HEMOSTASIS Haemostasis or Hemostasis (Greek: aimstasis, from ama "blood" + stsis "stagnation") Functions of Hemostasis

flow through the injured vessels

exaggerated or impaired it will cause either thrombosis or hemorrhage respectively; BLEEDING CLOTTING Hemostatic Process HEMOSTASIS is a balance between thrombosis and hemorrhage COMPONENTS of HEMOSTATIC SYSTEM 1. ENDOTHELIAL CELLS 2. PLATELETS 3. CLOTTING FACTORS 1) INTACT ENDOTHELIAL CELLS SMOOTH surface not conducive to clot formation a layer of glycocalyx on the endotheliumthat repels clotting factors and platelets display membrane proteins (thrombomodulin) that inhibit clotting store von Willebrand factor in cytoplasmic granules make prostacyclin ~ inhibits platelet aggregation SUBENDOTHELIAL CELLS extracellularmatrix proteins (collagen)that normally do not contact blood Willebrand factor (vWF) that binds to both platelet receptors and collagen/subendothelial cells 2. PLATELETS -10 days) enucleated cells that are fragments of larger progenitor cells called megakarocytes thrombopoietin and other growth factors platelets released from the bone marrow stay in the circulation; sequestered in a splenic pool that is freely exchangeable with circulating platelets Platelet specific granules 1. dense granules : ADP/ATP, serotonin, calcium. 2. alpha granules contain: a. adhesive proteins (von Willebrand factor, fibronectin) b. coagulant proteins (factor V, fibrinogen) c. growth factors/selectins (PDGF, P-selectin)

3. Plasma coagulation factors Nomenclature ulation factors have various names but an internationally standardized nomenclature system is using Roman numeral designations. A lower case a indicates the active factor (e.g. factor IXa) in the plasma except factors III & IV. of reaction in the coagulation system Coagulation Factors

-Prower Factor II Hageman Factor PROENZYMES are each activated by proteolytic cleavage to become enzymatically active. The activated factors are denoted by the subscript "a". The enzymes of the coagulation cascade all belong to the family of serine proteases exemplified by trypsin. Vitamin K dependent clotting factors - vitamin K dependent COFACTORS r the optimal activity of the enzymes of the clotting cascade

EVENTS IN HEMOSTASIS HEMOSTASIS: 3 phase process I. Primary hemostasis

II. Secondary hemostasis

III. Tertiary hemostasis

PRIMARY HEMOSTASIS I. VASCULAR CONSTRICTION traumatized, the greater the degree of spasm. This local vascular spasm can last for many minutes or even hours. VASCULAR CONSTRICTION FACTORS The contraction results from nervous reflexes, local myogenic spasm, and local humoral factors from the traumatized tissues and blood platelets. originate from the traumatized vessel or from nearby tissues. myogenic contraction of the blood vessels initiated by direct damage to the vascular wall.

substance thromboxane A2. Vasoconstriction: Serotonin ---- constricts blood vessels Thromboxane A2 --- is a powerful vasoconstrictor and platelet aggregating agent II. FORMATION of PLATELET - Platelet adhesion - Platelet secretion - platelet aggregation FORMATION OF THE PLATELET PLUG A. Platelet adhesion surface: sub endothelial collagen a disc shape to a slightly broader, plate like form to increase surface area FORMATION OF THE PLATELET When platelets come in contact with a damaged vascular surface, such as the collagen fibers in the vascular wall or damaged endothelial cells, they immediately change their characteristics. PLUG Platelet Changes: 1) begin to swell 2) assume irregular forms with pseudopods 3) their contractile proteins contract forcefully and cause the release of granules that contain multiple active factors; 4) they become sticky so that they stick to the collagen fibers; 5) they secrete large quantities of ADP; 6) their enzymes form thromboxane A2 (secreted into the blood). FORMATION OF THE PLATELET PLUG A. Platelet adhesion adhesion. Thrombin Fibronectin vWF - is the largest component of factor VIII and secreted by platelets and by vascular endothelial cells. B. Platelet secretion (platelet release reaction) -> stimulates aggregation Platelets contain 3 types of secretory granules: -granules : - platelet specific proteins (Plt factor 4, -thromboglobulin) - Platelet derived growth factor - coagulation proteins found in plasma (fibrinogen & von Willebrands factor) granules containing ATP, ADP, Calcium & serotonin C. Platelet aggregation END RESULT: PLATELET PLUG

SECONDARY HEMOSTASIS Clotting takes place in 3 essential steps: 1. Formation of a complex of activated substances: prothrombin activator in response to Rupture of the vessel Damage to the blood itself 2. Prothrombin activator catalyzes the conversion of prothrombin to thrombin. 3. Thrombin acts as an enzyme to convert fibrinogen into fibrin fibers that enmesh (trap) platelets, blood cells & plasma to form the clot. I. How is prothrombin activator is formed? formed in two ways, which actually interact with each other. 1. Extrinsic Pathway: Begins with trauma to the vascular wall & surrounding tissue 2. Intrinsic Pathway: Begins with trauma to the blood itself or exposure of the blood to collagen of traumatized blood vessel wall. In both pathways , series of different plasma proteins blood clotting factors play major role. Most of these are inactive forms of proteolytic enzymes. The process of coagulation involves a cascade of reactions i.e. activation of one factor leads to activation of next. EXTRINSIC PATHWAY Begins with trauma to the vascular wall or surrounding tissue. This occurs in 3 steps: 1. Release of tissue factor: Traumatized tissue releases a complex of several substances: Tissue factor/ Tissue thromboplastin/ Factor III . This is composed of phospholipids + lipoprotein complex. 2. Activation of factor X: Tissue factor + factor VII (Stable factor) complexes and in presence of Ca,activates factor X (Stuart-Prower factor) to form Xa. 3. Formation of prothrombin activator: Xa (Activated factor X) + phospholipids released from tissue or platelets + factor V (Labile factor) forms a complex called prothrombin activator. ++

Intrinsic Pathway orexposure of the blood to the collagen cascading reactions.

1. Activation of factor XII & platelets 2. Activation of factor XI 3. Activation of factor IX 4. Activation of factor X: by IXa in presence of Ca++, VIIIa , phospholipids 5. Formation of Prothrombin Activator

DIFFERENCE BETWEEN EXTRINSIC AND INTRINSIC PATHWAYS Extrinsic pathway ature by the amount of tissue factor released from the traumatized tissues and by the quantities of Factors X,VII, and V in the blood ,clotting can occur in as little as 15 seconds Intrinsic pathway proceed, usually requiring 1 to 6minutes to cause clotting Calcium ions of almost all blood clotting reactions (intrinsic mechanism)

II. Conversion of prothrombin to thrombin converts prothrombin to thrombin in presence of ionic Ca++. occurs at the surface of platelets that has formed platelet plug at the site of injury. fibrinogen into fibrin fibers in 10-15 sec. formation of prothrombin activator as nextsteps occurs rapidly to form clot. Conversion of fibrinogen to fibrin: thread.

FSF) which is activated by thrombin acts as an enzyme to convert weak non-covalent hydrogen bonding to strong covalent bonds between fibrin monomer & also between adjacent fibrin fibers.

Coagulation Pathway

Roles played by Thrombin VIII, V & XIII. bin. (Amplification Effect)

TERTIARY HEMOSTASIS Clot retraction- SERUM After forming, blood clot retracts and pulls the edges of a broken vessel together ( role of thrombasthenin, actin and myosin) results to extrusion of serum ( lacks fibrinogen and clotting factors) FIBROUS ORGANIZATION (DISSOLUTION OF THE CLOT) Once a blood clot has formed, it can follow one of two courses: 1) it can become invaded by fibroblasts, which subsequently form connective tissue all through the clot 2) it can dissolve. FIBROUS ORGANIZATION (DISSOLUTION OF THE CLOT) hours after the clot is formed. This continues to complete organization of the clot into fibrous tissue within about 1 to 2 weeks. larger clot, such as blood that has leaked into tissues, special substances within the clot itself usually become activated, and these thenfunction as enzymes to dissolve the clot. LYSIS OF BLOOD CLOTS: PLASMIN SYSTEM

plasminogen is trapped in the clot along with other plasma proteins. This will not become plasmin or cause lysis of the clot until it is activated. slowly release a powerful activator called tissue plasminogen activator (t-PA) , so after the clot has stopped the bleeding, it eventually converts plasminogen to plasmin and removes the clot.(fibrinolysis) flow has been blocked by clots are reopened by this mechanism.

Why blood does not clot in circulation? ENDOGENOUS ANTICOAGULANTS Physiologic inhibitors of coagulation

1. Antithrombin III (AT-III) and inactivates it, preventing thrombin from converting fibrinogen to fibrin binds to activated factors (factors IXa, Xa, XIa and XIIa) rendering them inactive presence of Heparin (either naturally released from basophils or given therapeutically as an anticoagulant) -III levels are subject to an increased risk of thromboembolism even in cases of slightly reduced AT-III levels. 2. Protein C

Protein C anticoagulant functions is achieved by Protein S. Protein C and/or Protein S deficiencies have a thrombotic tendency. Protein C and Protein S with liver disease and disseminated intravascular coagulation (DIC).

3. Protein S

Acts as a cofactor to Protein C to enhance its ability to degrade factors V and VIII

4. THROMBOMODULIN Thrombomodulin thrombin complex activates PROTEIN C which inactivates FACTORS V and VIII
2

5. PROSTACYCLIN -aggregate action adjacentvessel

Natural anticoagulants effects of antithrombin III (together they inhibit IX, X, XI, XII and thrombin) ithromboplastin (inhibits tissue factors tissue thromboplastins) HEPARIN capillaries of the lungs and to a lesser extent in the capillaries of the liver. uantities of heparin might be needed in these areasbecause the capillaries of the lungs and liver receive many embolic clots formed in the slowly flowing venous blood; sufficient formation of heparin prevents further growth of the clots. ABNORMALITIES OF HEMOSTASIS CAUSES OF EXCESSIVE BLEEDING 1. Vitamin K Deficiency 2. Hemophilia 3. Thrombocytopenia Vitamin K deficiency number of platelets

Thrombocytopenia Hemorrhagic spots (petechiae) - petechiae - decreased production (toxins, radiation, infection, leukemias) - increased destruction (autoimmune processes) - increased PLTs consumption (DIC) when PLTs<50,000/L -400,000/L Disseminated intravascular coagulation (DIC) blood vessels sis, and depletion of coagulating factors generalized bleeding

- bacterial infections (endothelial damage) - disseminated cancers (release of procoagulants) - complications of pregnancy - severe catabolic states HEMOPHILIA

trauma

Thromboembolic condition blood vessel Causes: 1. Any roughened endothelial surface due to arteriosclerosis, infection or trauma. 2. Slow blood flow Use of t- PA or streptokinase in treating intravascular clot. ANTI-COAGULANTS FOR CLINICAL USE A. Heparin B. Coumarins: (e.g. warfarin) competes with Vit K C. Aspirins A. HEPARIN AS THERAPEUTIC ANTICOAGULANT antithrombin III, this increases effectiveness of antithrombin III in removing thrombin antithrombin III removes several other activated coagulation factors in addition to thrombin, further enhancing the effectiveness of anticoagulation. The others include activated Factors XII, XI,IX, and X.

III to inactivate Thrombin IIa

B. Coumadin (Warfarin, Dicoumarol)

factors (II, VII, IX, X) (Protein C & S) first) C. Aspirin the platelet enzyme cyclooxygenase, which is needed for proper platelet aggregation (platelet adhesion is unaffected) at least seven days tests should avoid aspirin ingestion for at least seven days Tests of coagulation Hemostasis Laboratory Tests to Evaluate

PLATELET COUNT

BLEEDING TIME eding; hemostatic process - 6 min Abnormal Bleeding Time

platelet function defect (see platelet aggregation)

salicylates (including aspirin). CLOTTING TIME test tube - 12 min PROTHROMBIN TIME Blood removed is immediately oxalated and Ca++ & tissue factor are added The time taken for coagulation is known as prothrombin time. evaluates the EXTRINSIC pathway

Activated Partial Thromboplastin Time (APTT) lipid substance and are added to the plasma sample; fibrinogen -35 secs common case of prolonged aPTT heparin!!! PROTHROMBIN TIME The normal PT is 11-15 secs. Prolonged PT : a deficiency in any of factors VII, X, V, prothrombin (factor II), or fibrinogen (factor I) The extrinsic factors not measured in the PT test are Factors III (Thromboplastin), and IV (Calcium). The PT is also used to monitor oral anticoagulant therapy such as warfarin. PROTHROMBIN TIME

1. vitamin K deficiency (vitamin K is a co- factor in the synthesis of functional factors II (prothrombin), VII, IX and X) 2. liver disease 3. Warfarin therapy (coumadin) 4. DIC 5. excesive heparin THROMBIN TIME primarily measures the level of fibrinogen -15 sec Prolonged TT: parin (much more sensitive to heparin than aPTT)

NORMAL PT PROLONGED aPTT factor VIII, IX, XI or XII deficiency INTRINSIC PATHWAY

PROLONGED PT PROLONGED aPTT factor X, V, II or I (fibrinogen) deficiency Common pathway

PROLONGED PT NORMAL aPTT factor VII deficiency ; factor XIII deficiency Extrinsic pathway