Lecture 20 Thrombotic Conditions and Laboratory Assessment Flashcards
What is a thrombus?
Thrombus is an abnormal formation of a platelet or fibrin clot obstruction of a blood vessel
What are some causes of a thrombus?
Many causes:
- Circulatory stasis
- Abnormality in the coagulation system & coagulation control mechanisms
- Abnormal platelet function
- Blood vessel wall disruption
- Leukocyte activation
Where does Venous Thromboembolic Disease (VTE) commonly occur and what are the symptoms?
- Most commonly occurs in the veins of the calves and upper legs
- Symptoms are localized pain, sensation of heat, redness and edema
What more serious complications can occur in Venous Thromboembolic Disease (VTE)?
- Fragments of the clots (thrombi) may separate and move quickly to chambers of the heart, the brain and lodge in the lung called pulmonary emboli (PE)
- These blockages cause ischemia (decreased blood flow)and necrosis of tissue behind the block causing heart attack, stroke, and impaired breathing
10% to 15% of PE patients die within 3 mos.
What is Arterial Thrombosis?
Arthrosclerotic plaques caused by accumulation of lipoproteins mainly LDL
Describe the mechanism/process by which Arterial Thrombosis occurs?
- Activated platelets, monocytes and macrophages embed fatty plaques suppress nitric oxide a vascular relaxing factor
- Small plaques rupture occluding arteries and exposing collagen and releasing Tissue factor
- Platelets binding to vWF will adhere to collagen, then aggregate forming platelet plug
- Tissue Factor combined with Factor VIIa will initiate the coagulation cascade causing a fibrin clot
What patients are frequently tested for Hypercoaguable states?
1) Venous or arterial thrombosis.
2) Transient ischemic attacks (TIA’s)
3) Peripheral Vascular Disease
4) Personal or family history of thrombotic events.
5) Prior to surgery, pregnancy, oral contraceptive use, estrogen therapy, Decreased antithrombin and people with personal or family history of thrombosis.
What is a natural anticoagulant in the body? What factors does it affect?
Antithrombin: Natural anticoagulant to activated forms of thrombin, Factors IX, X, XI and XII. Most important factors inhibited are IIa and Xa.
What defects and disease are associated with hereditary deficiencies of antithrombin?
Hereditary deficiencies both quantitative and qualitative. D.I.C and Liver Disease also causes low levels
What therapy can deplete levels of antithrombin if not monitored properly?
Heparin therapy over a sustained period can deplete AT causing increased risk of thrombosis.
How is antithrombin typically measured?
Usually a chromogenic assay: Amount of chromophore released is proportional to AT concentration.
What can give rise to heparin resistance?
Low antithrombin level can give rise to heparin resistance.
Note: Without adequate amounts of Antithrombin Heparin cannot work properly as an anticoagulant. If patients are not responding to Heparin therapy an Antithrombin level assay should be performed to rule out Antithrombin deficiency as the cause.
What does Antithrombin require for effective anticoagulant activity?
Aka Heparin co-factor
Most important coagulation inhibitor
In vivo heparin sulfate (from endothelial cells and platelets) + commercial heparin
↑ AT activity
Need to confirm what “Aka Heparin …” means.
Where is antithrombin made?
Synthesized in the liver
What factors does anti-thrombin inhibit?
Naturally inhibits coagulation by forming complexes with factors II, IX, X, XI, XII therefore neutralizing them
See slide 15 for a list of all of AT’s functions.
In what conditions is anti-thrombin deficient in?
Deficiencies in AT such as those conditions listed below can lead to severe, even fatal thrombotic events:
- Hereditary seen in 1:2000 to 1:5000 of the general population
- DIC
- Liver disease
- Nephrotic syndrome
What are Protein C and S?
Protein S is a cofactor to Protein C
When complexed, Protein C and S are potent inhibitors of Va and VIIIa
What is the impact of even an heterozygous deficiency in either S or C proteins?
Heterozygous deficiencies lead to a 2-10 fold increased risk of thrombosis
Deficiencies result in serious thrombotic disorders.
What vitamin is protein C and S dependent on?
Vitamin K dependent (like II, VII, IX, and X)
How does protein C and S degrade Va and VIIIa?
- Thrombin binds to endothelial receptor thrombomodulin
- Thrombin + thrombomodulin cleave Protein C to form activated Protein C (APC)
- Protein S complexes to activated Protein C (APC) which degrade Va and VIIIa
What is Factor V Leiden?
Factor V Leiden is a genetic mutation seen in 3-8% of Northern European Caucausians causing the Factor V molecule to be resistant to Activated Protein C causing a 3-18 fold increased risk of thrombosis.
What does the Activated Protein C Resistance test do?
Activated Protein C Resistance: test for Factor V Leiden Mutation which provides resistance of Factor V to Activated Protein C. Abnormalities should be confirmed by DNA testing.
What is looked at in Factor V Leiden DNA testing?
Factor V Leiden DNA testing: describes as single point mutation which provides the resistance to APC.
● DNA testing important to confirm heterozygous or homozygous state.
What test discovers a gene defect that leads to increased amounts of Prothrombin and increased risk of thrombosis?
Prothrombin Mutation Test (G20210A)
● Is also a gene defect which results to increased amounts of Prothrombin and increased risk of thrombosis.
● Presence detected by DNA testing.
How does increased levels of Homocysteine increase risk of thrombosis?
Hyperhomocysteinemia Prothombotic effects:
● Decrease of endothelial cell Tissue Plasminogen Activator (TPA) binding sites.
● Activation of Factor VIIa and V
● Inhibition of protein C and heparin sulfate
● Increased prothrombin fragments
● Increased blood viscosity
● Decreased antithrombotic activity due to changes in thrombomodulin function
How does oral contraceptives increase risk of thrombosis?
- Due to acquired resistance to activated Protein C
- Increase in factors II and VII and fibrinogen
Note: Increase risk especially in people with other conditions such as hereditary defects; i.e. Factor V Leiden, Protein C and S deficiencies etc.
What is the name of a group of inhibitors that bind with protein-phospholipid complexes and sometimes called non-specific inhibitors?
Lupus Inhibitors (anticoagulant):
Lupus inhibitors / antibodies produced in response to viral infections, drugs, tumor necrosis factor, autoimmune inflammatory disorders are a group of immunoglobulins which bind to a variety of protein-phospholipid complexes and are sometimes called non-specific inhibitors
These antibodies alter the phospholipid surface as described above.
What does Lupus Inhibitors (anticoagulant) inhibit?
- Inhibition of fibrinolytic system (factors XII and prekallikrein)
- Inhibition of prostacyclin release from enthothelial cells. (inhibitor of platelet aggregation)
- Inhibition of Protein C and S systems.
- Inhibition of Annexin V (significant role in the down regulation of extrinsic (tissue) pathway
- Inhibition of beta-2-glycoprotein I (acts with protein S)
Note: Scientist continue to investigate how lupus inhibitors cause thrombosis
What are the effect of Lupus inhibitors on coagulation tests?
Lupus inhibitors cause an elevation in coagulation tests due to interference with protein/phospholipid reactions in the coagulation cascade .
What is the purpose of Antiphospholipid Testing and what condition is it found in?
Demonstrates presence of antibodies to phospholipid (negatively charged lipids), commonly found in System Lupus Erythematosis (SLE).
What kind of antibodies can be found in Antiphospholipid Testing?
Antibodies can be IgM, IgG, IgA
What conditions are associated with positive Antiphospholipid Tests?
Positive tests are strongly linked to conditions of venous, arterial thrombosis, recurrent fetal loss, thrombocytopenia, cerebral infarction.
What is Heparin Co-Factor II?
Heparin Co-Factor II
Inhibits thrombin
Less affinity and reacts more slowly than AT
What does Alpha 2 Macroglobulin inhibit?
Alpha 2 Macroglobulin
Inhibits thrombin and Kallikrein
Slower than AT
What factor does Alpha 1 Antitrypsin inhibit?
Alpha 1 Antitrypsin:
Alpha globulin inhibits FXIa
Inactivates thrombin at a weak rate
What is the main inactivator of XIIa, XIa, and Kallikrien contact factors?
C1 inactivator
Main inactivator of XIIa, Xia, and Kallikrien contact factors
What is the principal regulator of the tissue factor pathway? Where is it synthesized?
Tissue Factor Pathway Inhibitor (TFPI):
Synthesized by endothelial cells
Principle regulator of the tissue factor pathway
Remember . . . . in vivo TF:VIIa activates IX and X
How does TFPI prevent activation of X and IX?
TFPI binds and inactivates Xa
TFPI:Xa complex will then bind to TF:VIIa and prevent activation of X and IX
List three anticoagulant therapy drugs and what they inhibit?
- Aspirin
Inhibits platelet aggregation - Heparin (unfractionated)
Inhibits factors II, IX, X, XI, XII - Warfarin
Inhibits factors II, VII, IX, X
How does aspirin work as an anticoagulant to treat thrombotic events? How long does it last?
Aspirin Anticoagulant Therapy
- Aspirin inactiviates cyclooxygenase
Blocking thromboxane A2 production - Interferes with platelet aggregation
- Irreversible - Lasts the rest of the platelets life, ~7-14 days
Can be used to treat thrombotic events
What does aspirin prolong?
Most common drug associated with prolonged Bleeding Time.
What does Heparin Therapy (Unfractionated) inhibit?
Inhibits intrinsic and common factors
1. Thrombin
2. IX
3. X
4. XI
5. XII
How can you measure heparin therapeutic range? What is the range?
Measured by APTT Test
Therapeutic range: 0.2 – 0.4 units/mL Unfractionated Heparin
What does heparin enhance?
Anti-thrombin enhancer
1. Heparin’s action on these factors (X11a, X1a, IXa, Xa, VIIa, Va, VIIIa, Thrombin) is mediated through the action of anti-thrombin (speeds up AT)
2. Exposes more of its active sites so they can bind to the activated coagulation factors more efficiently and render them ineffective.
3. Heparin increases ATIII effect several thousand times fold !!! ( 2 – 10,000 times fold).
See slide 31 for diagram.
What is heparin therapy the choice treatment of?
Choice for immediate treatment of:
1. Acute venous thrombosis
2. Pulmonary embolism (PE)
How is heparin therapy administered?
Administered:
1. Intermittent bolus (concentrated) injection IV every 4-6 hours
2. Continuous IV infusion
3. Subcutaneous injection every 12 hours
Dose too ↑ = bleeding; dose too ↓ = thrombosis
What can cause heparin resistance?
Heparin Resistance
Can be due to ↓ levels of anti-thrombin (<50%) and ↑ levels of stress factors I, V and VIII.
Note: May require higher doses of Heparin to overcome their pro-coagulant effect
What is the goal of the heparin anticoagulant therapeutic range?
Therapeutic range to provide proper anticoagulant effect and prevent bleeding should be established for each lab using present heparin stock
What test is used to establish the therapeutic range for heaprin?
Test used to establish therapeutic range is heparin response curve.
APTT times are performed on concentrations of heparinized plasma from 0.0 to 0.5 units of heparin/ml of plasma.
Therapeutic range of Heparin Response curve is APTT times typically between 0.2 to 0.4 units/ml.
How can too high of dose of heparin be reversed?
Too high doses of heparin can be reversed by administration of Protamine Sulfate
What occurs in heparin induced thrombocytopenia (HIT)?
Heparin Induced Thrombocytopenia (HIT)
HIT: condition where antibodies are produced against the heparin/platelet factor 4 complex on the surface of the platelet. This leads to aggregation of platelets, thrombocytopenia with arterial and venous thrombosis.
What blood count should be performed prior to and during heparin therapy?
Platelet count should be performed prior to and during heparin therapy to detect Heparin Induced Thrombocytopenia (HIT).
How does Heparin Induced Thrombocytopenia (HIT) lead to thrombocytopenia?
Aggregates cleared by spleen
Leading to thrombocytopenia
Note: Patients with HIT will require an alternate anticoagulant therapy
What is Low Molecular Weight Heparin?
- Produced from unfractionated heparin by chemical and enzymatic depolymerization = reduced size
- Reduced activity to bind thrombin but increased ability to bind Xa
- Effective antithrombotic agent especially in deep vein thrombosis (DVT).
What are the advantages of LMWH over standard heparin?
- Does not interact with platelets so less chance of bleeding.
- LMWH shows little or no binding to plasma proteins providing greater bioavailability and more predictable clearance from the circulation than standard heparin.
- Lower incidence of HIT. As a result laboratory monitoring is not necessary.
- LMWH is the heparin of choice for out-patients because it can be given once or twice per day.
If LMWH monitoring is necessary what assay should be performed?
In rare cases where LMWH monitoring is necessary it should be done with anti-Xa heparin assay, not the APTT.
What is Warfarin?
Oral anticoagulant (can be given to people not requiring hospitalization).
Used most commonly for treatment of venous thrombosis
Brand name: Coumadin
May also hear it as Coumarin
When can Warfarin be given as an oral anticoagulant?
Warfarin should only be started when therapeutic range of heparin is achieved.
- Heparin is faster acting
- Stable warfarin anticoagulant effect is not achieved for several days (36-72hrs) to (72-96hrs)
- Heparin is discontinued once stable anticoagulant activity from warfarin is achieved. There is usually a ≈5-7 day overlap.
What does Warfarin interfere with and how do you reverse it?
Interferes with Vitamin K metabolism
Reverse affects of warfarin by administering Vitamin K and Fresh Frozen Plasma.
What vitamin K dependent proteins does Warfarin interfere with?
Vitamin K dependent proteins
C – inhibits Va and VIIIa
S – cofactor of C
Z – cofactor to ZPI which inhibits Factor Xa
What is Warfarin’s effect of the rate of reduction and why is this important?
Warfarin’s effect on the rate of reduction is related to the metabolic half life of the affected factors. Protein C half-life is shorter than the half-life of any of the other factors that Warfarin inhibits. This is important because in the initial stages of Warfarin therapy patients are actually at an increased risk of thrombosis hence the need for a Heparin-Warfarin overlap period.
Note:
Factor VII: short half-life, decreases first 5-6 hrs
Protein C: shorter half-life 4-6hrs (inhibits Factor Va and Factor VIIIa)
Protein C decreases first, so patient is at greatest risk of thrombosis at the beginning of warfarin therapy
Factor IX: 28-40 hrs
Factor X: 40-50 hrs
Factor II: 48-60 hrs.
How long does it take for stable warfarin anticoagulation to be achieved?
Stable warfarin anticoagulant effect is not achieved for several days (36-72hrs) (72-96hrs)
