone marrow

Question 7

What are haemopoetic growth factors?

What do they do?

What are the growth factors for:
Erythropoiesis
Production of granulocytes
Megakaryocytopoiesis and platelet production

Where are the growth factors produced?

Answer 7

The growth factors Are glycoprotein hormones that bind to cell surface receptors
-These growth factors regulate proliferation and differentiation of HSCs

-erythropoiesis (RBC production) - under the influence of erythropoietin

Bone marrow production of Granulocytes(neutrophils, eosinophils and basophils) and monocytes: G-CSF, G-M CSF, cytokines e.g interleukins, granular site and granular site macrophage collagen colony stimulating factors

Megakaryocytopoiesis and platelet production: thrombopoietin

Production of haemopoeitic growth factors is by the cells of the bone marrow (except for eryhthropoeitin)

Question 8

Lymphoid differentiation pathways

Pluripotent haematopoietic stem cells

Answer 8

Pathway 1:

Pluripotent HSC ———> common lymphoid progenitor ——-> B cell progenitors in bone marrow ——-> mature B cells (antibody producing - part of humoral immunity response)

Pathway 2:

Pluripotent HSC———> common lymphoid progenitor ——-> T cell progenitors (thymus -> bone marrow) ——-> Mature T cells (cytokine producing- cellular immunity)

Pathway 3:

Pluripotent HSC ———> myeloid cells

Question 9

What happens in the late stage of myeloid progenitor cells?

What characterises young/immature RBCs?

What does the presence of nucleated RBCs in the blood indicate?

Answer 9

When the cells reach the late erythroblast stage the cell extrudes its nucleus (immature RBC)

polychromatic erythrocyte - characterised by its larger size and blue tinge. (Polychromasia - many colours, in this case it’s referring to the blue tinge on the erythrocyte due to the higher RNA content)

If nucleated red blood cells are present in the blood it means that there is a very high demand for the bone marrow to produce red blood cells, as immature red blood cells are being released prematurely into the circulation

Question 10

What 3 things are required for erythropoiesis?

What are the effects of being deficient in these substances?

Answer 10

Iron

• vitamin B12
• Folate

Low iron/B12/folate can lead to anaemia

microcytic anaemia caused by - iron deficiency - red blood cells are smaller and have areas of central pallor

Macrocytic anaemia caused by - B12/folate deficiency - larger red blood cells, these cells can grow but are unable to divide, the same applies to neutrophils on the granular sites, there are more lives (?) than expected = Megan aplastic anaemia.

Question 11

Erythropoietin

What is it?

Where Is it synthesised and in response to what?

What does the oxygen level of erythropoietin indicate?

How does erythropoietin increase the oxygen carrying capacity of the blood?

Answer 11

Eryropoietin is a glycoprotein synthesised in the cortical interstitial cells of the kidney in response to hypoxia

Their oxygen level may indicate a diminished number of red blood cells.

Erythropoeitin interacts with the erythropoietin receptor on red blood cell progenitor membranes and stimulates the bone marrow to produce more red blood cells. Resulting rise in erythrocytes increases the oxygen carrying capacity of the blood.

Question 12

Major functions of iron

Answer 12

Oxygen transport in Haemoglobin

It is a component of mitochondrial proteins - cytochromes a, b and c: for the production of ATP and Cytochrome and P450 for hydroxylation reactions (eg drug metabolism)

essential for synthesis of oxygen transport proteins, Haemoglobin, myoglobin

vital key factor for protein and enzymes involved in energy, metabolism, respiration, DNA synthesis, cell cycle arrest and apoptosis.
-essential for healthy skin, mucous membranes, hair and nails

Question 13

Where is iron absorbed in the body?

What are the two forms of dietary haem and what foods can you source them from?

Which is the better absorbed form and why?

Answer 13

Two forms of dietary iron are:

• haem - i.e animal derived(red meat poultry and fish), in ferrous (Fe2+) form, this is the best absorbed form
• non haem - mainly in ferric form (Fe3+) in food(plant based such as grains beans veg and seeds, + animal meat, dairy and eggs) and requires action of reducing subs (eg absorbing acid, vitamin C) for absorption

sources of non haem iron (e.g soya beans) often contain phytates which bind to iron and reduce its absorption.

Question 14

Iron homeostasis:

What are the effects of excess iron in the body, how does it cause these effects? How is iron absorption regulated?

What is iron bound to in the plasma?
How is iron transported from the liver to the bone marrow?

How is iron stored in the liver?

Answer 14

Excess iron is potentially toxic to organs like the heart and liver
-no physiological method by which iron is excreted…as iron can form free radicals that damage body tissues, so it’s important that iron overload is avoided by the tight regulation of iron absorption in the gut: only 1-2 mg per day is absorbed from diet

Iron in the plasma is bound to the transport protein transferrin, it delivers iron to the bone marrow for erythropoiesis and for its use in enzymes and muscles.

Iron is stored in the liver as the protein ferritin

Question 15

How is iron excreted from the body?

what is hepcidin, what does it do?

Answer 15

Most iron is recycled there’s a general tendency to conserve iron and although iron isn’t actively excreted from the body a small amount is lost through the shedding of skin

hepcidin - master regulating hormone of iron absorption and utilisation, acute phase protein, activated in chronic diseases esp in inflammatory states, this causes anaemia as there’s a reduction in iron supply and absorption and availability - anaemia of chronic disease.

When iron stores/ferritin are full there is upregulation of hepcidin expression and iron absorption is limited (related of iron is also blocked), whereas a requirement of increased erythropoiesis leads to a reduction in hepcidin and more iron absorption

Question 16

How does hepcidin regulate iron absorption?

What is ferroportin? What does it do?

What happens to iron in the duodenum?

Answer 16

Hepcidin synthesis is suppressed by erythropoeitic activity, this ensures iron supply by increasing ferroportin in the duodenum enterocyte, which increases iron absorption.

When storage is high, hepcidin synthesis is increased which binds and degrades ferroportin. This prevents the efflux of iron from the enterocyte, so its lost when the cell is shed into the gut lumen.

Iron is usually reduced to its ferris form in the duodenum, or taken taken up by the duodenal enterocyte, iron taken into the cell is stored either directly as ferritin (may be lost when enterocyte dies and shed into the gut lumen) or it is oxidised to the ferric form and transported to the plasma via ferroportin

Question 17

Why does vitamin B12 and folate deficiency inhibit DNA synthesis?

Which cells do vitamin B12 and folate deficiency affect?

Answer 17

Vitamin B12 (cobalamin) and folate are needed for dTTP synthesis, necessary for the synthesis of thymidine. Therefore definitely of vitamin B12/folate inhibits DNA synthesis.

Vitamin B12 and folate deficiency affects all rapidly dividing cells:
Bone marrow: cells can grow but are unable to divide normally
Epithelial surfaces of mouth and gut
Gonads

Question 18

Vitamin B12 is found in?

Folic acid is found in?

Answer 18

Vitamin B12 found (only in foods of animal origin except in fortified cereals): 
Meat 
Liver and kidney 
Fish 
Oysters and clams 
Eggs 
Milk and cheese 
Fortified cereals

Folic acid: 
Green leafy veg 
Cauliflower 
Brussels sprouts 
Liver and kidney 
Whole grain cereals 
Yeast 
Fruit 
^Intake of this may be in inadequate in frail elderly people

Question 19

How is vitamin B12 absorbed?

How could vitamin B12 deficiency arise?

How is folic acid absorbed?

When do folic acid requirements increase?

Answer 19

stomach B12 combines with insptrinsic factor (IF) made in the gastric parietal cells
Small intestine B12-IF binds to receptors in the ileum
Vitamin B12 deficiency may arise from:
-inadequate intake eg vegan use
-lack of acid in stomach (achlorhydria) eg if part of stomach removed, partial gastractomy surgery.
-inadequate secretion of IF: pernicious anaemia
(Autoimmune disorder)
-malabsorption eg coeliac disease

Folic acid is Absorbed in small intestine (mainly in duodenum)
Requirements increase:
Pregnancy
Folate supplementation required here
Increased red cell production e.g sickle cell anaemia

Question 20

Red blood cell destruction

Answer 20

Erythrocyte circulates for ~ 120 days
Then it’s destroyed by phagocytise cells of the spleen (macrophages)
Iron from haem returns to the bone marrow (via Fe- transferrin in plasma) where it’s recycled
Metabolism of haem produces bilirubin, Bilirubin (yellow compound) is excreted in bile

Question 21

What does erythrocyte function depend on?

What does a defect in one of these lead to?

Answer 21

Erythrocyte function depends on:
Integrity of the membrane
Hb structure and function
Cellular metabolism

A defect in any of these lead to shortened erythrocyte survival (haemolysis).

Question 22

Red cell membrane

Answer 22

Erythrocytes are biconcave in shape which helps their manoeuvrability through small blood vessels to deliver oxygen

Membrane consists of a lipid bilayer that is supported by a protein cytoskeleton and contains transmembrane proteins. These maintain the integrity and shape + elasticity/deformability of the red blood cell.

A (band 3) and B(rhesus) = transmembrane proteins
C(junctional) and D(spectrin) = skeletal proteins

Question 23

What does disruption of the vertical linkages in the RBC membrane lead to?

What are spherocytes? What does it lead to?

Answer 23

Disruption of vertical linkages in membrane (usually ankyrin/spectrin)/85 cases hereditary spherocytosis (autosomal dominant) - Cause of haemolysis.

Spherocytes are (rb) cells that are approx spherical in shape, consequently have a round, regular outline and lack central pallor. They are a result of the loss of cell by membrane without the the loss of an equivalent amount of cytoplasm so the cell is forced to get the rounder. Red cells become less flexible and are removed prematurely by the spleen - haemolysis.

Question 24

What does disruption of horizontal linkages in membrane produce?

When may elliptocytes occur? What do they look like?

Answer 24

Disruption of horizontal linkages in membrane produces hereditary elliptocytosis (don’t have an area of central pallor)

Elliptocytes may occur in iron deficiency - these have very pale hypochromic appearance

Question 25

Haemoglobin

Answer 25

main function of erythrocytes is to transport O2 from lungs to tissues
-O2 is transported by the haem moiety of HB from the lungs to the tissues
-each RBC contains around 300 million HB molecules
-HB A in adults is a tetramer - 4 subunits each composed of a globin chain 2 alpha 2 beta bound to a haem group
Each haem group consists of a ferrous iron ion (Fe 2+) held in a porphyrin ring
Each Fe2+ can bind to one o2 molecule (so each HB can carry 4 O2 molecules)

At birth HB we have feral HB which is 2 alpha and 2 gamma globin chains.

Binding of one oxygen molecule facilities binding of the second oxygen molecule - cooperativity, works due to induced conformational change in the structure of HB by the binding of an o2 molecule.

Question 26

Oxygen HB dissociation curve

Answer 26

Oxygen HB dissociation curve: 
Depends on: 
H+ conc/ pH
Structure of HB
Conc of 2,3-DPG

Right shift (easy oxygen delivery)
High CO2 low ph - Bohr effect
High 2,3-DPG
HbS

Left shift (gives up oxygen less readily)

• HbF
• CO

Question 27

Red cell metabolism

What are metabolic pathways highly adapted to?

Answer 27

Generation of ATP to meet energy requirements

Maintenance of: HB function, membrane integrity and deformability, RBC volume

Question 28

What is 2,3-DGP? What does it do?

What is G6PD? What does it do?

Where may oxidants be generated and how?

Answer 28

G6PD - glucose-6-phosphate dehydrogenase
Importsnt enzyme in the hexose monophosphate (HMP) shunt. HMP shunt is tightly coupled to glutathione metabolism, which protects red cell from oxidant damage
Oxidants may be generated in the blood stream, eg during infection, or may be exogenous e.g. drugs, broad beans
Deficiency of G6PD causes red cells to be vulnerable to oxidant damage.

Question 29

What are the effects of G6PD deficiency?

What are extrinsic oxidants?

Answer 29

Most common enzyme disorder - 400M people worldwide
X-linked inheritance - gene for G6PD is in the X chromosome so affected individuals are usually hemizygous males (but sometimes homozygous females)
G6PD deficiency usually causes intermittent, severe intravascular haemolysis as a result of infection or exposure to an exogenous oxidant
Extrinsic oxidants may be food stuff (eg broad beans) chemicals or drugs
Distribution parallels malaria: selective advantage, resistance to falciparum malaria
Episodes of intravascular haemolysis are associated w the appearance of considerable numbers of irregularly cobra red cells
Irregular in outline but are smaller than normal cells and have lost their central pallor
Usually result from oxidant damage to the cell membrane and to the HB
HB is dentures and forms round inclusions - Heinz bodies, which can be detected by a specific test

Question 30

Microcytic rbs?

Normocytic?

Macrocytic?

Answer 30

Microcytic - red cells that are smaller this normal or an anaemia w small red cells
Causes - iron deficiency, anaemia of chronic disease, defect in globin synthesis (thalasssaemia), defect in alpha chain synthesis/beta chain synthesis = alpha/beta thalassaemia

Normocytic - red cells that are normal size or anaemia w normal sized red cells

Macrocytic - red cells that are larger than normal/ anaemia w large red cells, macrocytic can be round, oval or polychromatic (young immature RBC)
Causes - vitamin B12 deficiency or folic acid (megaloblastic anaemia), liver disease and ethanol toxicity, haemolysis (results in macrocytic blood picture with the overall vol being larger) (polychromasia), pregnancy

Question 31

Colour of
:

Normal RBCs

Hypochromia

Polychromasia

What are reticulocytes? How can you detect reticulocytes? Why may reticulocytosis occur?

Answer 31

Normal RBCs have a third of the diameter that is pale, this is a result of the disc shape of the red cell; centre has less HB and is therefore paler

Hypochromia - larger area of central pallor than normal, results from a lower HB content and conc and a flatter cell.
Red cells that show hypochromia = hypochromic
Hypochromia and microcytosis often go together (iron deficiency and thalassaemia)

Polychromasia - (many colours), describes blue tinge to the cytoplasm of a RBC
Indicated red cell is young
Polychromatic cells are larger than normal red cells - i,e cause of macrocytosis
Young red cells: reticulocytes snd reticulocytosis

• new methylene blue (stain) can be used to detect young cells for reticulocytes, this stains for higher RNA content
• reticulocytosis refers to the presence of increased numbers of reticulocytes
• reticulocytosis may occur as a response to (increased output of RBCs from bone marrow) bleeding or haemolysis

Question 32

Diffs in RBC shape

Poikilocytosis

Target cells

Sickle cells

Fragments

Answer 32

Poikilocytosis - red cells that show more variation in shape than is normal, come in a variety of shapes -

target cells - red cells where there’s clan accumulation of HB in the centre of the area of central pallor, at occur in a number of diff conditions - obstructive jaundice, liver disease, haemoglobinopathies, hyposplemism (spleen doesn’t function properly or has been removed),

sickle cells - sickle/crescent shape, result from the polymerisation of HB S, which in the deoxygenated form is less soluble than HB A. HB S occurs when one or two copies of an abnormal beta globin gene are inherited. Sickle cell anaemia was first condition to be described as being caused by a specific protein defect (1948). Caused by mutation in The beta globin gene: charged glutamic acid residue in position 6 is replaced by an uncharged valine molecule.

fragments - schistocytes are small pieces of red cells, may result from a shearing process caused by the platelet rich blood clots in the small blood vessels e.g disseminated intravascular coagulation