lec 5/6/7: Coagulation and hemostasis

clot

end product of hemostasis and coagulation, this term is used many ways, can be normal or pathologic

thrombus

a solid mass of blood-borne elements formed within (and attached to) a vessel, synonymous with "blood clot", may be normal or pathologic

white thrombus

clot formed of mostly platelets and fibrin

red thrombus

clot formed with platelets, fibrin, and RBCs

thrombosis

inappropriate activation of hemostasis leading to pathologic thrombus formation. However, it is also used as a normal term when referring to endothelium and its properties of physiologically appropriate clotting

plasma

portion of blood without cellular elements, has clotting factors

serum

blood plasma that does not contain fibrinogen or clotting factors

hemostasis, inflammation, regeneration

what are the three core responses to injury?

platelets/thrombocytes

annucleate cells that function for hemostasis

Bone marrow --> Megakaryocytes ---> Capillaries --> fragmented into platelets

origin of platelets

Thrombopoietin (TPO)

stimulus development of platelets

circulate for 8-12 days then removed from circulation by spleen and tissue macrophages

lifespan of platelets

thrompoietin and IL3

maturation of progenitors to mekakaryocytes is dependent upon

resting vs activated platelet

Resting looks like a circle, activated looks like a splat

vWF binding to GPIb

-leads to increase in calcium ---> activation of prothrombin to thrombin

the first binding that allows activation of platelet

gray platelet syndrome

pts lack alpha granules in circulating platelets

-pts present with bleeding and marrow fibrosis

1. vasoconstriction at site of injury

2. primary hemostasis: platelet plug

3. secondary hemostasis: coagulation cascade

4. clot retraction: growth of fibrous tissue into the clot to close the breach

5. fibrinolysis

6. healing: cellular proliferation and remodeling

what are the fundamental steps of hemostasis

-endothelin from endothelium

-thromboxane A2 from platelets

-serotonin from platelets

POTENT vasoconstrictors

modulators of vasoconstriction in the first step of hemostasis

Weibel-Palade bodies

in the second step of hemostasis, injury releases these bodies containing vWF and P-selectin

von Willebrand factor

adhesion molecule that allows platelets to attach to exposed to attach to exposed matrix with ligand GpIib

-ADP

-platelet activating factor

-thromboxane 2

once platelet is adhered to GpIb, the platelet releases

platelet actiivating factor

-released by platelet after adhesion to GpIb

-promotes recruitment and aggregation of more platelets

•Increased shearing force on platelet/endothelium

•Vessel injury

•Humoral signals

triggers of platelet plug formation

platelet adhesion, platelet activation, platelet aggregation

what are the 3 steps of platelet plug formation

-AAA

-platelets adhere to exposed subendothelial tissue through key receptors

what is the first step of platelet plug formation

GPIa binds directly to collagen

this binding causes conformational changes in platelet and tis receptors, facilitating vWF-platelet initeraction

strongest adhesion point in the formation of platelet plug

-leads to conformational change that unmasks GIIb-IIIa binding sites

effects of GPIb binding to vWF onn exposed collagen

GPIb and GPIa

conformational changes in ______ during adhesion lead to platelet ACTIVATION

-receptors undergo conformational change involving calcium influx ultimately leading to release of granules by exocytosis

-activated platelets go dramatic shape change mediated by calcium and structural proteins

-activate platelets recruit and activate nearby platelets

mechanism of step 2 of platelet plug formation: activation

-ligands from subendothelial tissue in adhesion

-ADP from platelets: activates nearby platelets, increases TXA2

-thrombin

-serotoinin

key agonists of activation in platelet plug formation

calcium

secretory product of platelet activation that is required for platelet conformation change

Activation step of platelet aggregation exposes and activates platelet receptor GPIIb-IIIa which binds fibrinogen from the blood

-fibrinogen links platelets, creating a plug

mechanism of aggregation step in platelet plug formation

fibrinogen

-linker protein between platelets that creates a plug

-formed ini the liver and is released from platelets

relatively weak (but formed fast)

Minor injuries that occur daily may be healed merely by the platelet plug. In more severe injuries, the platelet plug is strengthened into a secondary hemostatic plug by coagulation.

strength of the platelet plug?

-site specificity!

-near by NORMAL endothelium have glycocalyx (heparin sulfate) to prevent non specific adhesion, vasodilators (PGI2, NO)

what prevents overzealous platelet plug formation?

formation of fibrin which becomes cross-linked and firmly stabilizes the platelet plug in a mesh like web

what is the goal of secondary hemostasis?

calcium and prothrombin activator

what is required to turn prothrombin into thrombin?

thrombin

turns fibrinogen into fibrin

within a few minutes of clot formation and is completed in 20-60 min

when does clot retraction occur?

thrombin!

requires calcium and platelets

clot retractiion is triggered by

actin, myosin, and thrombosthenin

clot retraction occurs due to the interaction between ____, ______, and ________ in the platelets

-expulsion of serum

-approximation of wound edges

results of clot retraction

liver

where is fibrinogen synthesized

-platelet aggregation (binds GPIb-IIIa)

-substrate firbin formation

-binding site for thrombin funciton

function of fibrinogen

fibrin fibers, which result ini soft clot (non-covalent bonds)

fibrin monomers spontaneously polymerize and form

Factor XIIIa

-factor that crosslinks the fibrni fibers and stabilizes the clot (covalent bonds)

source: plasma and platelet secretion

-activated by thrombin

source and activation of factor XIII

normal intact vascular wall is antithrombotic

-negatively charged via heparin sulfate

intirinsic tissue properties inhibiting clot formation

conversion of plasminogen to plasmin

first step of fibrinolysis

plasminogen

plasma protein made in the liver that circulates and gets trapped in the clot with other molecules

plasmin

plasma protein that converts stable fibrin to fibrin degradation products

altepase, reteplase, urokinase

name the t-PA analogs used for acute thrombotic events

D dimer

-measurement of fibrin degradation products

a2-antiplasmin (from liver)

as form of regulation of clot lysis, if plasmin isn't bound to fibrin, it is quickly bound to:

plasminogen activator inhibitors (PAI1 and PAI2)

proteins that inhibit activation of plasmin by inhibiting its activators

-produced by endothelial cells and inhibits tPA

protein C

activated ____________ inhibits plasminogen activator inhibitors, facilitating fibrinolysis

results in reduced a2-antiplasmin, causing excessive plasmin activity---> bleeding problems

how does liver disease effect plasmin activity?

1. vasoconstriction

2. primary hemostasis

3. secondary hemostasis

4. clot retraction

5. fibrinolysis

6. proliferation and remodeling

steps to repair of an injured vessel

thrombin

factor IIa is aka

factor VIII

factor ____ deficiency causes hemophilia A

-signifiacnt cause of inappropriate bleeding

hematoma

spilled blood that leads to swelling

ecchymosis

blood released into tissues without significant swelling

factor Xa

factor required to activate prothrombinase to prothrombin

Factors 2, 7, 9, 10

-protein C and S (anticoagulant factors)

Factors requiring activation by vitamin K

prothrominase

enzyme complex that converts prothrombin (factor II) to thrombin

factor Xa, factor Va, calcium, phospholiipipd

what are the key components of the prothrombinase complex?

factor Va

cofactor that stabiliizes/accelerates thrombin productioin

-component of the prothrombinase compplex

formation of prothrombinase

what is the rate limiting step of the common coagulation cascdee?

proteolyzes fibrinogen to fibrin

primary action of thrombin

factor VIIIA

factor that crosslinks fibrin into mesh of stable fibrin

-clot stabilization

intrinsic pathway

coagulation pathway that takes longer (1-6 min) and is measured by PTT

(you Play Table Tennis Inside)

•Involves Factors XII, XI, IX, VIII also HK and Kallikrien

factors inivolved in the intrinsic pathway

trauma to blood or blood exposure to collagen

what activates the intrinsic pathway?

extrinsic pathway

-coagulation pathway with rapid onset, measured by PT

-Play Tennis Outside

trauma to vessels and tissue--> tissue factor exposed

-inflammation can drive tissue factor expression

-factor XII is activated by negatively charged surface

the extrinsic pathway is activated by

lipoprotein on non-vascular cell membranes or exosomes

where is tissue factor of the extrinsic pathway from?

Factor VII, activating to VIIa

initiating the extrinsic pathway, tissue factor binds

TF-VIIa-Ca2+ complex

complex from the extrinsic pathway that activates factor X

all its factors are from blood

-in extrinsic pathway, tissue factor is NOT from blood

why is it called the intrinsic pathway?

High Molecular Weight Kininogen (HMWK)

co-activates Factor XII as an anchor and is a bradykinin precursor

Kallikrein

HMWK activates _____, which accelerates XIIa formation

HMWK anchored to XIa activates factor IX (christmas factor)

what cofactor activates factor IX?

IXa-VIIIa-Ca2+ (tenase)

intrinsic pathway complex that actiivates factor X

Factor VIIIa

serves as a cofactor to stabilize the initrinsic pathway complex to activate common pathway

thrombin

the major protease of the clotting cascade

-activates fibrinogen

-activates factor XIII

-activates factor Va and VIIIa

-paracrine effects on surrounding endothelial cells (NO, PGI2, tPA)

main functions of thrombin

thrombin (plays a role)

what actiivates factor VIII and V?

TFPI (tissue factor pathway inhibitor)

-anticoagulant factor that blocks formation of the TF-VIIa-Ca2+ pathway

-blocks factor VIII (extrinsic pathway )

antithrombin III

-inhibits factor Xa and thrombin

-complex with heparin sulfate enhances the binding to inhiibit coagulatiion

-SLOWLY neutralizes thrombin without heparin

Va and VIIIa (anticoag)

Protein C and S (bound to phospholipid and calcium) work together to inactivate what cofactors?

thrombomodulin-thrombin complex

what activates protein C?

heparin

catalyst for the binding of antithrombin III to thrombiin

-vitamin K dependent carboxylation reaction required for binding factors to calcium and platelet membranes for activation

what is the importance of vitamin K in activation of some clotting facotrs?

reduced vitamin K (by epoxide reductase in the liver) is active

what form of vitamin is active?

warfarin blocks epoxide reductase, which is necessary to activating vitamin K

how does warfarin work as an anticoagulant drug?

-intrinsic!

-PTT prolonged via interaction with antithrombin III

is heparin's affect on the intrinsic or extrinsic pathway?

extrinsic

-PT is prolonged

is warfarin's affect on the intrinsic or extrinsic pathway?

thrombocytopenia--- indicate splenectomy

thrombocytosis--- indicate splenomegaly

platelet findings associated with spleen pathologies

bleeding time (3mm puncture made with lancets)

-time wound takes to stop bleeding

best measurement of primary hemostasis (soft clot formation)

3-6 minutes

-abnormal is greater than 6 min

what is a normal bleeding time?