intro to hemostasis

Question 1

Its all about balance!

Answer 1

Primary and secondary hemostasis and fibrinolysis need to occur tightly regulated, coordinated fashion to ensure appropriate clot formation and dissolution (clotting vs bleeding)

Question 2

normal hemostasis: clotting

Answer 2

divided into 2 categories:
Primary: vasculature, blood flow, platelet count and function, extracellular matrix proteins–> PLATELET PLUG

Secondary: coagulation factors–> FIBRIN CLOT

Question 3

Initial steps to vasculature injury

Answer 3

Arteriolar vasoconstiction: reduces blood flow, mediated by endothelin

Exposure of subendothelial von Willebrand factor (vWf) and collagen

Question 4

Primary hemostasis

Answer 4

1. platelet adhesion (binding of platelet glycoprotein 1b (Gp1b), to vWF and exposed collagen
2. 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
3. Recruitment
4. Aggregation-Gp2b/3a binds fibrinogen, forming bridges between platelets. Thrombin stimulates (irreversible platelet contraction, conversion of fibrinogen to insoluble fibrin)

Question 5

Secondary hemostasis

Answer 5

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

Question 6

Clotting in vivo

Answer 6

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

Question 7

Clotting factors

Answer 7

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

Question 8

charge and secondary hemostasis

Answer 8

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

Question 9

Along with the clot hemostasis what else is happening concurrently

Answer 9

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

Question 10

Normal hemostasis: natural anticoagulants

Answer 10

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

Question 11

Laboratory screening for primary hemostasis vs Secondary hemostasis

Answer 11

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

Question 12

Morphology of hemostasis

Answer 12

Normal platelets (variably sized, smaller than RBC, granulated)

Abnormal (hypogranular- MDS, grey platelet syndrome), Giant (MPNs, bernard soullier disorder, MHY9 disorders)

Question 13

bleeding time

Answer 13

vascular integrity, platelet count and function. time when the pt stops bleeding, not really performed. 0-13 minutes

Question 14

Platelet function assay

Answer 14

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

Question 15

Clotting/ secondary hemostasis in vivo vs in the lab differences

Answer 15

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

Question 16

Prothrombin time (PT)

Answer 16

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

Question 17

Acitvated partial thromboplastin time (aPTT)

Answer 17

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

Question 18

mixing studies

Answer 18

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)

Question 19

labs for fibrinolysis

Answer 19

D- dimers

Measurable breakdown product of plasmin-induced fibrinolysis
Immunoassay

Question 20

thrombin time (TT)

Answer 20

add thrombin, measure time to form clot, abnormal/prolonged

Fibrinogen –> fibrn formation yes or no

Heparin will elongate, direct thrombin inhibitors

A/hypofibrinogenemia
Dysfibrinogenemia

Question 21

Fibrinogen activity

Answer 21

Reflects fibrinogen activity and concentration, reported in mg/dL, abnormal (low: ahypofibrinogenemia, high: inflammatory state- acute phase reactant)

Question 22

Bleeding disorders: defects in primary hemostasis

Answer 22

Skin, mucosal membrane hemorrhages (petechiae, epistaxis, menorrhagia) are common

Vascular abnormalities (ehlers- danlos)

thrombocytopenia

Platelet function disorders

Von willebrand disease

Question 23

bleeding disorders:Defects in secondary Hemostasis

Answer 23

Defects in secondary Hemostasis
bleed into soft tissues or joints
Clotting factor deficiencies (hemophilia, liver disease)
Exogenous (anticoagulants)

Question 24

Thrombocytopenia

Answer 24

<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

Question 25

immune thrombocytopenia pupura (ITP)

Answer 25

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

Question 26

PAthologic and diagnisis of immune thrombocytopenia purpura (ITP) and treatment

Answer 26

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

Question 27

other causes of thrombocytopenia

Answer 27

Dilutional (massive transfusion of trauma patients) | Sequestration (seen with hypersplenism, increased storage of PLTs)

Question 28

Platelet functional disorders

Answer 28

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)

Question 29

vonWillebrand disease

Answer 29

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

Question 30

What is vonWillebrand factor ?

Answer 30

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

Question 31

Von Willebrand disease types and treatments

Answer 31

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

Question 32

vWD lab evaluation

Answer 32

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

Question 33

Defects in secondary hemostasis, Single clotting factors vs multiple clotting factors

Answer 33

Single clotting factors- hemophilia | Multiple clotting factors- liver disease, vitamin K deficiency, exogenous anticoagulants

Question 34

hemophilia

Answer 34

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

Question 35

Clinical presentation of hemophilia and lab evals (A and B)

Answer 35

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%)

Question 36

Single factor deficiencies

Answer 36

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

Question 37

Coagulopathy of liver disease

Answer 37

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)

Question 38

Vitamin K deficiency

Answer 38

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

Question 39

Anticoagulants

Answer 39

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)

Question 40

virchows triad of thrombosis formation

Answer 40

Endothelial injury, abnormal blood flow, hypercoagulability

Question 41

Thrombophilia risk factors

Answer 41

a couple of risk factors can be either inherited or acquired (more commonly acquired) Protein C, S , anti Thrombin 3 deficiencies Hyperhomocysteinemia Dysfibrinogenemia

Question 42

sources of thrombi

Answer 42

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

Question 43

Site of thrombi by disease

Answer 43

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

Question 44

Factor 5 leiden

Answer 44

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)

Question 45

Factor 5 leiden testing

Answer 45

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

Question 46

Prothrombin 20210a mutation

Answer 46

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

Question 47

double heterozygotes

Answer 47

heterozygotes for factor 5 leiden and prothrombin gene mutation Greater risk of thrombosis

Question 48

AT3, protein C and Protein S deficiencies

Answer 48

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

Question 49

Antiphospholipid antibody syndrome

Answer 49

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

Question 50

Antiphospholipid antibody syndrome diagnostic and lab eval

Answer 50

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

Question 51

Diagnosis of lupus anticoagulant (LA) for antiphospholipid antibodies

Answer 51

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)

Question 52

Thrombotic states associated with thrombocytopenia

Answer 52

Heparin induced thrombocytopenia (HIT) Thrombotic microangiopathies: thrombotic thrombocytopenia purpura (TTP), Hemolytic uremic syndrome (HUS), disseminated intravascular coagulation (DIC) Disseminated intravascular coagulation (DIC)

Question 53

Heparin induced thrombocytopenia (HIT)

Answer 53

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)

Question 54

Thrombotic microangiopathies

Answer 54

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

Question 55

Typical Hemolytic UREMIC syndrome

Answer 55

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

Question 56

Disseminated intravascular coagulation

Answer 56

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