intro to hemostasis Flashcards
Its all about balance!
Primary and secondary hemostasis and fibrinolysis need to occur tightly regulated, coordinated fashion to ensure appropriate clot formation and dissolution (clotting vs bleeding)
normal hemostasis: clotting
divided into 2 categories:
Primary: vasculature, blood flow, platelet count and function, extracellular matrix proteins–> PLATELET PLUG
Secondary: coagulation factors–> FIBRIN CLOT
Initial steps to vasculature injury
Arteriolar vasoconstiction: reduces blood flow, mediated by endothelin
Exposure of subendothelial von Willebrand factor (vWf) and collagen
Primary hemostasis
- platelet adhesion (binding of platelet glycoprotein 1b (Gp1b), to vWF and exposed collagen
- Platelet activation (shape change, granule release). Shape change- increases their surface area, move negatively charged phospholipids to the surface (for binding Ca++), conformationa gange in Gp2b/3a. Granule release- alpha granules (fibrinogin, clotting factors), Dense granules (ADP, ATP, Ca++), Stimulate platelet activation–>
recruitment - Recruitment
- Aggregation-Gp2b/3a binds fibrinogen, forming bridges between platelets. Thrombin stimulates (irreversible platelet contraction, conversion of fibrinogen to insoluble fibrin)
Secondary hemostasis
goal is to strengthen the fibrin glue via generation of THROMBIN (factor 2 a) 1. tissue factor release 2. phospholipid complex expression 3. Thrombin activation 4. Fibrin polymerization REPEAT
Clotting in vivo
Vascular damage, exposes tissue factor, tissue factor Activates Factor 7 (7a)
Factor 7a stimulates Factor 9 (9a)
(9a) and Factor 8a Stimulates Factor 10
10 a and 5 a activate factor 2 (Prothrombin) to 2a (thrombin
2a catalyzes fibrinogen to fibrin
creating a Fibrin clot
Thrombin feeds back and creates positive feedback to keep the process going
Clotting factors
Produced in the liver hepatocytes (factor 8 is the exception, made by endothelial cells)
Clotting factors circulate in inactive forms, but are functional (Active forms are produced during the cascade)
Some are vitamin K dependent: 2, 7 9, 10, Protein C and Protein S (anticoagulants) carboxylated with a negative charge, Vit K is antagonized by warfarin
charge and secondary hemostasis
negatively charged clotting factors (2 7 9 10) also the vitamin k dependent clotting factors
Localized to the Ca which has been attracted to the negatively charged phospholipid surface of the platelets
Along with the clot hemostasis what else is happening concurrently
t-PA is being released- fibrinolysis (digest fibrinogen)
Thrombomodulin (blocks coagulation cascade)
Fibrinolysis is always occuring during hemostasis to keep the clot at bay
Limits the size of the thrombus and dissolution
tPA (endothelium) activates plasminogen into plasmin
Plasmin breaks up fibrin, producing fibrin degradation products (D dimers)
alpha antiplasmin inactivates free plasmin
Normal hemostasis: natural anticoagulants
Endothelial cell activation and release of mediators (PGi2, NO, ADP) which inhibits platelets
Activation of thrombomodulin, which leads to activation of protein C and S which inactivate F5a, F8a
Tissue factor pathway inhibitor (TFPI) released, inactivating F7a
Heparin-like molecules activated antithrombin 3, (AT3), which inactivates f2a, and others
Laboratory screening for primary hemostasis vs Secondary hemostasis
Primary hemostasis: platelet count and morphology, bleeding time, platelet function (PFA100, Platelet aggregation studies), vWillebrand studies (vW Ag, vW activity), fibrinolysin: D imer is free and means plasmin has occured
Secondary Hemostasis: Activated clotting time (ACT), Prothrombin time (PT), Activated partial thromboplastin time (aPTT), fibrinogen activity, thrombin time
Morphology of hemostasis
Normal platelets (variably sized, smaller than RBC, granulated)
Abnormal (hypogranular- MDS, grey platelet syndrome), Giant (MPNs, bernard soullier disorder, MHY9 disorders)
bleeding time
vascular integrity, platelet count and function. time when the pt stops bleeding, not really performed. 0-13 minutes
Platelet function assay
Gold standard=platelet aggregation study
You add agonist to platelets, measures PLT aggregation, manual, labor intensive (allows light to go through)
Screening: automated and fast, PFA100 test (screens for PLT functional defects and vonWillebrand disease, high sensitive for severe PLT function defect/ VWD, moderate sensitivity for mild to moderate disorders. False positives common, moderate sensitiveity to aspirin therapy
Antiplatelet medication monitoring: automated/fast, Verify Now, aspirin, clopidogredl responsiveness
Clotting/ secondary hemostasis in vivo vs in the lab differences
In the lab clotting is divided up in the extrinsic pathway, intrinsic pathway and the common pathway
Extrinsic pathway: TF activates 7, 7a activates 10
Intrinsic pathway: a reaggent stimulates 12, 12 activates 11, 11 activates 9, 9 along with 8a activates 10.
Common path:10 with 5a activates prothrombin into thrombin
Thrombin activates fibrinogen to fibrin
Prothrombin time (PT)
combine tissue factor, lots of phopholipid and calcium
Tests the extrinsic pathway and the common pathway
F7, F10, F2, F5, fibrinogen
Uses: screening for factor deficiencies, Warfarin monitoring
Reported in seconds and INR (normalizes across labs), not prolonged with heparin
Acitvated partial thromboplastin time (aPTT)
combines activator, low phospholipid, calcium
Tests the INTRINSIC and common pathway (F12, 11, 10, 9, 8, 2, 5, fibrinogen
Uses: screen for factor deficiencies, lupus anticoagulant, heparin monitoring
variably sensitive to warfarin
mixing studies
can be done with PT or aPTT,
Mix 1 part of the patient sample with one part of normal plasma
then run the PT or aPTT
Corrects with factor deficiencies
Fails to correct with inhibitors (drugs, lupus anticoagulant)
labs for fibrinolysis
D- dimers
Measurable breakdown product of plasmin-induced fibrinolysis
Immunoassay
thrombin time (TT)
add thrombin, measure time to form clot, abnormal/prolonged
Fibrinogen –> fibrn formation yes or no
Heparin will elongate, direct thrombin inhibitors
A/hypofibrinogenemia
Dysfibrinogenemia
Fibrinogen activity
Reflects fibrinogen activity and concentration, reported in mg/dL, abnormal (low: ahypofibrinogenemia, high: inflammatory state- acute phase reactant)
Bleeding disorders: defects in primary hemostasis
Skin, mucosal membrane hemorrhages (petechiae, epistaxis, menorrhagia) are common
Vascular abnormalities (ehlers- danlos)
thrombocytopenia
Platelet function disorders
Von willebrand disease
bleeding disorders:Defects in secondary Hemostasis
Defects in secondary Hemostasis
bleed into soft tissues or joints
Clotting factor deficiencies (hemophilia, liver disease)
Exogenous (anticoagulants)
Thrombocytopenia
<100k/uL
differential: decreased bone marrow production, increased destruction (immune mediated, non immune mediated), dilutional, sequestration
Mostly small vessel bleeds but risk of intracranial hemorrgage when platelets <10k
Causes of thrombocytopenia: Decreased PLT production due to decreased Megakaryocytes (aplastic anemia, vit B12/folate deficiency, leukemia/lymphoma, MDS, metastatic carcinoma, alcohol, toxins, drugs, infection
Increased destruction: immune mediated (immune thrombocytopenia purpura, heparin induced thrombocytopenia, transfusion/pregnancy allo immune thrombocytopenia), drugs (quinine, vancomycin)
Non immune mediated: Disseminated intravascular coaulation (DIC), Thrombotic thrombocytopenia purpura, hemopytic uremic syndrome, drugs
immune thrombocytopenia pupura (ITP)
Caused by auto antibodies made against PLT Ags (Gp2b/3A, Gp1B)
Destruction of PLTs occurs by phagocytosis
FC receptor mediated
Site is the spleen
Primary : etiology unknown
Secondary: associated with: lupus, leukemia (CLL/SLL), Drugs, Viruses (HIV hepatitis)
Clinical features: Skin, mucosal bleeds, normal spleen size
Variants: Acute (self limited, < 6 months, post viral, children, most common), Chronic (adults, usually females) persisten, >6 months
PAthologic and diagnisis of immune thrombocytopenia purpura (ITP) and treatment
Pathologic findings: thrombocytopenia (low), large platelets, normal to increased megakaryocytes in the marrow, white pulp expansion
Diagnosis: presumptive, diagnosis of exclusion, no diagnostic tests, normal clotting tests bone marrow
Treatment might not need: esp in kids where its a self limited disease, immunosuppressent to stop the antibody attack of platelets (steroids, IVIG, AntiCD20 Ab), thrombopoietin agonist, splenectomy
other causes of thrombocytopenia
Dilutional (massive transfusion of trauma patients)
Sequestration (seen with hypersplenism, increased storage of PLTs)
Platelet functional disorders
Heterogeneous group of inherited and acquired, qualitative disorders with variable clinical presentation
Bernard soullier (Gp1b deficient, inherited, severe), Glanzmanns thrombasthenia (Gp2b/3a deficiency, inherited, severe), Storage pool disease (granule/transient defects, inherited, variable clinical), Aspirin (COX inhibition, ascquired, variable), Uremia (acquired, variable)
vonWillebrand disease
Most common inherited bleeding disorder
Usually autosomal dominant
Incidence may be underestimated given the mild presentation, rarely can be acquired
Clinically heterogeneous: Mild, revealed with surgery, dental work, trauma
Moderate to severe: menorrhagia, spontaneous bleeding, dependent on clinincal type
What is vonWillebrand factor ?
A protein produced by endothelial cells, megakaryocytes, PLTs
Function: in the circulating Form, it stabilizes F8
in the fixed form: it adheres to collagen and PLTs through Gp1b
Von Willebrand disease types and treatments
Types: (LOW, Weird, NO- 1,2 ,3)
1- quantitative (most common 80%), Autosomal dominant, clincally heterogenous, usually mild
2- qualitative (2A- 10-15%), 2B (<5%), 2N, and 2M (rare). Moderate to severe bleeding, 2N is Autosomal recessive other are usually A/D
3- Complete absence of vWF, rare A/R, severe bleeding disorder, mimicking hemophilia
Treatment:
Desmopressin- stimulates vWF release, factor concentrates, Anti-fibrinolytics, oral contraception
vWD lab evaluation
Screen: CBC (normal Plt coount), PT (normal), aPTT (normal or prolonged), vWF levels, vWf activity (ristocetin), Factor 8 activity
Confirmatory studies: multimer electrophoresis, specific binding assayas, platelet aggregation
Defects in secondary hemostasis, Single clotting factors vs multiple clotting factors
Single clotting factors- hemophilia
Multiple clotting factors- liver disease, vitamin K deficiency, exogenous anticoagulants
hemophilia
Inherited factor deficiencies:
X linked recessive for hemophilia A, B
Mostly males are affected, females most often are only carriers, 30% are sporatic new mutations
Types:
Hemophila A-F8 deficiency, most common inherited disease, associated with life threatening bleeds
Hemophilia B- F9 deficiency
Hemophilia C- F11 deficiency Ar inheritance, not predictable bleeding
Clinical presentation of hemophilia and lab evals (A and B)
Spontaneous bleeds, hemarthrosis–> progressive deformity, deep muscle bleeds, easy bruising with trauma, bleeding tracts with factor levels
Lab evaluation:
APTT prolonged, PT normal
aPTT mixing shows corrections
Factor levels/Activities severe to mild depending on F8 /F9 (<1%, <5%, 50%)
Single factor deficiencies
Defects in factor 12 is asymptomatic (factor 12 is only in lab, not in vivo coag cascade)
Factor 5- topical bovine thrombin, 10 amyloid, 13- poor wound healing and infertility in men
Inhibitors (auto Abs against any clotting factor can develop and cause a deficiency in the setting of Auto immune disorders, medications, and malignancies
Coagulopathy of liver disease
Deficiency of multiple coagulation factors: Procoagulants (factors 2, 5, 7, 9, 10), Anticoagulants (antithrombin 3, protein C and protein S)
Thrombocytopenia: Hypersplenism (in pts with cirrhosis associated splenomegaly), decreased thrombopoietin production
Vitamin K deficiency
Fibrinogen abnormalities- low fibrinogen dysfibrogenemia- dysfunctional protein
Therapy of coagulation disorder- treatment not required if patient not bleeding, if bleeding Vit K replacement, rarely corrects PT, replacement of plasma, cryoprecipitate, platelets, therapy specific for bleed site
Can be seen in acute liver injury (hepatitis, toxins tylenol and alcohol), or can be seen in chronic like cirrhosis
PT, aPTT, TT are going to be prolonged
Low fibrinogen activity, Normalish platelets, low factor activities (except F8 and vwF - made by endothelial cells)
Vitamin K deficiency
Dependent factors 2 7 9 10, Protein C and S
Cofactor fo gamma carboxylation (gamma carboxylation and Ca is required for factor binding to phospholipid), provided by diet and bacterial flora
Vitamin K deficienct Drugs (oral anticoagulant warfarin inhibits clotting factors, antibiotics decrease bowel flora and therefore there is less vitamin K synthesis)
Malabsorpption of dietary deficiency, liver disease, newborns
Treatment: vit K replacement, fresh frozen plasma for emergencies, newborns recieve an injection at birth
Anticoagulants
multiple clotting factors inhibited (Warfarin- fs 2 7 9 10 c and S)
Heparin inhibits 2 9 10, accelerates antithrombin 3 activity
Single clotting factor inhibited (direct thrombin inhibitors (2)
Factor X inhibitors (10)
anticoagulant activity- INR (warfarin), PTT/antiXa assay for heparin, and Ptt (some direct thrombin inhibitors)
virchows triad of thrombosis formation
Endothelial injury, abnormal blood flow, hypercoagulability
Thrombophilia risk factors
a couple of risk factors can be either inherited or acquired (more commonly acquired)
Protein C, S , anti Thrombin 3 deficiencies
Hyperhomocysteinemia
Dysfibrinogenemia
sources of thrombi
Venous- develop at sites of stasis, travel in direction of blood flow, DVT occuring in leg veins, may travel through heart to pulmonary veins (pulmonary embolism), unusual locations (splenic v, cerebral vein, branhial v)
Heart: atrial or ventricular walls or valve leaflets. Right heart (embolus may detach and travel into lung),
Arterial - develop at sites of endothelial injury and turbulence, travel retrograde, atherosclerotiic plaque
Site of thrombi by disease
Factor 5 leiden, prothrombin 20210a mutation and AT 3, protein C and S deficiency will cause a venous thrombosis
Antiphohpholipid antibody syndrom and homocystein will cause either venous or arterial
Factor 5 leiden
essentially a messed up factor 5 disease
Most common inherited predisposition to thrombosis (venous)
Single point mutation in F5 gene G-> cleavage site for protein C (anticoagulant degrades factor 5 and 8),
2-15 % of caucasians are heteros, most patients will not have thrombosis unless coupled with other risk facots
Activated ptotein C resistance=mutated F5 is resistant to protein C inactivation, F5 is active for longer and promotes clotting
So protein C cant degrade factor 5 (because of the messed up cleavage site)
Factor 5 leiden testing
diagnosis: molecular analysis (PCR, microarray) for mutation (cheap and automated
Who should be tested- younger adults whove ever had a venous thrombosis (VT). recurrent VTs, VT famHx, VT in pregnancy or while on OCs, MI in fem smokers under 50
Prothrombin 20210a mutation
2nd most common inherited predisposition for thrombosis (venous), single base pair mutation in prothrombin (F2) gene
Heterozygotes vs homozygotes
Unclear mechonism of hypercoagulability (could be due to elevated ptothrombin levele
Diagnosis: molecular analysis for gene mutation
double heterozygotes
heterozygotes for factor 5 leiden and prothrombin gene mutation
Greater risk of thrombosis
AT3, protein C and Protein S deficiencies
May be inherited or acquired
Associated with venous thrombsis
inherited forms: account for 5- 15% of familial thrombosis risk higher after puberty
Acquired forms (consumptive (recent clot, DIC), decreased synthesis (liver failure or vit K deficiency)
Lab eval: AT3, C and S activities, S free Ag, do not evaluate during active thrombolytic event because these will be abnormally high
Antiphospholipid antibody syndrome
1-5% of gen pop, acquired autoantibodies against various phospholipid complexes
Associated with arterial AND venous thromboses with high recurrence rates and repeat miscarriages (DVTs and chronic pulmonary emboli, stroke, renal failure, interference with placenta growth and devolopment
Primary/unknown, secondary associated with lupus
Inhibits clotting in vitro, promotes clotting in vivo, looks like a blleding disorder in a tube, but in body is a clotting disorder
Antiphospholipid antibody syndrome diagnostic and lab eval
Diagnosis 2 criteria:
Positive lab test (only need 1): lupus anticoagulant test, antiphospholipid antibody ELISA
Postitive clinical findind: thrombosis, recurrent miscarriage
Lab eval: Lupu s anticoagulant: functional assessment, prolongation of clotting based tests
Anti-cariolipin, antiB2 glycoprotein ELISA (anti phospholipid), could be prolonged PTT
Diagnosis of lupus anticoagulant (LA) for antiphospholipid antibodies
Prolongation of 1 clotting assay, sensitive to lupus anticoagulants (dRVVT, aPTT) NOT NOT NOT PT
correction fo clotting time with excess phospholipid
Failure to correct on mixing study
Persistently present (>12 weeks)
Thrombotic states associated with thrombocytopenia
Heparin induced thrombocytopenia (HIT)
Thrombotic microangiopathies: thrombotic thrombocytopenia purpura (TTP), Hemolytic uremic syndrome (HUS), disseminated intravascular coagulation (DIC)
Disseminated intravascular coagulation (DIC)
Heparin induced thrombocytopenia (HIT)
Definition: thrombocytopenia (<150k/ul) or >50 % drop in platelet count during heparin therapy
1-5% of pts on heparin devlop HIT (5-10 days after initiation of heparin, uncommonly seen within 24 hrs)
Most common in unfractionated heparin, CARDAIC>SURGICAL>MEDICAL
Major clinical manifestation: thrombosis!- venous or arterial
Drug antibody production: Abs made against heparin-PF4 complex (IgGs), immune complexes bind to platelets through Fc receptor, Platelets activate and initiate clot formation
Diagnosis: Screen ELISA for PF4-heparin IgG, 95% senstivitve, 80% spec, Confirm - seraotonin release as, 88% 100%
Treatment stop the fucking heparin, non heparinoid anticoagulants (direct thrombin inhibitors, warfarin)
Thrombotic microangiopathies
disease charachterized by wide spread thrombi deposition in microvasculature, hemolytic anemia (MAHA, organ dysfunction, thrombocytopenia due to consumption
Thrombotic thrombocytopenic purpura- the classic thrombotic microangiopathy
Clinical emergency
Classic penta: feverm renal and neurologic impairment, thrombocytopenia, MAHA (micro angiopathic hemolytci anemia)
Acquired»»>congential
30-50 females> males
Deficiency of metalloproteinase ADAMTS (usually autoimmune-anti ADAMTS13 anti bodies), leads to large vWF multimers
Platelet activation and thrombi formation
Diagnosis: look for red cell fragments- schistocytes, decreased platelts, decreased ADAMTS13 levels
Treatment- rapid, plasma exchange- remove antibodies resupply ADAMTS 13
Typical Hemolytic UREMIC syndrome
Associated with shiga toxin producing E coli
Local toxic effects on clonic cells-> pain and bloody diarrhea
Systemic toxin on endothelial cells–> systemic disease
Renal dysfunction, CNS dysfunction may occur
look for the ecoli in the stool, no decrease in ADAMTS 213
Disseminated intravascular coagulation
characterized initially by clotting in the microvasculature, organ ischemia, followed by bleeding tendencies
Consuptive coagulopathy (widespread excessive clotting–> consumption of factors and platelets, clotting factors are consumed–> bleeding, platelets are consumes–> bleeding, activation of fibrinolusis –> bleeding
Associated with sever illnesses: sepsis, trauma, OBGYn, fat embolis, cancer (AML), burns ischemia
Acute or chronic (acute will be bleeding), chronic thrombotic,
end organ ischemia (kidney disease, brain mircoinfarct, hyaline membranes/ARDS, adrenal necrosis and hemorrhage, MI damage
MAHA: red cells lyse as they go through occlusiosn
Prolonged clotting, elevated D dimers, treat the unerlying cause
