Haematology Flashcards
What are the constituents of blood?
plasma = 55% - water, electrolytes, plasma proteins, clotting factors
RBC = 45%
platelets 0.5% - travel near edges
white cells
Describe the structure of a platelet…
spindle shaped
2-4um
no nucleus
extensive cytoplasm
Describe the production of platelets. what is life expectancy and how are they removed?
Thrombopoietin from liver/kidney during inflammatory states
causes megakaryocytes in bone marrow to differentiate into platelets
life expectancy = 7-10days
removed by macrophages in spleen
What is Von willebrand factor
strand like protein
when inactive remains curled up
when exposed to turbulent flow/sheer stress unwinds by mechanical force
can bind collagen and platelets to help platelet adhering to area of vessel wall injury.
normal values for WBC, Hb, Platelets , haematocrit , MCV
Hb 130-170 men, 115-150 women (g/L)
platelet 150-400 x 10^9 per L
MCV 80-100
haemocrit 0.4 to 0.5
WCC 4-10 x 10^9 /L
what is haemostasis?
the physiological process of stopping bleeding and first stage of vessel repair.
consists of 3 major steps
- vasoconstriction,
- platelet plug formation
- formation of fibrin clot (via coagulation cascade)
Give an overview of the process of haemostasis
- vessel injury and bleeding
- vasospasm - mediated by endothelin - reduces blood loss and blood flow
- collagen from vessel wall (subendothelial) exposed
- PLATELET ADHESION: vWF is circulating in blood curled up (also present in endothelium) sheer stress of wall injury (turbulent flow) causes it to unwind exposing binding site for platelets GP1b receptor and collagen. hence vWF bridges collagen and platelets allowing platelets to aggregate at site of injury
- PLATELET ACTIVATION: the binding of GP1b to vWF activates platelets. also other factors activate platelet. (thromboxane, ADP, Thrombin, GPIIa/IIIb - positive feedback mechanism).
- Activation results in release of dense bodies (ADP , Ca) and alpha granules (vWF, fibrinogen) and other mediators (thromboxane, 5HT3). Positive feedback mechanism to activate more platelets and more adhesion/ aggregation.
- PLATELET AGGREGATION - activation results in fibrinogen release and GPIIb/IIIa activation. now vWF and fibrinogen can bind this receptor and cause aggregation of platelets through bridging.
- FIBRIN FORMATION - mean while clotting cascade has activated and produces factor IIa. This converts fibrinogen to fibrin to stabilise the clot!
What can activate platelets and what does this result in?
Activation of platelets
- GP1b and vWF binding
- GPIIb/IIIa binding fibrinogen/vWF
- thromboxane A2 receptor - thromboxane is released as part of inflammation from arachidonic acid pathway & released by platelets.
- ADP - released by platelets binds P2Y12 (GPCR) - results in platelet activation.
- Thrombin via PAR receptor
activation results in release of ..
- alpha granules - fibrinogen and vWF - strengthen clot
- dense bodies - ADP, Ca
- other mediators
outcome…
a lot of the mediators result in positive feedback for more platelet activation
fibrinogen/vWF - aggregation and strengthening of platelet plug
Ca - bridges between negative clotting factors and negative charges on platelet membrane to bring 2 pathways in physical contact
thromboxane, 5HT3 cause vasoconstriction
ADP binding P2Y causes cytoskeletal changes
what is the coagulation cascade
Divided into intrinsic and extrinsic pathway
consists of a cascade of proteolytic enzymatic reactions which results in amplification at each stage such that high quantities of thrombin are made
the main goal is to produce factor IIa (Thrombin) which catalyses fibrinogen to fibrin to stabilise the platelet plug and create a clot (soluble fibrinogen to insoluble fibrin)
Describe steps of coagulation cacade
Extrinsic pathway
- Tissue damage causes activation of factor III (tissue factor) to IIIa
- IIIa –> VIIa
Intrinsic pathway (triggered by mediators in blood e.g. Ca)
- F12 –> F11 –> F9 –> F8
- 3a, 7a (extrinsic) form complex which can catalyse X to Xa
- F8a is a cofactor for F9a (intrinsic) which also catalyses X to Xa
Xa –> 2a
2a –> fibrinogen to fibrin
fibrin polymerises - insoluble - strong clot
factor 13 forms cross bridges within fibrin to stabilise further
what are the fibrin , thrombin, TF , Ca known as in terms of the clotting factors..
Fibrinogen - factor 1 (fibrin 1a)
Thrombin = 2/2a
TF = 3/3a
Ca = factor 4
what is meant by secondary homeostasis ?
this describes the second part of haemostasis whereby platelet plug is matured by cleaving fibrinogen to fibrin tpo form a clot.
where is tissue factor found?
extracellular matrix of smooth muscle within vessel wall so exposed when vessel is damaged
what are the roles of thrombin?
cleaves fibrinogen to fibrin - stabilises clot
activates factor 13 which forms cross bridges to further stabilise the clot
can also activate more of the previous factors - positive feedback
bind PAR receptor on platelets to activate platelets
activates protein C - controls the amount of clotting by inhibiting clotting factors 5 and 8
what can inhibit platelet aggregation/activation?
NO
prostacyclin
what is the difference between PT and APTT?
PT = prothrombin time = test of extrinsic pathway (add tissue factor to sample and time how long for clot to form)
APTT = activated partial thromboplastin time = test of intrinsic pathway (add phospholipids and an activator to see how long it takes clot to form)
both also test common pathway
one T = the other T is an ex so its on its own.
what is the role of vitamin K
responsible for activation of clotting factors - gives them a negative charge such that Calcium can bridge them to platelets
gamma-glutamyl carboxylase - adds carboxyl group to create this negative charge
vitamin K is oxidised in this reaction and needs to be recycled via vit K epoxide reductase
normal value for PT and APTT
PT = 12-13 sec
APTT = 30-50 sec
what is the thrombin time?
thrombin added to sample and time how long the common pathway takes
why may factor Xa assays be measured?
see effectiveness of LMWH
particularly useful for those in extremes of weights were may not be getting the full benefit.
what is an INR?
international normalised ratio
ratio of PT compared to average PT of a control sample
what increases PT?
anything affecting common pathway
- Xa inhibitors- factor x
- LMWH - factor X and II
- Warfarin / liver disease / vit K deficiency - factor 10, 2
anything affecting extrinsic
- warfarin - factor 7
DIC - consumption of clotting factors
what increases APTT?
Anything affecting common pathway
- Xa inhibitors- factor x
- LMWH - factor X and II
- Warfarin / liver disease / vit K deficiency - factor 10, 2
intrinsic pathway -
haemophilia - 8 or 9
DIC
what might delay thrombin time?
thrombin time is just measuring common pathway - adds in thrombin and measures how long. so anything below this that is abnormal can cause issues.
e.g. fibrinogen deficiency
heparin and dabigatrin as these block thrombin (IIa)
What is fibrinolysis?
the breakdown of a clot whereby fibrin is degraded by plasmin
plasminogen –> plasmin (protease)
much slower than coagulation/ clot formation and hence allows vessel to heal.
activated by tissue plasminogen activator (tPA) which is slowly released from damaged tissue which converts plasminogen to plasmin
what is d dimer?
by product of breakdown of fibrin clot
what is thrombolysis?
This is a medical management whereby the fibrinolysis pathway is utilised to breakdown a life threatening clot. (P.E, MI, limb ischaemia, stroke)
e.g. thrombolytic drugs include alteplase, streptokinase
what is ROTEM?
Diagnostic test to evaluate the haemostatic process in whole blood to aid transfusion therapies
provides a evaluation of clot formation, stabilisation and dissolution
a sample of blood is rotated and causes blood to clot which changes the viscoelastic properties of the sample which can be measured by resistance to rotation
describe what each part of a rotem describes…
R time = stimulation until beginning of clot formation. determined by clotting factors. if prolonged need FFP
K time = time for width of graph to reach a certain point and is a measure of time taken to reach a particular firmness of clot. determined by fibrinogen. hence if prolonged give cryoprecipitate
alpha angle = rate of clot formation. determined by fibrinogen. if low give cryoprecipitate
max amplitude (MA) - max extent of clot. determined by platelets. if low give platelets
LY30 - lysis at 30mins - measures clot breakdown. if high give TXA
what is DIC? causes?
disseminated intravascular coagulopathy
widespread activation of clotting cascade
microclots and depletion of clotting factors so bleeding.
infection/ sepsis
trauma
malignancy
transfusion
placental abruption/ amniotic fluid embolism.
what are the lab findings in DIC?
high APTT, PT , TT
low platelets
low fibrinogen
high d dimer
define thrombocytosis and thrombocytopenia?
> 450
<150
classify the cellular components of blood…
Classify drugs that work on the thrombotic system..
Also mention TXA = which promotes thrombosis by blocking the conversion of plasminogen to plasmin and hence the breakdown of fibrinogen.
these are all active drugs - can also mention replacement of certain products (platelets, factors) to improve thrombosis
Classify the antiplatelet drugs
Can be classified via mechanism.
Their mechanism works through inhibiting platelet activation and aggregation through various platelet receptors
most of these acts via Gi to activate platelets.
Activation
- P2Y12 receptor (ADP receptor) = clopidogrel, prasugrel, ticagrelor
- Thromboxane receptor = reduced thromboxan production via COX inhibitors.
- phosphodiesterase inhibitors = Dipyramiole (increases cAMP , opposite to Gi)
Aggregation and activation
- GPIIb/IIIa receptor = tirofiban
Classify the anticoagulant drugs
can be divided into two broad groups - parental and oral
parenteral
- direct thrombin (F2a) inhibitors = Bivalirudin
- indirect thrombin inhibitors = unfractionated heparin, LMWH (enoxaparin, dalteparin), fondaparinux
oral anticoagulants
- coumarins = warfarin
- direct Xa (DOAC) = apixaban, edoxaban, rivaroxaban.
- direct thrombin (IIa) = dabigatran
tell me about clopidogrel..
ADP receptor anatagonist works via inhibiting platelet activation through irreversible antagonism.
commonly used in high risk CVS events such as MIs and Strokes, stents or peripheral vascular disease to prevent further thrombosis and ischaemia.
75mg OD is usually given but often a loading dose of 300mg.
Pharmacodynamics:
- It reduces platelet activation and thus has some side effects related to this bleeding & bruising (type A reaction)
- it can also lead to thrombotic thrombocytopenia pupura - rare side effect where platelets form small clots throughout the body and platelet levels are depleted. (type B)
pharmacokinetics..
- It is taken orally and absorbed from GI tract
- mostly broken down by esterases but some converted to active form.
- prodrug and converted to active form by CYP450
- genetic polymorphisms - some people cant convert to prodrug v. well.
- omeprazole also inhibits this conversion
- 98% PB
- renal excretion.
tell me about aspirin…
commonly used antiplatelet - irreversible COX inhibitor COX1 > COX2
used for pain, antiplatelet actions in prevention and treatment of MI, strokes, PVD and also for its antipyretic effects.
can be given PO, PR usually 75mg or 300mg loading dose in acute setting (has a quicker onset)
pharmacodynamics:
- COX inhibition results in less prostaglandins (pain and inflammation), less thromboxane (less platelet activation)
- thus side effects can be type A - bleeding, bruising, GI ulceration, renal dysfunction and worsening of asthma (leukotrienes)
- or type B - allergy, reyes disease in children
pharmacokinetics:
- absorbed from stomach and intestine
- pKa 3.5, acid. acidity of stomach promotes absorption.
- 80% PB
- converted to salicyclates in liver and GIT
- renal excretion.
Tell me about dipyramidole?
Phosphodiesterase inhibitor
inhibits platelet activation
Main role is in stroke prevention and treatment. particularly when clopidogrel is contraindicated.
what are the main uses of GPIIb/IIIa inhibitors?
tirofiban
IV infusion, very short half life
used for coronary stenting
Tell me about unfractionated heparin
Heparin is an endogenous molecule normally produced by mast cells and works via binding ATIII to potentiate its inhibition of factor 2,10, 9.11 and 12.
It has now mostly been replaced by LMWH due to monitoring issues however still used in special circumstances e.g. acute limb ischaemia and vascular surgery.
given as IV infusion and requires close monitoring of APTT and platelet count.
Issues:
- type A - bleeding
- type B - hypotension with large IV bolus, hypersensitivity, osteoporosis, and heparin induced thrombocytopenia.
can you tell me about heparin induced thrombocytopenia?
This comes in 2 forms
type 1 = non immune, mild, self resolves, no need to stop infusion. develops few days after.
type 2 = immune mediated - IgG antibodies to platelet factor 4 (PF4) which results in platelet activation and destruction which inturn causes strokes and bleeding. , severe and develops on day 5-10.
can be distinguished by timing, level of platelet drop and is there is thrombosis.
do you know a reversal agent for heparin?
protamine sulphate.
heparin is very negatively charged and acidic and can be neutralised by protamine which is basic and positively charged.
tell me about LMWH…
LMWH is a newer form of heparin which has been fractionated and hence smaller.
IT also binds and potentiates ATIII however due to its size is limited to actions only on 2a and 10a.
it has become popular due to its more predictable nature and thus doesnt need monitoring and its improved side effect profile.
given subcutaneously
and needs adjustments in obesity and renal failure.
its downfalls
- cant be reversed by protamine (or atleast only slightly)
- can be underdosed in obesity - factor Xa levels can be checked in those where underdosing may occur
which anticoagulants can be used in pregnancy?
heparins because they dont cross placenta - although LMWH much more favoured in comparison to unfractionated due to monitoring and HITT
aspirin low dose can also be used and is used for pre-eclampsai
avoid warfarin - teratogenic
avoid DOACs
Tell me about warfarin
Commonly used oral anticoagulant. although more recently, its role is being replaced by newer agents such as the DOACs.
It is a coumarin based molecule
Works via inhibition of Vitamin K epoxide reductase. This inhibition prevents the reduction of vitamin K such that it cannot be replenished for its use in carboxylation of clotting factor and hence their activation
It mostly affects the production of factors 10, 9, 7 and 2 but also protein C and S
It only affects the synthesis of new clotting facotrs and hence takes around 3-5 days to work and should be bridged by LMWH. Also this bridging is useful as initially the inhibition of protein C and S gives pro-coagulant effects
It is taken orally and titrated to the INR and the range of INR depends on indication e.g. AF is 2-3 whereas a mechanical valve is 3-4.
pharmacodynamics
- bleeding
- N&V & diarrhoea
- teratogenic
pharmacokinetics
- good oral absorption
-metabolised by CyP450 and subject to inducer/inhibitors.
- many DDI = plasma protien binding (NSAIDs, phenytoin), CYP450 inhibitors (erythromycin) inducers ( phenytoin, COCP)
how is a bleeding patient who normally takes warfarin managed?
stop warfarin
high INR no bleed - oral vit K
bleeding, mild = vitamin K IV
major bleed = Prothrombin complex concentrate (beriplex) or FFP
what DOACs do you know? when are these indivaated
apixaban, edoxaban, rivaroxaban
indicated in venous thrombosis e.g. P.E, DVTs, prophylaxis for AF
which DOACs can be reversed?
Dabigatran = monoclonal Ab
rivaroxaban and apixaban = Andexanet alpha
which drugs are fibrinolytics ?
these are drugs that promote breakdown of fibrin clot.
normally plasminogen –> plasmin. this step is activated by tissue plasminogen activator.
plasmin cleaves fibrin.
there are a number of drugs that mimic tPA including streptokinase, alteplase
how does transexamic acid work?
anti-fibrinolytic agent
blocks conversion of plasminogen to plasmin to prevent breakdown of fibrin
aims to prevent/ help with haemorrhage
used in major trauma and traumatic brain injuries after CRASH 2 and 3 studies. also often used in orthopedic surgery.
name of 2 models of haemostasis
The Classical (Cascade) Model of Hemostasis
* clotting cascade
The Cell-Based Model of Hemostasis
* newer model incorporates the role of cells (especially platelets and endothelial cells)
previous Q on iron metabolism/ storage - how much iron, transferrin and ferritin in the body?
iron - around 3g (4g in male)
ferritin - males up to 400ng/ml
females 15-150ng/ml
transferrin - normal saturation of iron around 20-50%
what are transferrin and ferritin ?
ferritin is the intracellular storage protein for iron. used as a marker of iron deficiency/ overload
transferrin is the mechanism for transport of iron in the blood.
where is iron stored?
mostly in liver, bone marrow and spleen
how is iron absorbed and recycled?
absorbed from duodenum and upper jejunum
absorbed as Fe2+ and enhanced by vit C
hepcidin is a protein made by the liver that inhibits absorption for regulation
old damaged RBC are engulfed by macrophages and the iron in them is recycled.
what are the causes of iron deficiency anaemia?
excess loss
* menorrhagia, GI malignancy
insufficient production
* dietary - iron lack
* malabsorption - low stomach acid
is blood loss always pathological?
no - mensturation, venepuncture e.g. for haemochromatosis
