Thrombophilia Flashcards
What is thrombophilia
Disorders of the haemostatic system which are likely to predispose to thrombosis
Hereditary or acquired
Heritable thrombophilia is an inherited tendency for venous thrombosis (DVT with or without PE)
Relates to an increase risk of venous thrombosis and may be asymptomatic
Can occur as a result of genetic factors, acquired changes in the clotting mechanism or more commonly an interaction between genetic and acquired factors
First described deficiency of anti-thrombin as a risk factor for thrombosis
Deficiencies of protein C and Protein S have since been found to be associated with familial thrombosis
What is Heritable thrombophila
Heritable thrombophilia is an inherited tendency for venous thrombosis (DVT with or without PE)
Relates to an increase risk of venous thrombosis and may be asymptomatic
What can cause thrombophilia
Can occur as a result of genetic factors, acquired changes in the clotting mechanism or more commonly an interaction between genetic and acquired factors
What deficiencies are associated with thrombophilia
First described deficiency of anti-thrombin as a risk factor for thrombosis
Deficiencies of protein C and Protein S have since been found to be associated with familial thrombosis
What initiates thrombosis
Rupture of the plaque, exposing material to subendothelium in the blood
What does thrombosis cause
Platelet plasma coagulation factor activation which results in fibrin formation
The end result is a thrombus that can obstruct the artery or an embolus breaks off and lodges in the heart or brain, causing tissue death
What are the risk factors for thrombosis
Hypercholesterolemia
Hypertension
Smoking
Physical inactivity
Obesity
Diabetes
Inflammatory processes related to artherosclerosis
What is deep vein thrombosis
A blood clot that forms in a deep vein of the leg or pelvis either partially or totally blocking the flow of blood
What is pulmonary embolism
Deep vein thrombosis (blood clot) or part of it, breaks off from the vein
The break away clot travels through the bloodstream to the heart and migrates towards the lung
The clot blocks a vessel in the lung interupting blood supply
What is venous thrombosis
Occurs when activation of blood coagulation exceeds ability of the anticoagulant/inhibitors and fibrinolytic system to prevent the formation of fibrin
Post thrombotic syndrome can result in significant morbidity
Venous thrombosis is a multi-causal disease i.e. gene-environment interactions
Case fatality ranges from 1 to 5%, incidence and fatality are age dependent
How frequent is venous thrombosis
1.17 per 1000 per year
How frequent is DVT
0.48 per 1000 population/year
How frequent is pulmonary embolism
0.69 per 1000 population/year
Write about DVT
Unlikely to cause acute complications
Above knee DVT could cause PE within the lungs, life threatening>shortness of breath and chest pain
What causes thrombosis
Inherited defects
Acquired factors
Multiple defects
What are the eight main risk factors for thrombosis
Atherosclerosis
Acquired thrombophilia
Surgery trauma
Estrogens
Malignancy
Inflammation
Immobility
Hereditary thrombophilia
What are the five strongest risk factors for thrombosis
Hip or leg fracture
Hip or knee replacement
Major General surgery
Major trauma
Spinal Cord Injury
What are the inherited anticoagulant protein deficiencies associated with venous thrombosis?
Antithrombin
Protein C
Protein S
What are the genetic defects associated with venous thrombosis?
Factor V Leiden
Prothrombin gene mutation
How does the clot form in VTE
(5)
Local venous stasis is necessary
Increased turbulence of blood around valves
Endothelial damage causing platelet activation
Localised trapped activated coagulation factors
Low shear rates of blood flow
What causes vascular wall injury
Trauma or surgery
Venepuncture
Chemical irritation
Heart valve disease or replacement
Artherosclerosis
Indwelling catheters
What causes circulatory stasis
Atrial fibrillation
Left ventricular dysnfuction
Immobility or paralysis
Venous insufficiency or varicose veins
Venous obstruction from tumour, obesity or pregnancy
Write about antithrombin
Serine protease inhibitor produced in the liver
Most potent serine protease in coagulation
Responsible for 80% of inhibition that takes place
It’s primary targets are FIIa, FXa, FIXa (FXIa and TF-FVIIa)
It inhibits free FIIa and FXa more easily than that bound in complexes: acts as a scavenger
Antithrombin forms a stable covalent complex with its substrate is then rapidly cleared from the circulation
It’s rate of activity is enhanced 1000x by heparin
Write about antithrombin deficiency
More thrombogenic that PC or PS deficiency
Family studies suggest that AT deficiency is more severe than PC or PS
Majority of patients experience thrombosis before age 25 years
More than 250 mutations have been reported. Homozygosity is rare
Write about thrombosis before age 25 years
Slightly lower in pre-menopausal women than in men of similar age
Lower in COCP use than in non COCP use
Low levels in patients on heparin and in patients with current thrombosis
Significant decreases in DIC, liver diseases, nephrotic syndrome
What are the three different antithrombin deficiency types
Type I
Type II
Type III
Write about Type 1 antithrombin deficiency
Low antithrombin functional activity
Quantitative reduction in antigen and function
Major gene deletions and point mutations
Write about type II antithrombin deficiency
Reduced antithrombin functional activity
Qualitative defect with abnormal AT protein
Reactive site defect - reduced ability to inhibit thrombin or FXa with or without heparin
Heparin binding site defect - reduced ability to bind and be activated by heparin
Pleiotropic effect where mutations produce multiple effects on the structure-function relationship of the molecule
Results from single base substitutions in the coding regions
Write about antithrombin deficiency treatment
Warfarin
Low molecular weight heparin or unfractionated heparin
In some cases antithrombin concentrates and LMWH therapy may be considered
LMWH therapy with regulated dose of warfarin
INR aim 2.0-3.0
Aim is to initially get antithrombin activity over 120% and then maintain activity at 80%
Write about the protein C pathway
The protein C pathway regulates coagulation on phospholipid surfaces
It limits the procoagulant activity of FVa and FVIIa
What are the components of the protein C pathway
Protein C (PC)
Activated protein C (APC)
Protein S (PS)
Thrombomodulin (TM)
Thrombin (FIIa)
Endothelial protein C receptor (EPCR)
How does increased generation of thrombin affect the protein C pathway
Increased generation of thrombin results in increased affinity for thrombomodulin on the endothelial cell surface
FIIa-TM complex prevents binding of FIIa to procoagulant substrates and activated platelets
FIIa-TM complex alters substrate specificity of FIIa allowing activation of PC
TM bound FIIa increases its susceptibility to inhibition by AT
What activates protein C pathway
Activated by the FIIa-TM complex on the surface of the endothelial cells
The inhibitory effects of Protein C are facilitated by the cofactor Protein S
Write about protein C deficiency
(5)
Autosomal dominant inheritence
Relative risk for thrombosis is 10 fold
Type 1 and Type II, Type 1 is more common
Reduced level in Warfarin therapy, DIC, liver disease
Adult heterozygous protein C deficiency patient normally have 60% activity
Write about type I protein C deficiency
Quantitative defect
Characterised by the parallel reductions of functional and immunological protein C
Write about type II protein C deficiency
Qualitative defeect
Functional level is significantly lower than the immunological protein C level
How is protein C deficiency treated
(4)
LMWH or heparin
Once the therapeutic anticoagulation has been achieved with these patients, warfarin can be introduced
Target INR of 3.5 (2.0-3.0)
Side effects of high doses of warfarin treatment is warfarin necrosis
Write about protein S
Vitamin K dependent glycoprotein produced in liver, endothelial cells and megakaryocytes
PS is a non-enzymatic cofactor for APC-mediated inactivation of FVa and FVIIIa
60% of the total plasma PS is complexed with C4b-binding protein (C4bBP) and has no cofactor activity, 40% is free PS remains uncomplexed and is the active moiety
The bioavailability of PS is linked to the concentration of C4bBP and acts as an important regulatory protein in the APC pathway
Write about the action of protein S
(4)
Free PS binds strongly to negatively charged phospholipids on the surface of activated platelets
It forms a Ca++ dependent complex with APC
APC/PS complex will inactivate FVa
Regulation of FVIIIa requires APC/FV/PS
What are the three types of protein S deficiency
Type I, II, III
It has been suggested that type I and type III defects are phenotypic variants of the same genetic disorder
Write about type I Protein S deficiency
(3)
Quantitative deficiency resulting in reduced production of structurally normal protein
Deficiency in total and free PS
Variety of mutations
Write about type II deficiency
Qualitative defect
Missense mutations
Write about type III deficiency
Qualitative deficiency with reduced free PS antigen and normal total PS antigen
Missense mutations
Write about protein S deficiency in women
Lower levels in women
Separate reference range for male/female
Level falls progressively during pregnancy
Reduced levels in women using estrogen containing OCP or HRT
What might reduce protein S levels
(4)
Warfarin therapy
Antiphospholipid syndrome
DIC
Liver disease
Write about factor V Leiden and APCR
(4)
The gene for FV is on chromosome 1
FVa is required for the activation of FII by FXa in the prothrombinase complex
Factor Va is cleaved and inactivated by APC and PS to produce FVi
A mutation in gene for F5 results in production of factor V that is resistant to cleavage by APC
What is APCR
APCR is defined as an impaired plasma anticoagulant response to APC added in vitro
Write about APCR and the FVL mutation
APC resistance co-segregated with thrombosis in families with familial VTE
Mutant FV Leiden has normal procoagulant activity but impaired response to APC
The mutation was first identified in 1994
In the FV gene there is a G (guanine) to A (adenine) base substitution
This results in the amino acid exchange from Arginine to Glutamine at position 506 in the factor V protein
Write about the FVL mutation FVR 506Q
5% of Caucasians
More common in Northern Europeans than in south
15% of patients presenting with first VTE episode
Heterozygous carriers 3-8 fold increased risk (90-95%)
Homozygous carriers 80 fold increased risk (5-10%)
Acquired thrombosis risk factors
The effect of factor V Leiden is strongly enhanced by use or oral contraceptives
What are some acquired thrombosis risk factors
Smoking
OCP
Recent Surgery
What strongly enhances factor V Leiden
Oral contraceptives
Write about the prothrombin gene
Chromosome 11
Vitamin K dependent protein produced in the liver
Guanine to adenine base substitution at base 20210 of the prothrombin gene
Elevated plasma prothrombin levels and increased risk of venous thrombosis
Write about mutations in the prothrombin gene
Prevalence in northern europe is 3% and in 4% of patients with first VTE episode
Relative risk is 2 fold in heterozygotes. Risk for homozygotes is unkown although asymptomatic homozygotes have been described (mild defect)
15-40% of patients with heterozygous FVL are also heterozygous for prothrombin mutation
What are the five main BSH guidelines
Results don’t predict likelihood of recurrence of thrombotic episode
Indicated for patients with first time thrombotic episode under the age of 40
Family history of unprovoked thrombotic events
Indicated for patients with three or more early pregnancy loses
Testing should only be carried out when testing is going to influence treatmnet
When should you not do a thrombophilia screen?
(4)
Reason for thrombosis known such as central venous catheter
Upper Limb Thrombosis
Hospital acquired thrombosis
Retinal vein occlusion
What are three acquired venous thromboembolism causes
Environmental
Iatrogenic
Disease related
What four environmental factors might caused venous thromboembolism
Age
Pregnancy and post-partum
Immobility
Dehydration
What three iatrogenic factors can cause acquired venous thromboembolism
Postoperative immobilisation
Indwelling venous devices
Pharmacological (COCP, HRT, Tamoxifen, chemotherapy)
What four fisease related factors might cause acquired venous thromboembolism
Antiphospholipid syndrome
Malignancy and inflammatory states
Intravenous drug users
Thrombotic thrombocytopenic purpurra
What are some other names for antiphospholipid syndrome
Hughes Syndrome
Sticky Blood Syndrome
What is antiphospholipid syndrome
Acquired autoimmune clinical syndrome with features of thrombosis (venous, arterial and micro-vascular) and/or pregnancy complications and failure
What can antiphospholipid syndrome cause
Venonus thrombosis -> lower limb DVT +/- PE
Arterial thrombosis -> cerebral vasculature: TIA, stroke
Microvascular thrombosis is least common but can present at the potential lethal catastrophic antiphospholipid syndrome
What are the two types of APS
Primary APS
Secondary APS
What is primary APS
Arterial occlusion
Venous thrombosis
Recurrent miscarriage
Sterile endocarditis
What is secondary APS
SLE
RA
SS
Temporal arthritis
What are antiphospholipid antibodies
A heterogenous family of antibodies that react with proteins which bind to negatively charged phospholipids
B2 - GPI most reactive
How do we investigate PAS
Persistent laboratory detection of antibodies, 12 weeks apart
Rules out the possibility of a transient positivity of antiphospholipid antibodies induced by infections or drugs
What antibodies are used in the investigation of APS
Antibodies directed against proteins that have the property of binding to negative charged phospholipids
Lupus Anticoagulant (LA)
Anticardiolipin antibody (aCL) (IgG, IgM)
Anti-B2-Glycoprotein I antibody
What is lupus anticoagulant
Recognised as a strong risk factor for thromboembolic events and pregnancy morbidity
LA tests are sensitive to antibodies to B2GPI and Prothrombin
What does the presence of anti-phospholipid antibody mean
They slow clot formation prolonging clotting time, impairment of the assembly of the prothrombinase complex
How should you select patients for APS testing
Do not test randomly on patients, patient should have significant probability of having APS, Unexplained prolonged APTT
Low, moderate or high probability
What would put someone at low risk for APS
VTE, arterial thrombosis in older patient
What would put someone at moderate risk of APS
Abnormal APTT in asymptomatic patient
Recurrent early pregnancy loss
Provoked VTE
What would put someone at risk for APS testing
Unprovoked VTE
Unusual thrombosis
Late pregnancy loss
Thrombosis in patients with auto-immune diseae
What are anticardiolipin antibodies
The most sensitive test for APS diagnosis
ELISA test
Test is not influenced by warfarin and heparins
Low, moderate or high aCL
What are anti-B2-glycoprotein-I antibodies
Detected using ELISA and are included in the classification criteria
There is no standardisation for this methodology
What are some diagnostic criteria for lupus anticoagulant
Prolongation of phospholipid dependent coagulation test
Demonstration of an inhibitor
Confirmation of phospholipid dependent nature of the inhibitor
Perform 2 screening tests based on different principles, no single test is sufficiently sensitive to LA
Risk of false positives if more than 2 tests used
Some methods are unsuitable for testing when the patient is on warfarin or heparin
Guidelines issued by ISTH, BSH and CLSI
What are two tests used to detect lupus anticoagulant
Dilute Russell viper venom time
Sensitive APTT with reduced phospholipid and silica as activator
Write about the lupus anticaogulant screen
Positive DRVVT ratio is > 1.2
Russell’s Viper Venom activates factor X leading to a clot formation in the presence of FV, prothrombin, phospholipid and Ca++
LA prolongs the DRVVT by binding to the phospholipid and preventing the action of RVV
DRVVT is measured in presence of low dose and high dose phospholipid
The final result is expressed as a ratio of both clot times
The excess phospholipid will neutralise the LA and normalise the DRVVT
This gives the tests it’s specificity
Write about the use of APTT with Sillica as the activator
Use reagent with low concentration of phospholipid
Use a combination of two reagents
- Low concentration of phospholipid
- High concentration of phospholipid
Calculate ratio of Silica time of patient results to normal plasma for both reagents and then overall ratio for both phospholipid reagent
Improve specificity for LA
A ratio < 1.16 is normal
What is needed for the laboratory diagnosis of APS
DRVVT positive
SCT postitive
Interpretation - Lupus Anticoagulant detected
In order to fulfil laboratory criteria for APS a positive result on two occasions more than 12 weeks apart must be obtained
What is needed for diagnosis of acquired thrombophilia
Increased factor VIII levels
Hyperhomocysteinaemia attributable to non genetic causes
Elevated fibrinogen levels
Fulfils at least one clinical and one laboratory criteria
What are some criteria for diagnosis of APS
LA: persistence in positivty on two or more occasions at least 12 weeks apart
Coagulation test:
- Lupus anticoagulant
Immunology test:
- Anti-Cardiolipin antibody
- B-GPI antibody
