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Flashcards in Coagulation Deck (84):
1

What is hemostasis?

A series of physiologic processes that prevent bleeding when a blood vessel is damaged and, at the same time, keep blood in a fluid state

2

primary hemostasis

platelet plug formation

3

secondary hemostasis

coagulation cascade

4

fibrinolygsis

removal of clot

5

What converts prothrombin to thrombinf?

prothrombinase complex

6

What converts fibrinogen to fibrin

thrombin (IIa)

7

What does thrombin do?

coagulation, platelet activation, factor 8 activation, fibrinolysis, antigcoagulation etc....

8

What are natural inhibitors of coagulation? (inhibit clotting)

TFPI - binds factor 7a
Protein C serine protease - cleaves 5a, 8a
- cofactor protein S is crucial for its function
- both are vit K dependent
Antithrombin
- binds/inactivates serine proteases of the coaglation cascade
- activity augmented by heparin

9

What vasodilates the vessel wall?

nitric oxide
prostacyclins

10

What are anticoagulant properties that effect hte vessel wall?

heparan sulfate
thrombomodulin (protein C pathway)
TF pathway inhibotr
VW factor

11

What are components of platelets?

membrane receports
stored components
thromboxane

12

what is in the plasma?

VWF

13

What are disrdoers of primary hemostasis?

disoders of blood vessels
VW disease
thrombocytopenias
platelet dysfunction

14

What are bleeding sxs in disorders of primary hemostasis?

• Petechiae (pinpoint, nonraised intradermal hemorrhage)
• Easy bruising
• Epistaxis (nose bleeding)
• Menorrhagia (heavy menstrual bleeding)
• Unexpected pre or post operative bleeding
• Severe spontaneous bleeding

15

function of VWF?

mediates platelet adhesion at sites of vascular injury by binding to connective tissue and platelets

binds and stabilizes factor 7

16

What produces VWF?

vascular endothelium and stored in weibel palade bodies --> multimerization

17

What degrades VWF?

ADAMTS13

18

What is VW disease?

• Inherited bleeding disorder caused by deficiency or dysfunction of von Willebrand factor (VWF)
• Prevalence of symptomatic patients ~0.0023-0.01%
• Prevalence by screening populations to identify people with ~0.6-1.3%
• Autosomal inheritance

19

What are initial screenings assays for VWD?

• VWF antigen (VWF:Ag)
– Measures the amount of protein
– Antigen level many not equal activity level
• Ristocetin Cofactor Activity (VWF:RCo)
– Activity of VWF that causes binding of VWF to platelets in the presence of ristocetin resulting in platelet agglutination
– Measured by various assays
– Large coefficient of variation
• Factor VIII
– Clot based assays

(blood doesn't clot well)

20

What are tests for sub-typing VWD?

Multimeric analysis of VWF
• Type 1, 2N – all multimers are present
• Type 2A and 2B – largest are missing
• Type 2M – all multimer are present or increased largest
• Type 3 – too little VWF to measure

Ristocetin-induced platelet aggregation (RIPA):
• Differentiates Type 2A from 2B
• Increased sensitivity to ristocetin (“gain of function”) = Type 2B

21

What is bleeding time?

• Cutismadeinforearm,earlobe,orfingertip blot with filter paper
measure bleeding time when bleeding stops

Disadvantages
• Labor intensive
• Accuracy depends on operator skills
• Hard to standardize
• Not a specific indicator of platelet function
• Poor indicator of surgical bleeding risk!
• ShouldnotbeusedasascreenforVWD!

22

What are the classificaitons of VWD subtypes?

1 Partial quantitiative def of VWF
2 Qualittative VWF defect
2A No large multivers
2B No large multivers
2M Multivers are normal or incraesed high molecular weight
2N decreased affinity for factor 7
3 Deficiency of VWF

23

What is hte prevalence of VWD subtypes?

1 60-80%
2 7-30%
3 5-20%

24

14 y/o female with menorrhagia
VWF:AG 28% low
VWF:RCo 27% low
Multimer analysis: all multimer sizes present

Diagnosis?

T1 VWD

25

5yo boy w/ frequent nose bleeds
VWF:Ag normal
VWF:RCo LOW

Multimer analysis: large multimers are mising

Ristocetin induced platelet aggregation study POSITIVE

2A or 2B (large multimers missing)

2B: POSITVE ristocetin
often has decreased platelet count

26

What are the functions of platelets?

• Adhere to sites of vascular injury
– Glycoprotein Ia/IIa + collagen
– Glycoprotein Ib/IX/V + von Willebrand factor

• Release contents from granules
– Dense granules
• 2-7 per platelet
• ADP, ATP, calcium, serotonin

-- Alpha granules=
• 50-80 per platelet
• PDGF, insulin like growth factor 1, TGFβ, platelet factor 4, thrombospondin, fibrinogen, von Willebrand factor, fibronectin, factor V
• Express P-selectin and CD63

27

Platelet functions?

• Aggregate together to form a hemostatic platelet plug
– Glycoprotein IIb/IIIa + fibrinogen
– Glycoprotein Ib + von Willebrand factor
• Procoagulant surface for activated coagulation protein complexes on the phospholipid membrane

28

What are antiplatelet drugs?

cyclooxygenase - aspirin
ADP receptor/P2Y12 inhibitors - clopidogrel, prasugrel, ticagrelor, ticlodipine
glycoprotein IIb/IIa- abciximab, epitifibatide, tirofiban
phophodisterzase - cilostazol
adenosine reuptake - dipyridamole

29

What is a PFA?

• Measures closure time
• Simulatesprimaryhemostasisbycreatinghigh shear flow in capillary tube
• Membrane coated with agonists: – Collagen and ADP (C/ADP)
– Collagen and epinephrine (C/EPI)
• Plateletsgetactivatedbytheagonistsatthe membraneadhere on the membrane surfaceaperture is occluded by the platelet plug
• Instrumentsensesthetimeittakesbloodflowto stop

30

What is secondary hemostasis?

coagulation cascade
series of enzymatic steps to amplify a minor insult into formation of a fibrin plu

31

What proteases and cofactors are responsible for secondary hemostasis?

• Serine proteases with different protein domains
– Factors XII, XI, X, IX, VII,II (prothrombin), XIII
• Cofactors - Factors V, VIII, tissue factor
– Enhance the efficiency of the coagulation factors

32

What initiates coagulation?

• Tissue Factor (TF) and Factor VII binding
• TF expressed on damaged or stimulated cells
• <5% Factor VII circulates in activated (VIIa) form
• Forms a complex in the presence of calcium
• Converts Factor X to Xa directly or indirectly through Factor IX to IXa conversion

33

What are contact factors?

Prekallikrein (PK) is a serine protease that complexes with the cofactor high molecular weight kiniogen (HMWK)

PK is cleaved by factor 12a > kallikrein > activation of kinin pathway

34

What does factor XI do?

Activates factor 9
Activated by factor 12 and thrombin (creates a feedback loop)

35

What forms the xase complex?

• Factor IXa forms a complex with Factor VIIIa as cofactor
• Calcium ions required
• On a phospholipid surface • Activates Factor X to Xa

36

What forms the prothrombinase complex?

Factor Xa binds to Va as cofactor • Calcium ions required
• On a phospholipid surface
• Converts prothrombin (Factor II) to thrombin (Factor IIa)

37

What activates factor 8?

circulates as inactive form bound to VWF

activated by thrombin and factor Xa

38

What activates factor 5?

– Activated by several factors including thrombin, Factor Xa, plasmin, and others
– Can serve as a cofactor for Factor Xa without activation

39

What does thrombin do?

• Converts fibrinogen to fibrin
• No cofactor for enzymatic function needed
• Soluble fibrinogen cleaved by
thrombin turns to insoluble fibrin clot
– Thrombin cleaves off fibrinopeptide A and B from fibrinogen
– Fibrin monomers spontaneously polymerize to fibrin polymers
– Factor XIII stabilizes the clot by forming glutamyl-lysine bridges

40

What regulates thrombin?

• Positive feedback by activating Factors V, VIII, XI and XIII
• Negative feedback by activating Protein C and fibrinolysis
• Activates platelets

41

What is the testing principle to screen for coagulation disorders?

• Collect sample in citrate containing tube 
binds Ca2+blocks clot formation
• Centrifuge to separate plasma from
cellular elements
• Plasma + add calcium to overwhelm
citratestarts clot formation
– Add additional factors to speed up process
and drive down a particular pathway
• Measure how long it takes to form a clot

42

APTT

Intrinsic pathway

43

PT/INR

Extrinsic pathwayy

44

Thrombin time

common pathway

45

What is PT

• Measures the time (seconds) from VIIa/TF binding to clot formation
• Thromboplastin reagent (Tissue factor + phospholipid) + Calcium added to citrated plasma
• Tests the EXTRINSIC and COMMON pathways
• Used commonly to monitor warfarin therapy

46

When is hte extrinsic pathway initiated?

when blood is exposed to TF on damaged endothelium

--> PT assay

47

What can cause a prolonged PT?


 Inhibitor of Factor X, VII, V, II, or fibrinogen
 Antiphospholipid antibodies, such as Lupus anticoagulants
 Medications
 Warfarin (a vitamin K antagonist)
 Heparin (only at very high levels)
 Anti-Xa inhibitors (e.g. Rivaroxaban, Apixaban, Edoxaban)
 Direct thrombin inhibitors (eg. Dabigatran, Bivalirudin, Argatroban)
 Liver disease
 Consumption of factors as may be seen with disseminated
intravascular coagulation (DIC) or bleeding  Dilution
 Vitamin K deficiency
 Dysfibrinogenemia
 “Crap in a tube” = underfill, contamination from IV fluids, contamination from heparin in line, delay in testing, collecting in wrong tube, etc.

48

Why was INR developed?

to standardize PT testing and monitor pts on warfarin therapy

49

INR formula

INR = sample PT/mean PT x ISI

50

What is aPTT?

• Measuresthetime(seconds)clotformsthrough the activation of the contact and intrinsic pathways to clot formation
• Citratedplasmaismixedwithphospholipid (partial thromboplastin) and a contact activator + calcium
• TeststheINTRINSICandCOMMONpathways
• Used commonly to monitor unfractionated heparin therapy

51

What initiates aPTT measurement?

activation of FXII and contact factors on negatively charged phospholipid surfaces

52

What causes a prolonged aPTT?

• Inhibitor of Factor XII, XI, X, IX, VIII, V, II, or fibrinogen
• Deficiency of Prekallikrein and High Molecular Weight Kininogen (may not always be detected)
• Von Willebrand disease (due to low Factor VIII)
• Antiphospholipid antibodies, such as Lupus anticoagulants
• Medications
– Warfarin (a vitamin K antagonist) – Heparin
– Anti-Xa inhibitors (e.g. Rivaroxaban, Apixaban, Edoxaban)
– Direct thrombin inhibitors (e.g. Dabigatran, Bivalirudin, Argatroban)
• Liver disease
• Consumption of factors as may be seen with disseminated intravascular coagulation (DIC) or bleeding
• Dilution
• Vitamin K deficiency
• Dysfibrinogenemia
• “Crap in a tube” = underfill, contamination from IV fluids, contamination from heparin in line, delay in testing, collecting in wrong tube, etc.

53

What is used to assess the common pathway?

V, X, II, fibrinogen

Russel viper venom
activates factor X directly
DRVVT assay

54

What does TT measure?

• Measures the conversion of fibrinogen to fibrin (last step in the aPTT, PT/INR, and DRVVT)
• Detects abnormalities in fibrinogen to fibrin conversion

55

What causes a prolonged TT?

– Hypofibrinogenemia
– Dysfibrinogenemia
– Unfractionated heparin
– Direct thrombin inhibitors (hirudin, argatroban, dabigatran) – Fibrin degradation products (e.g. D-Dimer)

56

What has no effect on thrombin time?

– Warfarin
– Direct Xa inhibitors
– Factor II (factor 2) deficiency

57

Ho do you determine if a pt has a factor deficiency vs an inhibitor?

mixing study!

Mix pt plasma w/ normal plasma.
If aPTT corrects --> factor deficiency
If aPTT does NOT correct --> inhibitor

58

inheritance for hemophilia A and B?

x-linked recessive

59

Hemophilia A

def factor 8
1/5000 live male births

60

hemophilia B

def factor 9
1/30,000 live male births

61

what percentage of hemophilia A are new spontaneous mutations?

30%

62

What are clinical featurs of hemophilia A and B?

Intra-articular bleedings
– knees, elbows, ankles, shoulders, wrists
• Intramuscular hemorrhage 30% of bleeding events (compartment syndrome)
• Hematuria
• Intracranial hemorrhage (50% are spontaneous in adults)
• Gastrointestinal and oropharyngeal bleeding

63

How do you diagnosis w/ lab tests hemophilias?

• Isolated prolongation of aPTT
• Specific clotting factor assays are used for diagnosis (Factor VIII or IX)
– Severe <1%
– Moderate 1-5% – Mild 5-30%
• Can not distinguish Hemophilia A and B without labs

64

What is dyfibrinogenemia?

Protein does not function properly
• May be asymptomatic, hemorrhagic, or thrombotic
• Functional assays (activity) show lower level than antigen assays
• May or may not prolong clot times

65

Causes of elevated fibrinogen levels?

increasing age, females, pregnancy, acute phase reaction, smoking, exercise

66

Causes of reduced fibrinogen levels?

congenital, liver disease, disseminated intravascular coagulation (DIC), dilution, fibrinolysis

67

What is Owren's disease (factor V)?

acquired associated with exposure ot bovin thrombin in cardiac surgery

68

How do you treat factor VII (alexander's)

Treate with NovoSeven (recombinant FVIIa)

69

How do you treat factor X def?

Prothrombin complex concentrates
associated wtih amyloidosis
acquired form is more common

70

Factor XI def?

– 1 in 100,000
– Usually mild bleeding disorder
– FXI level does not correlate as well with bleeding tendency
– Bleeding
• Primarily trauma related
• Surgical bleeding at sites with fibrinolytic activity (tooth extraction, tonsillectomy, nasal surgery, prostatectomy)

71

FActor XII def?

1 in 1 million
NO clinical signficance

72

What does factor XIII do?

crosslinks strands of fibrin -> stabilizes clot (fbirin stabilizing factor)

73

Sxs of factor XIII def

– Delayed bleeding, 12-36 hour after trauma
• Umbilical stump
• Bleeding after circumcision
• Spontaneous intra-cranial bleeding
• Superficial bruising, subcutaneous hematomas – Miscarriages
– Poor wound healing

74

How do you test for factor XIII def?

• Standard screening tests (PT, aPTT,
TT) are normal!!
• Screening test: clot solubility test (dissolve the clot with 5M urea or acid)
• Confirm with FXIII antigen or activity test

75

A patient has a prolonged PTT but normal PT and TT. Which of the following may explain this set of results?
A. Factor II (Factor 2) deficiency B. Factor V (Factor 5) deficiency C. Factor VII (Factor 7) deficiency D. Factor IX (Factor 9) deficiency

Factor 9

76

A patient has a prolonged INR and PTT, but normal TT. Which of the following may explain this set of results?
A. Fibrinogen deficiency
B. Factor V (Factor 5) deficiency
C. Factor VIII (Factor 8) deficiency D. Factor XIII (Factor 13) deficiency

Factor 5

77

Thrombin forms clot formation/thrombosis where as PLASMIN causes...

clot lysis/bleeding

Fibrinolysis

78

What is plasmin?

• Plasminogen --> plasmin
• Proteolytic enzyme
• Activated by thrombin
• Degrades cross-linked fibri > fibrin degradation products (FDP) appear in circulation

79

What is a d dimer?

a fibrin degradation product

80

What regulates fibrinolysis?

plasminogen is activated by tPA

81

What is tPA?

– synthesized in endothelial cells
– half life about 5 min
– inhibited by plasminogen activator inhibitor-1 (PAI-1)

82

What does alpha-2 plasmin inhibiotr do?

inhibits plasmin in circulation

83

What are the TEG parameteres?

• Rtime
– Measuresthetimetofibrinformationwithinaclot
– Determinedbyclottingfactorsandinhibitorbalance(eg.heparin)
•  angle
– Anglebetweentheinitialslopeofthetracingandthehorizontal – Representsspeedofclotstrengtheningbyfibrincross-linkage
•K
– TimefromtheendofRuntiltheclotreaches20mm – Representsthespeedofclotformation
• Maximum amplitude (MA)
– Thestrengthoftheclot
– Dependsontheintegrityofplateletfunctionandfibrinogenbinding
• Ly30 and Ly60
– Lysis of the clot at 30 minutes and 60 minutes
• Coagulation Index (CI)
– Overallassessmentofcoagulation
– CI = −0.6516R − 0.3772K + 0.1224MA + 0.0759α − 7.7922
– Values +3.0 = hypercoagulable

84

What is platelet mapping?

Compares maximum amplitudes of response to various agonists
– Baseline = MATHROMBIN
– Activator (Reptilase + Factor XIII) = MAFIBRIN
– Activator +ADP = MAADP
• Used for ADP receptor inhibitors (e.g. clopidogrel)
– Activator + Arachidonic Acid = MAAA
• Used for Aspirin