16/17 Hemostasis

Definition of homeostasis

• Control of bleeding following injury

• Dissolution of clot following tissue repair

• If balance is not maintained bleeding or clotting to death can occur

• Excess bleeding- trauma, major surgery, hemophilia

• Clotting- stroke, MI, thrombosis

Major systems involved in hemostasis

• Vascular- lumen is lined with flattened endothelial cells which become thrombogenic when damaged

• Platelets- part of primary haemostatic plug

• Coagulation- cascade of enzymes that enable formation of fibrin clot

• Fibrinolytic- clot dissolution

Minor systems involved in hemostasis

• Kinin- family of plasma proteins that activate vasodilation, blood coagulation and fibrinolysis

• Complement- activates inflammation, opsonisation, direct killing of microbes by lysis

• Serine protease inhibitors- serine proteases cleave peptide bonds, inhibitors regulate coagulation cascade e.g. anti-thrombin

Four main processes in coagulation

• Constriction of blood vessels which limits blood flow to area

• Platelet activation by thrombin leads to formation of platelet plug

• Formation of fibrin clot

• Dissolution of clot by plasmin (fibrinolysis)

Primary hemostasis

• Constriction of injured vessels

• Exposure of subendothelial collagen

• Adhesion and aggregation of blood platelets on damaged surface

• Formation of hemostatic plug

Secondary and tertiary hemostasis

• Formation of thrombin catalysed by the surface of activated platelets

• Formation of thrombin via activation of factor 7 by tissue factor (3)

• Conversion of fibrinogen to fibrin catalysed by thrombin

• Formation and stabilisation of fibrin clot

• Tertiary- clot dissolution

Structure of blood vessels

• Smooth muscle - vasoconstriction

• Connective tissue contains collagen for platelet activation, more in veins

• Walls have a thin layer of endothelium that is non-thrombogenic to ensure smooth blood flow and stop unwanted thrombosis

• Lumen inside where blood flows

Role of blood vessels in hemostasis

• Vasoconstriction minimises blood loss

• Diverts blood from injured vessel to intact ones

• Collagen fibers become exposed when endothelium is damaged which causes thromboplastin and von Willebrand factor to leak from the cells

• Platelets activate after coming into contact with collagen fibers and vWF

• Coagulation system activated when blood proteins contact thromboplastin (tissue factor / F3)

Structural functions of platelets

• Cellular fragments of megakaryocytes

• Alpha granules contain adhesive proteins, coagulation and fibrinolytic factors, growth factors, cytokines, chemokines (recruit to damaged area)

• Dense granules contain ADP/ATP for recruitment of more platelets, Ca++ for platelet activation and fibrinogen attachment, serotonin for vasoconstriction, histamine for platelet aggregation

• Surface connected tubular system for secretion of coagulation factors and rapid uptake of calcium

• Surface receptors specific for vWF and fibrinogen

Role of platelets in hemostasis

• Adhesion by exposure to subendothelial connective tissue which contains collagen fibers and vWF, platelets adhere to damaged endothelium

• Shape changes from normal round / oval shape to irregular with many projection after encountering collagen fibres, irreversible once prolonged stimulation causes degranulation

• Release of alpha and dense granules via tubular system

• Aggregation once fibrinogen binds to platelet receptors, also induced by ADP, collagen, thrombin, vWF

• Results in primary hemostatic plug which is fragile, conversion of fibrinogen to fibrin stabilises it (facilitated by thrombin)

Coagulation cascade

• Generates thrombin, converts soluble fibrinogen to fibrin

• Enmeshes platelet aggregates at the site of vascular injury which creates a more stable hemostatic plug

• Reaction occur on exposed collagen

Coagulation factors

• Plasma proteins - most are serine proteases except fibrinogen, F5, F8

• Synthesised in the liver, vWF made by endothelial cells and megakaryocytes

• Vitamin K is necessary for y-carboxylation of some factors (2, 7, 9, 10) which is a post-translational modification that converts the factors to their functional form

• If vitamin K is deficient, the proteins cannot bind calcium and are non-functional

Coagulation factor names

Intrinsic pathway

• Initiated by contact of flowing blood with foreign surface e.g. collagen, phospholiipid

• This activates F12 to F12a which also links intothe fibrinolytic, complement and kinin systems

• Prekallikrein and HMWK enhance activation

• F11 is activated by F12a, F9 is activated by F11a

• F9a combines with F8 in the presense of calcium and phospholipid to activate F10

Extrinsic pathway

• Tissue factor (F3) released from injured cell wall activates F7 to form F7a in presence of calcium

• F3a and TF complex which activates F10 and F9 (both commmon and intrinsic pathways)

• TF pathway inhibitor regulates this complex

• This system allows for quick fibrin production on the surface of injured cells and accelaration of intrinsic pathway

• Both pathways always work together in vivo

Common pathway

• Once factor 10 is activated (F10a) it forms a complex with F5, prothromin (F2), calcium and phospholipid

• This complex assembles on the surface of activated platelets and converts prothrombin to thrombin F2a

• Thrombin converts fibrin to fibrinogen which allows the fibrin strands to polymerise, it also regulates factors 5 and 8

• Thrombin activates F13 to F13a which catalyses the formation of crosslinks between fibrin molecules to stabilise the clot

Fibrinolytic system

• Plasminogen is an inactive form of plasmin which has a high affinity for fibrinogen and fibrin

• Plasminogen activators like tissue plasminogen activator are found in tissues and secretions

• Once plasmin is activated, it digests the clot from the inside to outside

• Fibrinolytic inhibitors e.g. a1-antiplasmin prevent uncontrolled proteolytic activity

Regulatory proteins

• Serine protease inhibitors (serpins) regulate coagulation activity

• Antithrombin is present in plasma and neutralises F10a and thrombin as well as most active serine proteases of the system

• Protein C is a plasma protein that is activated by thrombin and inactivates F5 and F8

• Thrombomodulin is a membrane bound glycoprotein lining vascular endothelium which binds to thrombin and converts it to a less active form, also activates protein C

• Others include heparin co-factor 2, a2-macroglobulin, a1-proteinase inhibitor