Constituents Of Blood Flashcards
Anaemia
haemoglobin level in blood below the normal level for age and sex
• male <130g/L + female <110g/L
classified according to the size of the red cells into macrocytic, normocytic and microcytic
• The Reticulocyte count, morphology of the red cells and changes in the white cell and/or platelet count help diagnose anaemia
Features of anaemia
shortness of breath on exertion, pallor of mucous membranes, malaise and angina in older people and tachycardia. Potentially also jaundice, leg ulcers
Causes of anaemia
Corpuscular (mainly haemolysis):
• membrane (eg spherocytosis, PNH)
• Haemoglobin (eg haemoglobinopathy)
• Enzymes (eg G6PD, PK)
Extra-corpuscular:
• Production mismatches- hypoplastic (not enough), dyshaemotpoeitic (ineffective production)
• Increased removal of erythrocytes- acute blood loss (haemorrhage), haemolytic anaemia, auto or alloimmune, mechanical damage, other (eg heat, chemicals)
• Deficiencies of iron, folate (macrocytic anaemia) or vitamin B12 (pernicious)
• Redistribution- hypersplenism
Classifying anaemia-hypoxia
Hypo regenerative (don’t produce red blood cells) vs hyper-regenerative (loading reed blood cells)
Reduced production vs increased destruction
Corpuscular (problem with cells) vs extra-corpuscular
Haematocrit (PCV)
ratio of volume of red blood cells to total volume (45%)
Reticulocyte
final precursor to erythrocyte. Contains some rRNA and is able to synthesis haemoglobin. Circulates in the peripheral blood for 1-2 days before maturing
Erythropoesis
regulated by the hormone erythropoietin (a heavily glycosylated polypeptide)- secreted by the kidney in response to hypoxia
• It stimulates Erythropoesis by increasing the number of progenitor cells
• Hypoxia induces synthesis of hypoxia-inducing factors (HIF-1 alpha and beta) which stimulate erythropoietin production and also new vessel formation and transferrin receptor synthesis, and reduces hepcidin synthesis, increasing iron absorption.
• Erythropoietin production increase in anaemia
Erythrocytes
120 day lifespan
• 6-8um diameter
• Anuclear biconcave disc- flexible
• Contain haemoglobin and glycolytic enzymes
• Able to generate ATP by anaerobic glycolytic pathway
• Oxygen and carbon dioxide transport
• removed in spleen, liver, bone marrow (reticuloendothelial system) and through blood loss
Haemoglobin
capable of reversibly binding to oxygen
• haemoglobin synthesis occurs largely in the mitochondria by a series of biochemical reactions commencing with the condensation of glycine and succinyl co-A under the action of the rate limiting enzyme aminolavulinic acid synthase (ALA)
• Protoporphyrin combines with iron in the ferrous state to form haem
• A tetramer of 4 globin chains each with its own haem group then form to make a haemoglobin molecule
• mutations or absence of alpha or beta chains in adults referred to as thalassemia (potentially causing anaemia)
Thalassemia
mutations or absence of alpha or beta chains in adults referred to as thalassemia (potentially causing anaemia)
innate immunity
neutrophils, eosinophils, basophils, macrophages, mast cells- non-specific immunity (cytokines, phagocytosis, cytotoxicity)
Adaptive immunity
B cells, T cells- antigen-specific immunity (cytokines, antibodies, cytotoxicity)
Myelopoiesis
production of white blood cells, hormonal factor: granulocyte-macrophage colony stimulating factor (GM-CSF)- only stimulate production of myeloblastic white blood cells not lymphoid cells
Neutrophils
phagocytotic and release chemo- and cytokines to induce inflammation
Monocytes function
mature into either macrophages or dendritic cells (both antigen presenting)
Basophils
mature into mast cells.
Express surface IgE and release histamine- role in allergies an immunity (anaphylaxis)
Stain blue
Eosinophils function
Antagonistic to basophils: decrease mast cell histamine secretion
particular role in fighting parasitic infections but also wide range of regulatory functions
Where are Granulocytes produced
Made in the bone marrow under the control of a variet of growth factors and have a short lifespan in the blood stream before entering tissues
Lymphocytes
immunologically competent white cells that are involved in antibody production (B cells) and with the body’s defence against viral infection or other foreign invasion (T cells)
• Arise from haemopoietic stem cells in the marrow- T cells processes in the thymus
Only blood cells that divide
B cells
secrete antibodies specific for individual antigens (M, A, G, E, D)
20% of lymphocytes
T cells
further subdivided into helper (CD4+) and cytotoxic (CD8+) cells- they recognise peptides in HLA antigens
• Natural killer cells are cytotoxic CD8+ cells that kill target cells with low expression of HLA molecules
80% of lymphocytes
Immune response
occurs in the germinal centre of lymph node and involves B cell and T cell proliferation, somatic mutation, selection of cells by recognition of antigen on antigen-presenting cells and formation of plasma cells or memory B cells
Plasma
fluid component of blood- 55%
• Transportation medium which contains water, salt, glucose and proteins
Serum
Blood plasma without clotting factors
Cryoprecipitate
centrifuged blood plasma containing clotting factors and fibrinogen
Plasma proteins
• albumin: produced in the liver. Determines oncotic pressure of blood, keeps intravascular fluid within that space, lack of albumin leads to oedema, seen in liver disease and nephrotic syndrome ( loss of protein in urine)
• carrier proteins
• Coagulation proteins
• Immunoglobulins- produced by activates B cells
Platelets
circulate in an inactive form
• Anucleate cells- structure: plasma membrane, cytoskeleton, dense tubular system, secretory granules ( alpha, eg VWF, PF4, plasminogen; dense eg serotonine; lysosome; peroxisome)
• 2-5um
• Lifespan- 7-10 days
Where are platelets derived from
megakaryocytes- enter endomitosis where the chromosomes duplicate but the cells do not divide (polyploidy)
• Membrane blebbing process- platelets break off from the megakaryocytes. 1 megakaryocyte can produce up to 4000 platelets
• Stimulated by thrombopoietin
5 stages of platelet plug formation
- Endothelial injury
- Exposure
- Adhesion
- Activation
- Aggregation
Endothelial injury
• smooth muscle cells and nerves detect injury
• trigger reflexive contraction of vessel (vascular spasm)- reduces blood flow and loss
• Secretion of nitric oxide and prostaglandins stop- secretion of endothelin begins → further contraction
Exposure
• damage to endothelial cells exposed collagen
• Damaged cells release Von Willebrand factor (binds to collagen)
Adhesion
GP1B surface proteins on platelets bond to Von Willebrand factor
Activation
• platelet changes shape (smooth to spiculated, increasing their surface area), releases more Von Willebrand factor, serotonin, calcium, ADP, thromboxane A2 (positive feedback loop)
• ADP and thromboxane A2 result in GPIIB/IIIA expression
Aggregation
GPIIB/IIIA binds to fibrinogen and links platelets → platelet plug
3 Anti-clotting systems
- Tissue factor pathway inhibitor inhibits the tissue factor–factor VIIa complex.
- Protein C, activated by thrombin, inactivates factors VIIIa and Va.
- Antithrombin III inactivates thrombin and several other clotting factors.
Fibrinolytic system
Clots are dissolved by the fibrinolytic system.
• A plasma proenzyme, plasminogen, is activated by plasminogen activators to plasmin, which digests fibrin.
• Tissue plasminogen activator is secreted by endothelial cells and is activated by fibrin in a clot.
Anti-clotting drugs
• Aspirin inhibits platelet cyclooxygenase activity thereby inhibiting prostaglandin and thromboxane production—this inhibits platelet aggregation.
• Oral anticoagulants and heparin interfere with clotting factors—they prevent clot formation.
• Recombinant tissue plasminogen activator (t-PA) is a thrombolytic—it dissolves blood clots after they are formed.
Platelet plug formation
- Injury to a vessel disrupts the endothelium and exposes the underlying connective-tissue collagen fibres.
- Platelets adhere to collagen, largely via an intermediary called von Willebrand factor (vWF), a plasma protein secreted by endothelial cells and platelets. This protein binds to exposed collagen molecules, changes its conformation, and becomes able to bind platelets- vWF forms a bridge between the damaged vessel wall and the platelets
- Binding of platelets to collagen triggers the platelets to release the contents of their secretory vesicles, which contain a variety of chemical agents. Many of these agents, including adenosine diphosphate (ADP) and serotonin, then act locally to induce multiple changes in the metabolism, shape, and surface proteins of the platelets, a process called platelet activation. Some of these changes cause new platelets to adhere to the old ones, a positive feedback phenomenon (platelet aggregation), which rapidly creates a platelet plug inside the vessel.
- Chemical agents in the platelets’ secretory vesicles are not the only stimulators of platelet activation and aggregation. Adhesion of the platelets rapidly induces them to synthesize thromboxane A2, a member of the eicosanoid family, from arachidonic acid in the platelet plasma membrane. Thromboxane A2 is released into the extracellular fluid and acts locally to further stimulate platelet aggregation and release of their secretory vesicle contents
Platelet biochemistry
Activation → adhesion → aggregation → amplification pathways
Which clotting factor is tested for as part of a prothrombin time test
Factor VII as part of extrinsic clotting pathway
Blood coagulation: clot formation:
- Blood is transformed into a solid gel when, at the site of vessel damage, plasma fibrinogen is converted into fibrin molecules, which then bind to each other to form a mesh.
- This reaction is catalyzed by the enzyme thrombin, which also activates factor XIII, a plasma protein that stabilizes the fibrin meshwork.
- The formation of thrombin from the plasma protein prothrombin is the end result of a cascade of reactions in which an inactive plasma protein is activated and then enzymatically activates the next protein in the series.
• Thrombin exerts a positive feedback stimulation of the cascade by activating platelets and several clotting factors.
• Activated platelets, which display platelet factor and binding sites for several activated plasma factors, are essential for the cascade. - In the body, the cascade usually begins via the extrinsic clotting pathway when tissue factor forms a complex with factor VIIa. This complex activates factor X, which then catalyzes the conversion of small amounts of prothrombin to thrombin. This thrombin then recruits the intrinsic pathway by activating factor XI and factor VIII, as well as platelets, and this pathway generates large amounts of thrombin.
• The liver requires vitamin K for the normal production of prothrombin and other clotting factors.
Plasminogen
precursor for an enzyme that lyses clots
Anticlotting Systems:
- Tissue factor pathway inhibitor inhibits the tissue factor–factor VIIa complex.
- Protein C, activated by thrombin, inactivates factors VIIIa and Va.
- Antithrombin III inactivates thrombin and several other clotting factors.
• Clots are dissolved by the fibrinolytic system.
• A plasma proenzyme, plasminogen, is activated by plasminogen activators to plasmin, which digests fibrin.
• Tissue plasminogen activator is secreted by endothelial cells and is activated by fibrin in a clot.
Anticlotting drugs:
• Aspirin inhibits platelet cyclooxygenase activity thereby inhibiting prostaglandin and thromboxane production—this inhibits platelet aggregation.
• Oral anticoagulants and heparin interfere with clotting factors—they prevent clot formation.
• Recombinant tissue plasminogen activator (t-PA) is a thrombolytic—it dissolves blood clots after they are formed.
Clotting screen:
• PT- prothrombin time (activity of Factor VII)
• APRT- activated partial thromboplastin time (activity of factor IX and factor XI)
• Fibrinogen
• Used to measure activity of medications eg warfarin which inhibits Factor VII, so increased PT/ heparin » elevated APRT
• Used to show haemophilia A/B/C- raised APRT without elevated PT
Constituents the coagulation cascade:
A series of proteolytic enzymes that circulate in plasma in an inactive form and generate thrombin when activated
Functions of the liver
• responsible for synthesising coagulation factors and fibrinogen
• Filtration of portal blood
• Detoxification
• Biliary drainage
• Glucose regulation
• Protein, amino acid, vitamin, fat synthesis and storage
Extrinsic coagulation pathway
damage to the endothelial lining of vessels releases tissue factor which activates factor VIIa. Factor VIIa has direct effect on factor Xa to initiate common part of cascade
Intrinsic coagulation cascade
blood contracting endothelial collagen outside the lumen activates factor XIIa (serum protease) → XIa → IXa. Factor Xa then activated by IXa along with cofactors VIII, phospholipids and Ca2+
Common pathway of coagulation cascade
Xa converts prothrombin (factor II) into thrombin (factor IIa)
• Thrombin converts fibrinogen (I) into fibrin (Ia) and activates factor XIII - leading a the crosslinking of fibrin and a clot
• Thrombin also gives positive feedback on factors V, VII, VIII and XI but also prevents over-coagulation by activating plasmin which is fibrinolytic
Order of clotting factors in intrinsic coagulation pathway
12 → 11 → 9 → 8 → 10
thrombin
fibrinogen is converted into fibrin molecules, which then bind to each other to form a mesh.
This reaction is catalyzed by the enzyme thrombin, which also activates factor XIII, a plasma protein that stabilizes the fibrin meshwork.
Which vitamin is required by the liver for normal production of prothrombin and other clotting factors
Vitamin K
Platelet activation:
Shape change =
• Smooth discoid → spiculated and pseudopodia
• increases surface area
• Increases possibility of cell-cell interactions
Glycoprotein IIb/IIIa receptor =
• on surface of platelet (50000-100000 copies on resting platelet)
• Activation increase number of receptors and affinity of receptor for fibrinogen
• Fibrinogen links receptors, binding platelets together (platelet aggregation)
• Also known as integrin (alpha)IIb(beta)3
Release of alpha and dense granules
Why does the platelet plug not spread along the normal endothelium
the latter secretes prostacyclin and nitric oxide, both of which inhibit platelet aggregation.
COX-1
mediates GI mucosal integrity and thromboxane A2-mediated platelet aggregation (+ vasoconstriction)
COX-2
Mediates inflammation. Involved in prostacyclin production, which inhibits platelet aggregation and affects renal function
Arachidonic acid
converted into prostaglandins by COX (cyclooxygenase)
Platelet receptors
After atherosclerotic plaque rupture- platelets adhere to damaged vessel wall
• Collagen receptors bind to subendothelial collagen which is exposed by endothelial damage
• GPIIb/IIIa also binds to von Willebrand factor (VWF) which is attached to collagen
• Soluble agonists are also released and activate platelets leading to shape change, cross-linking of GPIIB/IIIa and platelet aggregation
• Aspirin inhibits an amplification pathway
Platelet purinergic receptors: ADP-induced platelet aggregation
• platelet P2Y receptors- different G proteins link to different signalling pathways
ADP activates P2Y1
• Causes platelet activation
• Results in GPIIb/IIIa fibrinogen cross-linking and aggregation
ADP activates P2Y12
• amplification of platelet activation, aggregation and platelet dense granule release
• Inhibits adenylate cyclase so drop in cAMP levels
ADP activates P2Y1
• Causes platelet activation
• Results in GPIIb/IIIa fibrinogen cross-linking and aggregation
ADP activates P2Y12
• amplification of platelet activation, aggregation and platelet dense granule release
• Inhibits adenylate cyclase so drop in cAMP levels
Which granules release ADP
Dense granules
Thrombin activates protease-activated receptors (PAR) on platelets
This leads to platelet activation and release of ADP, which amplifies this activation
Amplification loops:
• Dense granules release ADP, which causes further activation
• Activation of GPIIb/IIIa also amplifies platelet activation
Platelet procoagulant activity:
activated platelets catalyse thrombin generation, creating an amplification loop that also links with coagulation (the production of fibrin)
1. Aminophospholipids maintained on inner layer phospholipid bilayer- asymmetry
2. Platelet activation eg thrombin activating PAR1
3. Ca2+ release from intracellular stores
4. Activates enzyme scramblase which flips translocase enzyme so aminophospholipids flip to outer surface
5. Allows promthrombin complex to be assembled
6. Catalysed thrombin production via prothrombinase
• Inhibition of translocase and activation of scramblase leads to expression of aminophospholipids on the outer platelet membane, which allows assembly of prothrombinase complex and generation of thrombin
Additional platelet receptors
feed into these amplification loops eg release of thromboxane A2
Platelet activation causes:
• Shape change
• Cross-linking of GPIIb/IIIa
• Platelet aggregation
Fibrinolytic system
• A dynamic interaction between fibrinolytic and anti-fibrinolytic factors is designed to maintain homeostasis i.e. haemostasis without thrombosis
• Stop blood clotting
Fibrinolytic pathway
tPA
Plasminogen
Plasmin
Fibrin
Fibrin-degradation products
Platelets and inflammation:
Platelets have pro-inflammatory and prothrombotic interactions with leukocytes and release inflammatory mediators from alpha granules
Platelets and inflammation: monocytes
Cytokines e.g. chemotactic molecules
Proteolytic Enzymes
Pro-thrombotic molecules : Tissue factor
Adhesion Molecules e.g. PSGL-1
Platelets and inflammation: alpha granules
Inflammatory mediators (vWF, thromboxane A2)
Adhesion Molecules e.g. P-Selectin binds to PSGL-1 on monocytes (pro-inflammatory)
Coagulation Factors (fibrinogen)
Platelet alpha granules
Mediate expression of surface P-selection and release of inflammatory mediators
Overview: platelet aggregation
• Mediated by GPIIb/IIIa activation (final common pathway)
• Cross linking of platelets by fibrinogen
Overview:platelet adhesion
• Collagen and VWF exposed
• Mediated by GPIb, alpha2beta1 and GPVI
Overview: platelet granules
• Mediators of vessel response to injury
• Inflammatory mediators released from a granules
Overview: Thrombin → Coagulation cascade:
• Activated platelets support thrombin generation
• Inhibited by anticoagulant drugs
Inhibitor of thromboxane A2 amplification pathway
Aspirin
Inhibitors of P2Y12 amplification pathway
Clopidogrel, Prasugrel, Ticagrelor
Percentage of blood made up of erythrocytes
43%
Percentage of blood made up of white blood cells
1%
Eosinophil structure
Bilobed nucleus
IgE receptors
Stain orange
Granulocyte
Neutrophil granules
1’- lysosomes- myeloperoxidase and acid hydrolases
2’- inflammatory mediators
3’- gelatinases and adhesion molecules
Where are Kupffer cells found
Liver
Where are microglia macrophages found
CNS
Monocyte structure
Reuniform nucleus
Agranulocytes but have granules
Lifespan of erythrocytes
120 days
Sites of erythropoesis
In utero- yolk sac ——> liver and spleen
Children - all bones
Adults - REFS (ribs, ends of long bones, flat bones of skull and pelvis, spine)
Where are RBCs destroyed
Liver, bone marrow and spleen
Where is erythropoietin made
Kidney
HbA
2 alpha chains
2 beta chains
HbF
2 alpha chains
2 gamma chains
HbA2
2 alpha chains
2 delta chains (delta = disease)
Thalassemia
Congenital mutations of alpha or beta chains
Thrombopoesis
Megakaryocytes ——> endomitosis - chromosomes duplicate without cell division ——> membrane blebbing
Where is thrombopoeitin produced
Liver and kidneys
Activation shape change
Smooth discoid ——> spiculated with pseudopodia (increased surface area)
Haemophilia
Inability to make clotting factors
Type A = factor 8 deficiency
Type B = factor 9 deficiency
Thrombocytopenia
Platelet deficiency
Decreased production, increased destruction and altered redistribution
Pseudothrombocytopenia
Platelet clumping
Primary haemostats is
- Injury- vasoconstriction via endothelin release
- adhesion- exposed collagen —> vWF —> GP1b —>platelet
- Activation- pseudopodia and granule release
- Aggregation- platelet-GP2b/3a-fibrinogenGP2b/3a-platelet
Vitamin K dependant factors
10, 9, 7, 2
Virchow’s triad
Circulatory stasis + endothelial injury + hypercoaguable state = thrombosis
Overview of coagulation cascade
Intrinsic: 12 —> 11 —> 9 —> 8 —> 10
Extrinsic: 3 (tissue factor) + 7a —> 10
Common: 10 —> 5 —> 2- thrombin (from prothrombin) —> 1- fibrinogen —> fibrin mesh
Degradation of fibrin
Plasminogen —> plasmin
Due to 7a (intrinsic) and endothelium —> tPA (extrinsic)
What are required for factor 9a and 10a function
Phospholipids
Ca2+
Factor 5
+ve feedback: thrombin and phospholipids bilayer
Thrombin —> PAR1 or PAR4 receptors—> influx of Ca2+
1. Blocks translocase
2. Activates scramblase
Aminophospholipid expression on outer platelet membrane
Prothrombinase complex assembly
Prothrombin ——> thrombin
Low dose aspirin
Inhibits COX1 so less thromboxane A2—> less activation/aggregation
High dose aspirin
Inhibits COX1 and COX2
Reduction in aggregation mediated by platelets and inhibition of prostacyclin mediated inhibition, which means more aggregation
Malabsorption of what substance is most likely to cause excessive bruising and recurrent nosebleeds
Micelles- contain fat-soluble vitamins (ie vitamin K) which is essential for formation of Factor VII, IX, X and prothrombin
Deficiency of vitamin K= excessive bleeding due to reduced clotting factors
Role of factor VIII
Converts the loose fibrin meshwork into a more stable network to allow entrapment of red blood cells (now the clot can begin to form)
Which clotting factors are vitamin K dependent
Factor II, VII, IX, X
The gene coding for alpha subunits of haemoglobin is found on which chromosome
16
What is tested for when diagnosing pernicious anaemia
Anti-intrinsic factor antibodies
When someone starts to bleed, a platelet plug begins to form. This is due to the coagulation cascade, where a series of factors combine and catalyse to eventually form a stable fibrin clot. One of these coagulation factors is thrombin. Which of the following processes shows the formation of thrombin in the cascade?
Xa + Va catalyse II → IIa
