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Blood Cells Simon Hunt Dunn School of Pathology http://users.path.ox.ac.uk/~svhunt/internal/bloodcells OR http://www.weblearn.ox.ac.uk/bodington/site/medsci/undergrad/med/teach/lectsupp/hunt/ Images from Wheater’s Functional Histology, 4 th edn

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blood cells

Blood Cells

Simon Hunt

Dunn School of Pathology

http://users.path.ox.ac.uk/~svhunt/internal/bloodcells

OR http://www.weblearn.ox.ac.uk/bodington/site/medsci/undergrad/med/teach/lectsupp/hunt/

Images from Wheater’s Functional Histology, 4th edn

Some electron micrographs from the collection of the late Drs Poole and French, Dunn School

purposes of this presentation
Purposes of This Presentation
  • How structure relates to function in erythrocytes (red blood cells)
    • To the level of syllabus section 5.4.1
    • As living and dying ultra-specialised (“differentiated”) cells
  • Ditto for leukocytes (white blood cells)
    • To the level of syllabus section 5.4.2
    • A functional “Who’s Who?”
  • How we find out about the physiological properties of blood cells
    • Illustrated by some disease conditions

Blood Cells

erythrocytes greek red hollows

7.2mm

Erythrocytes – Greek: “Red Hollows”
  • Pink stain – eosin, not haem
    • Haemoglobin is a basic protein
    • Binds acid dyes (e.g. eosin)
  • Scanning e-m
    • Always black & white

Blood Cells

blood cells computer enhanced
Blood cells, computer-enhanced

© Dennis Kunkel Microscopy, Inc http://www.denniskunkel.com

Blood Cells

erythrocytes can form rouleau pl rouleaux
Erythrocytes can form Rouleau (pl. Rouleaux)
  • Like a stack of coins
    • Only when blood flow is slow

http://www.finchcms.edu/cms/anatomy/histology/histology/blood/blood.html#blood

Blood Cells

rouleaux 2
Rouleaux 2
  • by fluorescence
    • Red shows DNA stain of leukocytes
    • Shows that rbc have no nuclei

http://www.cyto.purdue.edu/flowcyt/educate/photos/cells/images.htm

Blood Cells

erythrocyte shape

(a):discocyte, (b) stomatocyte and (c) echinocyte

from Harriet Gershon www.els.net

Erythrocyte shape
  • Anucleate (no nucleus)
    • also, no mitochondria
  • Shape depends on water content
    • Osmotic effects of solutes, especially ions
  • Shape maintained by cytoskeleton
    • Spectrin, ankyrin, & other membrane proteins

Blood Cells

advantages of being anucleate
Advantages of being anucleate
  • Better surface-volume ratio
    • About 25% greater than if an equivalent sphere
    • Improves gas exchange
  • Improves deformability
    • To fit through capillaries
      • diameter of true capillaries is only 5-10 mm, often less
  • Less work for heart as a pump
    • Pumps approximately 3 kg of erythrocytes per minute
    • 40% of total mass would be nucleus
    • Saves pumping 1 to 1.5 tons per day

Blood Cells

disadvantages of being anucleate
Disadvantages of being anucleate
  • No further protein synthesis or repair
    • Finite cell lifespan
      • 120 days on average
    • Requires vast new replacement cell production
      • Red blood cell production = “erythropoiesis”
  • Equally true for blood platelets
      • Platelet production = “thrombopoiesis”

Blood Cells

consequences of lacking mitochondria
Consequences of lacking mitochondria
  • Have to survive on anaerobic metabolism (glycolysis)
    • Energy needs not great, mainly ion pumps
    • Depends solely on blood glucose for energy supply
    • the glycolytic intermediate, 2,3-bisphosphoglycerate (2,3-BPG), is produced by an erythrocyte enzyme
      • BPG shifts dissociation curve to unload O2 from HbO2

Blood Cells

erythrocyte c ontents 1
ErythrocyteContents 1
  • Haemoglobin ~750 gm per adult body
    • Globin protein, alpha2beta2,~650 million molecules per cell
    • Haem prosthetic group
    • One Fe2+ per haem, ~2.2 gm per whole body
      • About 2/3 of all body iron
      • Must not oxidise to Fe3+ ( “methaemoglobin”)
        • cell needs reducing conditions
  • Functions in O2 and CO2 transport
    • Dr Dorrington’s lectures later this term

Blood Cells

erythrocyte c ontents 2
ErythrocyteContents 2
  • Glucose metabolising enzymes
    • Anaerobic glycolysis (Embden-Meyerhof Pathway)
    • Pentose Phosphate shunt
      • Uses G6PDH, Glucose-6-Phosphate Dehydrogenase, an X-linked enzyme
      • generates NADPH, slows build-up of oxidised proteins associated with erythrocyte ageing
      • thus maintains Glutathione, a Cysteine-containing tripeptide, in the reduced state

G6PDH deficiency is revealed by a serious haemolytic crisis when broad beans or the (now-obsolete) anti-malarial Pamaquine are ingested

Blood Cells

erythrocyte c ontents 3

[Na+] = 6 mM

[K+] = ~100 mM

ErythrocyteContents 3
  • Ions, especially K+
    • Maintained by membrane-associated ATP-dependent Na+-K+ ion exchanger

PLASMA concns:

[Na+] = 140 mM

[K+] = 3.5 - 5 mM

Blood Cells

membrane protein functions
Membrane protein functions
  • Cytoskeletal proteins maintain shape
    • spectrin, ankyrin, Band III
  • Channels, pores or pumps
    • Cations, anions, water, glucose
  • Glyoproteins & glycolipids: display extracellular carbohydrate
    • glycophorins maintain net negative charge
    • blood group substances
  • Regulatory proteins
    • complement-absorbing components
      • anti-inflammatory action

Blood Cells

osmotic effects
Osmotic effects

“Crenated”

From http://arbl.cvmbs.colostate.edu/hbooks/cmb/cells/pmemb/osmosis.html

Blood Cells

hypotonic haemolysis

http://www.physiology.rwth-aachen.de/user/martin/DuUndDeinBlut/Bl-Terminologie-d.htmlhttp://www.physiology.rwth-aachen.de/user/martin/DuUndDeinBlut/Bl-Terminologie-d.html

Hypotonic haemolysis
  • “Hypo-” means lower than normal
    • i.e. solute concentration outside is less than inside cell
      • allow for ions; count all osmotically active particles; use “Osmolarity”
    • water potential outside cell > intracellular
  • Cell membrane semi-permeable
    • Water permeates via “Aquaporin” proteins
    • >250 times cell volume crosses membrane per second

Blood Cells

osmotic fragility test

Normal

http://hsc.virginia.edu/medicine/clinical/pathology/educ/innes/text/rcd/membrane.html

Diseased - fragile

Osmotic fragility test

Blood Cells

erythrocytes can deform

DHAG e-m#328

Erythrocytes can deform
  • To squeeze through arterioles or capillaries
  • Tend to keep to central axis of vessel
    • Plasma-rich at circumference
  • Blood is “visco-elastic”
    • Not easy to find good artificial substitutes

NB rbc are electron-dense – why?

Blood Cells

rheology the flow properties of blood suspensions
Rheology - the flow properties of blood suspensions
  • The Fåhraeus-Lindqvist effect(small-diameter phenomenon)
    • rbc concentration is lower near the vessel wall and higher in the centre
      • hence vessel haematocrit decreases in the smaller branches of the vasculature i.e. more fluid and fewer cells in capillaries
      • hence get misleading blood cell concentrations if capillaries are sampled

Blood Cells

anomalous viscosity of blood
Anomalous viscosity of blood
  • Viscosity of blood increases with decreased velocity
    • Blood flow is low in small vessels (1 mm/sec) –
      • viscosity can increase 10 times just because of slow velocity
      • due to adherence of RBCs to each other (form rouleaux) and to vessel walls
      • shear forces no longer enough to deform RBC, so they appear more rigid
  • Effect is even more noticeable
      • if membrane more rigid, e.g. Spectrin defect, or becomes crenated
      • in aged erythrocytes
      • if there are inclusions inside cells, e.g. sickled cells
      • if rbs are enlarged osmotically
  • In what circumstances might this matter clinically?

Blood Cells

hereditary spherocytosis 1

Spectrin defect

Ankyrin defect

Band III defect

Spectrin deficiency

Decreased rbc deformability, osmotic fragility

Splenic conditioning: further loss of membrane surface area

Rbc entrapment in splenic cords

Macrophage removal of severely abnormal rbc

http://hsc.virginia.edu/medicine/clinical/pathology/educ/innes/text/rcd/membrane.html

Hereditary spherocytosis 1

Loss of membrane surface area  micro-spherocytosis

Blood Cells

summary erythrocytes
Summary - erythrocytes
  • Vast numbers, steady turnover
    • No nucleus, no mitochondria
    • Glucose essential as energy source
      • To maintain reducing conditions, and for ion pumps
    • Cells become “aged” as oxidation products build up
  • Lipid bilayer membrane, with attached & inserted glycoproteins
    • Cytoskeleton
      • Biconcave disc, but deformable
      • Flexibility important for proper plasma flow
    • Semi-permeable properties  lysis if not in isotonic medium
  • Will examine erythropoiesis in next lecture

Blood Cells

neutrophils 1
Neutrophils 1

Blood Cells

neutrophils 2
Neutrophils 2
  • The most abundant leukocyte in blood
  • Filled with granules
    • Lysosomes  fuse with ingested phagosome
    • Secrete toxic chemicals
  • Very active migration
    • Sensitive to chemotactic factors which attract them to infection site

Blood Cells

neutrophils functions as effector cells
Neutrophils: functions as effector cells
  • Synonyms
    • Polymorphonuclear leukocytes, = PMNs or “Polys”
  • Don’t stain strongly with either eosin or basic dyes
  • Raised numbers (“neutrophilia” during acute bacterial infections
    • Increased mobilisation from extensive reserves
    • Increased production from progenitors
  • Adhere to vessel walls and transmigrate to areas of infection in tissues (acute inflammation)
    • Engulf bacteria, kill rapidly with very toxic molecules (incl strong oxidisers comparable to bleach)
    • Collateral damage to host cells, plus dead bugs,  pus
  • Defects in adhesion molecules, or in killing mechanism,  serious pyogenic (pus-forming) infections

Blood Cells

eosinophils
Eosinophils
  • Generally larger than neutrophils
  • Stain orange-pink with eosin
    • Contain abundant basic protein
  • Elevated levels in
    • tropical parasite infections
      • defence against single-celled and multicellular parasites
    • Chronic allergic conditions
      • May reduce hypersensitivity via histaminase

Blood Cells

eosinophil e m
Eosinophil e-m
  • Large ovoid granules, very electron dense
  • Actively phagocytic
  • Passively adsorb certain kinds of antibodies
    • helps them recognise targets
  • Can exocytose (spit out) toxic substances
    • Different chemicals from neutrophils

Blood Cells

basophil
Basophil
  • Least common leukocyte
  • Stains with basic dyes
  • Precursor of mast cells in tissues
    • Mast cells release histamine etc in allergies

Blood Cells

monocyte
Monocyte
  • ~5 – 10% of wbcs
  • Nucleus often kidney-shaped
  • No obvious granules
  • Precursor of macrophages in tissues
    • Macro = “big”; phage = “eat”

Blood Cells

lymphocyte
Lymphocyte
  • Next most common after neutrophils
  • No obvious granules
    • Except for Natural Killer subset
  • Subsets
    • T, B

Blood Cells

lymphocytes

Autoradiograph by J.L. Gowans

Lymphocytes
  • Vary in size
    • Small = very dormant, out of cycle, long-lived
      • After stimulation  memory cells
    • Large = rapidly dividing
      • Incorporate DNA synthesis precursors
      • “Lymphoblasts”
      • Intermediates before full maturation
        • To antibody-forming cells in tissues (very rare in blood)
        • To cytotoxic T lymphocytes
  • These are from lymph (hence no rbc)

Blood Cells

lymphocytes patrol continuously
Lymphocytes patrol continuously
  • To connect functionally all the dispersed lymphoid tissues
    • Blood  lymph  blood …….
  • Leave blood at special endothelium only on certain venules, within lymph nodes, tonsils & other lymphoid organs
  • Ensures rare clones with a given specificity have a good chance of encountering their specific stimulus

http://www.geocities.com/CapeCanaveral/Hangar/1962/page3.html

Blood Cells

lymphocytes emigrate via high endothelial venules
Lymphocytes emigrate via High Endothelial Venules
  • To seek antigens outside the blood stream, which have been filtered, processed and presented by cells in lymphoid tissues

Blood Cells

summary of leukocytes 1
Summary of leukocytes 1
  • Granulocytes include:
    • Neutrophils
      • Move promptly into tissues to deal with any noxious event harmful to the body: infection, tissue damage and so on
      • Engulf, kill and digest invaders  inflammation and perhaps pus
    • Eosinophils
      • Defence against some parasites, in collaboration with antibodies
      • May diminish some immediate-type hypersensitivity reactions
    • Basophils
      • Move into tissues to become mast cells

Blood Cells

summary of leukocytes 2
Summary of leukocytes 2
  • Monocytes
    • Precursors of tissue macrophages, slower to act than neutrophils
  • Lymphocytes
    • Each call has one specificity for antigen, therefore needs to continuously patrol to meet up with the right molecule
      • perpetual motion into and out of vasculature - “lymphocyte recirculation”

Blood Cells

reading and online materials
Reading and online materials
  • Wheater’s Functional Histology, 4th edn, chapter 3
  • http://www-micro.msb.le.ac.uk/MBChB/2a.html
    • Leicester, immune system cells only, not complete haematology
  • http://www.medinfo.ufl.edu/year1/histo/review/lab11.html
    • Florida, in-depth, with self-test Qs at: http://www.medinfo.ufl.edu/year1/histo/quiz/mh10.html
  • http://www.ultranet.com/~jkimball/BiologyPages/B/Blood.html#RBCs
    • (textbook account, excellent hyperlinked glossary)
  • http://www.aum.iawf.unibe.ch/HemoSurf/Demo_E/Gallery/NRB_fr.htm
    • (proper haematologist’s site)

Blood Cells