Heamostasis Flashcards
NORMAL HAEMOSTASIS
Haemostasis requires the interaction of three compartments
• The blood vessels
• The platelets
• The coagulation factors (soluble plasma proteins)
Must be tightly controlled!!
HAEMOSTASTIC MECHANISM
1)Haemostatic mechanism is three distinct phases:
• Primary haemostasis
• Interactions between blood vessels, platelets and vWF
2)Secondary haemostasis
•Pathways of coagulation to generate fibrin strands
3)Fibrinolysis
• Biochemicalsystemthatdegradesthefibrinclot
• Primary haemostasis
Interactions between blood vessels, platelets and vWF
Secondaryhaemostasis
Pathwaysofcoagulationtogeneratefibrin strands
Fibrinolysis
Biochemical system that degrades the fibrin clot
Layers of the blood vessel wall
Intima=inner layer, comprised of endothelial cells on a basement membrane of collagen and elastin fibres
Media=smooth muscle layer,responsible for regulation vascular tone
Adventia=outer layer comprised of collagen and fibroblasts to protect and anchor the vessel
Platelets
-no nucleus
-higher than normal thrombocytosis
-lower than thrombocytopenia
-circulate for 7/10 days
-produced from megakaryocytes
-100 billion produced a day
-major players of the haemostatic response,they adhere to the injured vasculature and to prevent bleeding
THROMBOPOIESIS
Phase 1-megakaryocyte maturation
-Endomitosis (DNA replication without cell division)
-Cytoplasm enlargement (cytoskeletal proteins and
platelet granules)
Phase II - Platelet generation
-Mature megakaryocytes extend long branching processes (proplatelets)
-Organelles and granules are transported to proplatelets
-Driven by cytoskeletal rearrangements
Platelet structure
-Small
-Disc shape
-No nucleus
-Plentiful
-it has dense granules,a-granules
-it has glycogen stores
-it has dense tubular system
-has an open canalicular system
Granules in platelets
-they can’t synthesise as they don’t have a nucleus
-granules allow storage of molecules that the platelet needs to perform their functions
-platelets contain 3 types of storage granules:dense,a-granules,Lysosomes
The role of platelets
• Their primary physiological role is in blood clotting
• Detect damaged vessel endothelium
• Accumulate at the site of the vessel injury
• Initiate blood clotting to block the circulatory leak
Platelets in a healthy blood vessel
• Platelets circulate close to the vessel wall
• Endothelium provides a barrier preventing platelets from coming into contact with prothrombotic matrix proteins eg
collagen
• Endothelium release nitric oxide, prostacyclin and CD39, which keep platelets in a non-active state
•in a healthy blood vessel there is an inhibition of platelet activation;vasodilation
What do platelets adhere to
• Von Willebrand factor (VWF) circulates in plasma and is released from damaged endothelial cells
• VWF binds collagen
• Platelets adhere to VWF allowing an indirect interaction with collagen
• Initial VWF-platelet interaction is weak tethering
• Allows more stable direct interactions between collagen and platelet receptors (GPVI and α2βI)
• Signalling through GPVI leads to platelet spreading, activation and secretion.
PLATELET GRANULE SECRETION & TXA2 RELEASE
• Platelet activation by collagen and thrombin leads to secretion of granules
• The potent platelet agonist Thromboxane A2 is also released
Platelet aggregation
• More platelets become activated due to the release of ADP and TXA2 and are recruited to the site of damage
• Activated platelets bind fibrinogen linking (aggregating) them together
• Activated platelets bind fibrinogen via GPIIbIIIa.
• Form platelet-fibrinogen-platelet bridges.
Primary hemostasis
- Collagen (among other ECM proteins) is exposed following damage to the vasculature.
- vWF (von Willebrand Factor) – a plasma protein, binds to collagen and via interaction with
its receptor GPIb ‘tethers’ the platelets to the site of damage. - This tethering, slows the platelets down as they flow past, and initiates platelet ‘rolling’
- This enables the collagen receptor GPVI to bind to the collagen (strong interaction).
- Binding of collagen to GPVI triggers a signalling cascade (see later) which leads to:
• Granule secretion - releases pro-thrombotic factors such as ADP and that positively
feedback onto the platelets
• Production of TxA2 - positive feedback onto the platelets
• Platelet shape change – increases the surface area of the platelet
• Integrin activation – leads to platelet aggregation via the formation of fibrinogen
bridges.
Dense granules(mostly involved in primary haemostasis)
ADP
ATP
CALCIUM
SEROTONIN
A granules
AIIbβ3 (GPIIb-IIIa)
P-selectin
Fibrinogen
Von Willebrand factor (vWF)
Coagulation factors
Other pro-activatory factors, cytokines etc
Lysosomes
Contain:hydrolytic enzymes
Elastase
Collagenase
Heparinase
What happens when there is a damage to the blood vessels
-sticky collagen is exposed
-endothelial cells damage causing them to become activated(releasing VWF)
-VWF factor binds to collagen through a receptor(GPIb).This happens in a really fast flowing blood
-Platelets adhere to VWF allowing an indirect interaction with collagen
-The initial VWF-platelet interaction is weak tethering,it slows the fast flowing blood and allows for more stable direct interactions between collagen and platelet receptors(GPVI. And a2BI).Signallimh through GPVI leads to platelet spreading,activation and secretion.
-platelets spread by forming pseudopia and then lamellopodia in order to maximise SA to minimise blood loss
-platelet activation by collagen and thrombin leads to to secretion of granules(TXA2,ADP,5-HT)|.Theu are secondary mediators which play part in the positive feedback mechanism:platelets that are flowing by get activated and grow the clot forming a platelet plug
-txa2 causes vaso-constriction, it is also a platelet agonist=it activates platelet potency.
Platelet aggregation:
-Activated platelets bind fibrinogen via GPIIbIIIa. Forming platelet-fibrinogen-platelet bridges.(fibrinogen is bi-valent so it can have 2 platelets binding)
Platelet plug formation:
-Activated platelet expose phosphatidylserine (PS) providing phospholipid surface coagulation as it changes the charge of the membrane (secondary haemostasis)
• Soluble fibrinogen is converted to insoluble fibrin stabilising the thrombus
Platelet plug formation
• Activated platelet expose phosphatidylserine (PS) providing phospholipid surface coagulation (secondary haemostasis).(PS in the platelet membrane is flipped to the outside of the platelet).Catalysing coagulation by providing the PL surface for the prothrombuinase and tenses complexes
• Soluble fibrinogen is converted to in soluble fibrin stabilising the thrombus
Sample collection for platelet tests-pre analytical variables
• Appropriate needle size to avoid ‘traumatic’ venepuncture
• Avoid haemolysis
• Use polypropylene or siliconized glass tubes/syringes
• Minimise time between sampling and analysis
• Keep tubes at room temperature
Tubes used for full blood count
EDTA tube(anticoagulant)
Strong CA2+chelator
Tubes used for platelet function assays
Citrate tube
Love ph
Mild ca2+ chelator
PLATELET TESTS – FULL BLOOD COUNT
• PLT count (number) - 150 – 400 x 109/L • Thrombocytopenia <150 x 109/L
• Thrombocytosis >400 x 109/L
• MPV (size) (PDW, P-LCR) • 8-12 fL
• Indication of size
• Measured in femtolitres (fL)
• High MPV → CML, hyperthyroidism, risk factor for CVD • Low MPV → Drug toxicity, chronic kidney failure
• Immature Platelet Fraction (reticulated platelets) (1-7%) • Immune thrombocytopenia eg when bone marrows releases platelets that are not ready
