Circulation disturbances - Hemostasis

Pathology - Lec 12 - Exam 2

hemostasis

arrest of bleeding → physiologic response to vascular damage & a mechanism to seal an injured vessel to prevent blood loss

Normal hemostasis is the result of what?

a series of complex, well-regulated processes that have 2 main functions

What is the main function of hemostasis under normal conditions?

maintains blood in fluid state

What is the main function of hemostasis under abnormal conditions?

rapid formation of a local hemostatic plug at a site of vascular damage

What is the big picture process of hemostasis?

• transient vasoconstriction (brief reflex)

• platelet aggregation

• formation of platelet plug (1^o hemostasis)

• formation of a fibrin-platelet aggregate (2^o hemostasis)

• dissolution of fibrin-platelet aggregate (thrombolysis/ fibrinolysis)

What causes the brief reflex vasoconstriction at the initial site of vascular injury?

endothelin release from damaged endothelial cells

serotonin & TXA₂ from platelets

What are the physiologic goals of transient vasoconstriction?

• reduce volume of blood flowing through the affected area

• bring opposing endothelial surfaces closer together

Disruption of the endothelium exposes what?

• collagen, fibronectin, & other ECM glycoproteins & proteoglycans

• subendothelial von Willebrand factor (vWF)

What releases von Willebrand Factor and what does it do?

activated endothelial cells & platelets

coats subendothelial collagen

What is vWF necessary for?

firm platelet adhesion

What are the 5 main steps of 1^o hemostasis?

1. platelet adhesion (vWF attracts)

2. shape change

3. granule release (ADP, TXA₂)

4. recruitment

5. aggregation (hemostatic plug)

What are the 4 main steps of 2^o hemostasis?

1. tissue factor stimulates CoAg cascade

2. phospholipid complex expression

3. thrombin activation

4. fibrin polymerization

What function does fibrin serve in 2^o hemostasis?

creates a firm net that stabilizes platelets

What 2 substances initiate antithrombotic events? 

1. tPA → fibrinolysis

2. thrombomodulin → blocks CoAg cascade

T/F: Endothelium is a dynamic interface during hemostasis.

TRUE

A normal endothelium promotes what? How?

smooth, non-turbulent blood flow

vasodilation, antithrombotic & profibrinolytic (prevent clotting)

What types of injury activates the endothelium?

oxidative stress, hypoxia, inflammation, infectious agents, and tissue injury

An injured (activated) endothelium promotes what?

• vasoconstriction

• enhance platelet adhesion & aggregation

• stimulate coagulation

• prothrombotic & antifibrinolytic (avoid breakdown)

What are platelets?

annucleate cell fragments derived from megakaryocytes circulating in blood

What do platelets adhere to?

exposed subendothelial collagen, laminin, fibronectin, etc

What are the membrane receptors that various glycoproteins bind to?

collagen, vWF, thrombin, fibrin, etc

T/F: An intact endothelium promotes interaction.

FALSE - prevents interaction

Platelets provide a surface for the assembly of what?

coagulation factors → IX & X

What are the 2 types of cytoplasmic granules?

1. alpha → growth factors, CoAg factors

2. dense → adenine nucleotides, Ca²⁺, inorganic phosphates, serotonin

coagulation

process by which blood changes from a liquid to a gel, forming a blood clot

What are the 3 classical coagulation pathways?

1. intrinsic

2. extrinsic

3. common

Coagulation factors can be:

• serine protease proenzymes

• non-enzymatic proteins 

• platelet phospholipids

• transmembrane cell surface receptors

• calcium

What are serine protease proenzymes?

enzymatic factors → vitamin K dependent factors (2,7,9,10)

What synthesizes serine protease proenzymes?

the liver

T/F: The circulating ½ life of serine protease proenzymes is long.

FALSE - short = hours to days

Which factor has the shortest ½ life?

factor VII → 4-6 hours

T/F: Serine protease proenzymes require activation to become proenzymes or zymogens

TRUE

The vitamin K dependent factors are activated by what?

vit K dependent carboxylation of glutamic acid residues on the factors 

T/F: The vitamin K dependent factors are only found in the intrinsic and common pathways.

FALSE - found in all 3

The intrinsic pathway involves which substances?

high molecular weight kininogen, prekallikrein, & factors XII, XI, IX, VIII

T/F: The intrinsic pathway exhibits contact activation.

TRUE - HMWK, prekallikrein, XII, & XI

What activates the intrinsic pathway?

negatively charged substances → collagen, activated platelets, endotoxin

The intrinsic pathway is a series of activation events:

factors IX & VIII activate factor X → start of the common pathway

Which pathway usually initiates coagulation in vivo?

the extrinsic pathway

Explain the initiation of coagulation in vivo.

tissue factor (factor III) contacts & complexes w/ factor VII (or VIIa) → factor III-VIIa complex can activate factor IX (intrinsic) and factor X (common)

What is tissue factor?

CoAg factor III → cell surface glycoprotein expressed on subendothelial tissue

T/F: Factor X can be activated by either the intrisic or extrinsic pathway.

TRUE - binds to factor V

Where can factor V be found?

circulating in plasma

within platelet alpha granules

Factor X binds to what to form the prothrombinase complex?

Factor X-V-Ca²⁺ (10,5,4)

What does prothrombinase complex converts what?

prothrombin (factor II) to thrombin (factor IIa)

Thrombin converts what?

fibrinogen (Factor I) to fibrin (Ia)

Which factor cross-links fibrin?

factor XIII (fibrin stabilizing factor)

T/F: The extrinsic, intrinsic, and common pathways happen in a stepwise mannor.

FALSE - all pathways occur simultaneously

What are the 3 main platelet events in 1^o hemostasis?

1. platelets recognize endothelial damage

2. platelets become activated

3. recruitment & aggregation of additional platelets to form platelet plug

How do platelets recognize endothelial damage?

• change in blood flow

• attracted to negatively charged ECM matrix

• adhere to exposed vWF

What happens after platelets become activated?

• results in shape change → increase surface area

• release of secretory granules

• phospholipid activiation/ phosphatidylserine exposure → binds CoAg factors

What are the 2 main steps of platelet aggregation?

1. fibrinogen (circulating plasma glycoprotein) bridges loosely link platelets → form loose aggregate

2. platelet contraction & polymerization of fibrinogen to fibrin consolidates aggregate to a dense plug

T/F: In many cases, fibrin is necessary for complete hemostasis.

TRUE - mediators from endothelial cells, activated platelets, and the ECM trigger activation of the CoAg cascade & the formation of fibrin for 2^o hemostasis

The initiation of blood coagulation… (sorry this is the big bolded statement on slide 28)

occurs via the extrinsic tenase (VII, X) complex following tissue injury. However, sustained generation of factor Xa is dependent upon the intrinsic tenase (VIII, IX, X) complex which is 50x more efficient and not as susceptible to inhibition.

What are the 3 steps of the cell-based model of coagulation?

1. initiation

2. amplification

3. propagation

Cell-based model of coagulation: initiation

exposure of tissue factor (III) on perivascular fibroblasts/ activated endothelium

Cell-based model of coagulation: amplification

activation of small amounts of thrombin/ activation & aggregation of platelets (P)

Cell-based model of coagulation: propagation

activation of large amounts of thrombin/ increased numbers of activated platelets (AP)/ formation of fibrinopeptides and fibrin

Cell-based model of coagulation: written out fully (sorry for the word vomit) - slide 29

Vascular damage results in tissue factor (TF) exposure and production of thrombin and factor IXa via the extrinsic pathway (initiation). The small amounts of thrombin formed during initiation cleave von Willebrand factor (vWF) from factor VIII, enhancing platelet adhesion to the damaged site. Thrombin activates platelets (P), which result in degranulation and the aggregation of additional platelets. Thrombin also results in the generation of factors Va, VIIIa, and XIa (amplification). During propagation, activated platelet (AP) membranes support assembly of intrinsic tenase complex (IXa/VIIIa) that is rapidly induced from factor XIa. Factor Xa and its coreactants form prothrombinase to generate large amounts of thrombin, which is released directly onto platelets to cleave fibrinogen into fibrinopeptides, which cross-link to form fibrin.

What is the stimuli of extrinsic coagulation?

release of tissue factor (III)

What releases tissue factor (III) thus stimulating extrinsic coagulation?

activated endothelium, activated platelets, perivascular cells, ECM

What is the stimuli for intrinsic coagulation?

binding of factor XII to negatively charged surfaces

What are the negatively charged surfaces that can trigger intrinsic coagulation via binding of factor XII?

ECM, activated platelets, endotoxin

Initiation of coagulation via the extrinsic pathway:

• release of tissue factor

• factor VII circulates in blood plasma (1% active VIIa)

• factor VII, with Ca²⁺ forms a TF:VII complex on the surface of the injured area (phosphatidylserine helps with orientation)

• TF:VII is activated toTF:VIIa (may be autoactivated or activated by many factors)

• TF:VIIa activates factors X and IX

Amplification of coagulation via the common pathway:

• active factor X (Xa) binds to factor V, Ca²⁺, and phospholipids to form prothrombinase complex

• factor Xa cleaves prothrombin (II) into thrombin (IIa)

• thrombin converts fibrinogen (I) to fibrin (Ia), activates platelets and factors XI, VIII, V, and XIII

Propagation of coagulation via the intrinsic pathway:

• contact activation of factor XII - also activation of factors XI & VIII by thrombin (factor XII can autoactivate)

• conversion of proenzymes to active enzymes occurs on platelet surface

• activation cascade of XII → XI → IX

• factor IX complexes with factor VIII = intrinsic tenase complex (+ Ca²⁺ & phospholipids)

• intrinsic tenase complex activates factor X

T/F: It is harder to stop the intrinsic pathway.

TRUE

Explain the formation of fibrin.

• factor Xa converts prothrombin (II) to thrombin (IIa)

• thrombin cleaves fibrinogen (I) into fibrin (Ia)

• factor XIII is activated by thrombin (& Xa)

• factor XIIIa cross-links & stabilizes fibrin

T/F: Cross-linked fibrin is soluble.

FALSE - insoluble

Freshly formed fibrin threads are ________, causing them to adhere to ____________.

sticky

each other, platelets, blood cells, tissues, & foreign substances

Over time, fibrin ________.

contracts 

The contraction of fibrin does what 2 things?

• allows continued blood flow

• brings edges of defect closer together

thrombolysis

removal/ dissolution of fibrin-platelet aggregate (thrombus) → fibrinolysis

What prevents permanent occlusion of the vessel after injury?

balance between formation of the fibrin-platelet aggregate for adequate hemostasis and removal of aggregate to allow for healing and return of normal blood flow

What is responsible for fibrinolysis?

plasminogen

Fibrinolysis is activated by what?

• tissue plasminogen activator (tPA)

• urokinases

• factor XIIa

Fibrinolysis is inactivated by what?

• a2-antiplasmin

• a2-macroglobin

Fibrinolysis is regulated by what?

• thrombin activatable fibrinolysis inhibitor (TAFI)

• plasminogen activator inhibitors (PAIs)

T/F: Coagulation factors are only activated when needed.

FALSE - continuously activated, primed for rapid response

What helps limit the hemostatic reactions?

proteins that inhibit or degrade CoAg factors that circulate in the plasma & are released at sites of vascular injury

Coagulation regulation is the balance between which 3 pathways?

1. hemostatic

2. fibrinolytic

3. anticoagulant

How does blood flow contribute to the regulation of coagulation?

dilution → molecules removed from the site and from circulation by the liver and spleen

What is the most potent and most significant coagulation inhibitor?

antithrombin III

What are the other anticoagulant molecules?

• heparan sulfate 

• tissue factor pathway inhibitor

• protein C - protein S - thrombomodulin system

Protein C and protein S are _________ dependent.

vitamin K

What is the function of the anticoagulant system?

inactivate or inhibit factors involved in coagulation

Wha are the endothelial surface members of the anticoagulant system?

• heparan sulfate

• thrombomodulin

What are the circulating members of the anticoagulant system?

• antithrombin III

• tissue factor pathway inhibitor

• protein C

• protein S

What is the end result of the anticoagulant system?

conversion of thrombin (IIa) from procoagulant to anticoagulant

Which factors are inhibited as a result of the anticoagulant system?

Xa, VIIa, VIIIa, V

Regulation of coagulation requires the integration between pathways. An example of this is thrombin. Explain how thrombin can be both a procoagulant and anticoagulant.

procoagulant:

• cleaves fibrinogen (I) to fibrin (Ia)

• activates factors V, VIII, XI, and XIII

• activates platelets

anticoagulant:

• high concentrations destroys factors V and VIII

• activates protein C when bound to thrombomodulin

A prothrombotic event is also ___________.

proinflammatory

T/F: There is no overlap between coagulation and inflammation.

FALSE - large overlaps → complement cascade