hemostasis_surgury 1

HEMOSTASIS

Department of Surgery

FEU-NRMF MEDICAL CENTER

HEMOSTASIS

balance of the physiological processesprevent excessive bleeding after vessel injurymaintain a viable circulation by keeping the

blood in an uncoagulated state“Self-sealing” system to prevent

excessive bleeding and potentially life threatening states

Four components

Vessel function constriction of injured vessels

Platelet function formation of platelet plugs

Coagulation formation of fibrin plugs

Fibrinolysis dissolution of blood clots

Normal Hemostasis

ExcessiveBleeding

ExcessiveClotting

VesselInjury

Means stopping of blood – prevents blood loss Imbalance in one direction may lead to excessive bleeding, imbalance in

the other may lead to thrombus formation

Overview of blood coagulation

PRIMARY HEMOSTASIS

SECONDARY HEMOSTASIS

Primary HemostasisFormation of platelet plug

Secondary HemostasisFormation of blood clot

Hemostasis

response is dependent on overlapping and interdependent responses

Vascular Endothelium

Blood vessel wall integrity is essential for prevention of blood loss – provides potent anticoagulation surface.

Vascular Endothelium

ECs form monolayer resting on continuous basement membrane; constitutes first barrier of defense from hemostasis and thrombosis

EC Products Secreted into the Subendothelium

EC surface products secreted into the blood stream

LUMEN

ECs provides numerous proteins to subendothelium essential for cell–to-cell interactions and formation of diffusion barrier to prevent blood loss.

Vascular Endothelium

EC Products Secreted into the Subendothelium

● basement membrane ● elastin ● fibronectin● collagen III and IV ● lamilin ● mucopolysaccharides● microfibrils ● vibronectin ● vWf

● protease inhibitors

These proteins essential for cell–to-cell interactions and formation of diffusion barrier to prevent blood loss into extravascular spaces.

Vascular Endothelium

ECs also secrete numerous substances into the vascular lumen to prevent clotting (e.g. heparin)

which promote fluidity of the blood

EC surface products secreted into the blood stream

● PGI2 ●glycosaminoglycans ● tissue factor● EDRF ● ATIII/heparin sulfate ● vWf● t-PA ● protein kinase/thrombomodulin ● Factor V● urokinase ● plasminogen activators ● inhibitors (PAI-1, PAI-2● nitric oxide ATIII)

● IL1, TNFα ● endothelin-1 ● PAF

Anticoagulants Procoagulants

These substances promote fluidity of the blood.

Vascular Endothelium

Normal endothelium acts as a potent anti-coagulation surface, upon stimulation or injury, endothelium transforms into a potent pro-coagulation surface as subendothelial collagen is exposed

Primary Hemostasis

Primary Hemostasis

response to vascular injury that produces a platelet plug at the site of damage

immediately limit bleeding through the formation of a loose platelet plug

Platelets

Minute round or oval discs (1-4 mm diameter).

Do not have a nucleus and can not divide.Formed from megakaryocytes in bone

morrowNormally 150-450,000 per ml of blood

Platelets

Adhering to the endothelial wall at the site of injury

Releasing potent anticoagulant compounds

Aggregating to form a plug Providing a phospholipid surface for

activated coagulation enzyme complexes

Platelets

Have many functional characteristics of whole cells Contains contractile proteins (actin, myosin and

thrombosthenin) Residuals of ER and Golgi apparatus Mitochondria and enzymes to form ATP and ADP Can synthesis prostaglandins (thromboxane A2) Forms fibrin stabilizing factor Forms growth factors

Platelets Cell membrane contains glycoproteins that reduces

adherence to normal endothelium yet promotes adherence to injured areas of vessel wall,

especially injured ECs and collagen of subendothelium. Membrane also contains phospholipids that play

activating role at multiple points in the clotting process. Platelets have a half life of 8-12 days Eliminated from the circulation mainly by tissue

macrophages, especially in spleen.

Platelets

Endothelial cell injury exposes subendothelial collagen which causes change in platelet shape and adherence.

vWf- bridges platelet membrane glycoprotein to the exposed collagen at site of injury.

Vascular Injury

INJURED

Vasoconstriction

an immediate reflex

Initial response to injurydiminishing blood

lossLocal constriction

of smooth muscle

Vasoconstriction

Transient, typically lasts minutes (severed limbs)

Result of release of local humoral actors, neural reflexes (caused by pain), local myogenic spasm

Small blood vessels, release of vasoconstrictor thromboxane A2 from platelets is primarily responsible

Platelet adhesion

Exposed collagen from the damaged site will promote the platelets to adhereundergo degranulation and release

cytoplasmic granules Occurs within 15 secs after injury

Platelet adhesion

Injury to intima

Subendothelial collagen

(Glycoprotein receptor Ib)

Platelet adhesionvWF

Von Willebrand factor (vWF)

Multimeric plasma glycoprotein required for normal hemostatic platelet plug formation

Forms a bridge between platelet glycoprotein IB and exposed collagen in the subendothelium

It is the “glue” that binds platelets to collagen.

vWF is also responsible for the binding and transport of factor VIII (antihemophilic factor), a procoagulant protein in plasma.

Platelet adhesion

cytoplasmic granules serotoninADP and Thromboxane A2

Platelet adhesion

serotonin, a vasoconstrictor ADP attracts more platelets to the area thromboxane A2 promotes platelet

aggregation, degranulation, and vasoconstriction

EM of normal and activated platelets

Normal Aggregated Platelets

ADP and thromboxane A2 promote more platelet adhesion therefore more ADP and thromboxane“Second wave”

Platelet plug

The positive feedback promotes the formation of a platelet plug

Primary platelet plug is only good for stopping bleeding in the first minute- it can not sustain

hemostasis. Heparin does NOT interfere with this reaction

Arachidonic acid pathway

ASPIRIN and other NSAID

Primary hemostasis

Defects : associated with mucocutaneous bleeding, characterized by epistaxis, ecchymosis, genitourinary bleeding, or gingival bleeding “first wave” of aggregation“second wave” ( granule release )

Secondary Hemostasis

Secondary Hemostasis

Cascade of enzymatic reactions that ultimately results in the conversion of fibrinogen to fibrin monomers

cross-linked into insoluble strands that serve to stabilize the loose platelet clot formed in primary hemostasis

Secondary Hemostasis

triggered by the release of tissue factor from epithelial cellsPlatelets, vascular wall and multiple

circulating or membrane-bound coagulation factors

Formation of Prothrombin Activator

Prothrombin activator formed in two ways:Extrinsic pathway Intrinsic pathway

In both pathways, inactive forms of blood clotting factors are converted to active forms.

Designated by Roman numerals, small “a” for active

The common pathway

p ro th ro m bin * th ro m bin

fib rino ge n

F ib rin m on om er

F ib rin po ly m erC L O T

* X a

X III X IIIa

Va V

C o m m o np a thwa y

Ca++

Note that both the Extrinsic and Intrinsic Pathways converge on Factor X

Prothrombin Activator Complex (rate limiting step)

Vessel Injury

Characteristics of clotting reactions in the cascade

A proteolytic enzyme generates the next enzyme in the cascade by cleavage of a proenzyme

The reactions occur on a phospolipid surface ( platelet membrane )

Each reaction requires a helper protien to bring the enzyme and substrate together

Clotting Factors in the BloodFactor Common Name Pathway Function

I Fibrinogen Common Fibrin clotII 1Prothrombin* Common Generates fibrinIII Tissue factor (Thromboplastin) Extrinsic Receptor for VIIaIV Calcium All Co-factorV Proaccelerin Common Receptor for XaVII 1Proconvertin* Extrinsic Activates IX and XVIII Antihemophilic factor Intrinsic Receptor for IXaIX 1Christmas factor* Intrinsic Activates XX 1Stuart factor* Common Activates prothrombin and VIIXI Plasma thrombin antecedent* Intrinsic Activates IXXII Hageman factor* Intrinsic Activates XIXIII Transglutamidase Common Cross-links fibrin

Von Willebrand factor Platelets Platelet activation and bindingPrekallikrein* Intrinsic Activates XIIKininogen Intrinsic Receptor for kallikrein and XII

Extrinsic PathwayProcess begins with trauma to vascular

wallinitiated by tissue factor release by damaged

tissue initiated by contact with Factor VII and

platelets with collagen in vessel wallClotting begins within seconds

Extrinsic Pathway Release of Tissue Factor (thromboplastin).

Functions as a proteolytic enzyme complex Activation of Factor X.

Tissue Factor further complexes with Factor VII which in the presence of Ca++ activates Factor X to Xa.

Formation of Prothrombin Activator Xa combines with phospholipids and Factor V to form

prothrombin activator complex In the presence of Ca++, this splits prothrombin into

thrombin in the Common Pathway

Extrinsic Pathway

Release of Tissue Factor (thromboplastin).Functions as a proteolytic enzyme complex

Extrinsic Pathway

Activation of Factor X. Tissue Factor further complexes with Factor

VII which in the presence of Ca++ activates Factor X to Xa.

Extrinsic Pathway

Formation of Prothrombin ActivatorXa combines with phospholipids and Factor V

to form prothrombin activator complex

Extrinsic and common pathways

pro th rom bin * th rom b in

fib rinogen

F ibrin m onom er

F ib rin po lym erC L O T

*X a

X

X III X IIIa

*V IIa

T F

In ju redC ells

V II

Va V

unk .

Extrinsicp a thwa y

C o m m o np a thwa y

= C a lcium & P L com plex

* = ac tive serine p ro tease Tissue Factor

Tissue Trauma

Intrinsic Pathway

Initiated by trauma to blood or exposure of blood to vascular wall collagen

Takes 1-6 minutes to cause clotting All components are “intrinsic to the circulating

plasma” and NO surface is required to initiate clotting

Intrinsic Pathway

Causes activation of Factor XII (to XIIa) and release of platelet phospholipids

Activation of Factor XI by Factor XII. This reaction also requires High Molecular Weight (HMW) kininogen and is accelerated by prekallikrein.

pro th rom bin * th rom b in

fib rinog en

F ibrin m onom er

F ib rin po lym erC L O T

*X a

X

X III X IIIa

*IX a

IX

*X Ia

X I

*X IIa

X IIW O U N D surface

*ka llik rein

k in inog en (H M W K )

prekallik re in

Va

V IIIa V III

V

Intrinsic p a thwa y

C o m m o np a thwa y

= C a lcium & P L com plex

* = ac tive serine p ro tease

Intrinsic and common pathways

Prothrombin Activator

Factor X activation

Extrinsic XaseTF- VIIa

Intrinsic XaseVIIIa-XIa

50x more effective at catalyzing factor X activation

Functions of thrombin

Conversion of fibrinogen to fibrin Activation of factors V, VIII, XI and XIII

Prothrombin unstable plasma protein that splits in half to

form thrombin. Made by liver and requires vitamin K

Common Pathway

In the presence of Ca++, this splits prothrombin into thrombin in the Common Pathway

Prothrombin Activator Complex

Thrombin Activation

co llagen

WO UNDv W F

endo th eliu m

p la te le tPL surfa c e

C a

C a

Va

Xa

C ircu la tio n

N H2C O O H

G la G la

P ro -T h ro m b in

SS

p ro teo ly tic cu t

N H2C O O H

G la G la

T h ro m b inP ro -

SS

Phospholipid surface

Common Pathway

Fibrinogen large plasma protein formed in liver Acted on by thrombin (removes 4 peptides) to form fibrin which

polymerizes into fibrin fibers

Common Pathway

Fibrin-stabilizing factor from platelets trapped in clot is activated which causes formation of covalent bonds between fibrin monomer molecules

BLOOD CLOT

INTRINSIC PATHWAY (12-11-9-8-10)

XII (Hageman Factor)

XIIa

XI XIa + Ca ++

IXa + VIII + Ca++ + phospholipid, F3,Plt

Xa

Prothrombin (Factor II)

Thrombin

Fibrinogen Fibrin ClotXIII a

EXTRINSIC PATHWAY

Tissue thromboplastin + VII + Ca++

V

All coagulation factors synthesized in the liver except for thromboplastin, Ca++

Factors 2,7,9,10 – vitamin K dependent

The coagulation system

Regulated system In addition to Clot formation :

Balanced the propagation of clotFeedback inhibition of cascadeMechanism of fibrinolysis

Feedback inhibition of cascade

Tissue factor pathway inhibitor (TFPI )Antithrombin IIIProtien C system

Tissue factor pathway inhibitor (TFPI )

pro th rom bin * th rom b in

fib rinogen

F ibrin m onom er

F ib rin po lym erC L O T

*X a

X

X III X IIIa

*V IIa

T F

In ju redC ells

V II

Va V

unk .

Extrinsicp a thwa y

C o m m o np a thwa y

= C a lcium & P L com plex

* = ac tive serine p ro tease Tissue Factor