CAPS 205 Blood - Lecture 3 PDF

Physiology of Blood 3 Coagulation CAPS 205  Learning Objectives Blood III Outline the maturation of megakaryocytes and the generation of platelets Outline the three components of the hemostatic response and discuss how t...

Physiology of Blood 3 Coagulation CAPS 205  Learning Objectives Blood III Outline the maturation of megakaryocytes and the generation of platelets Outline the three components of the hemostatic response and discuss how these components influence one another Describe the platelet response to vascular injury and explain how the response is isolated to the site of injury Describe the initiation and amplification of the clotting/coagulation cascade Outline the factors involved in the prevention of hemostasis (anticoagulation) Describe the common disorders of hemostasis and consider the complications associated with these disorders 2  Sliced Finger Example You are chopping vegetables to prepare dinner and you accidentally slice your finger. Your finger is still attached but is bleeding. Upon examination you are able to determine that you do not require stitches. Consider the following questions: What steps can you take to minimize your blood loss? Why are they helpful? After ten minutes or so you realize you are no longer bleeding. How did your body respond to prevent continued blood loss? 3  Thrombopoiesis Generation of Platelets The liver hormone, thrombopoietin, stimulates megakaryocyte production in the bone marrow Cell fragments (platelets) break off from the edges of megakaryocytes Platelet formation occurs in the bone marrow or quickly following entry into circulation Platelet characteristics: Disc shaped Enucleate 2-3 μm diameter Pseudopodia allows for shape alteration Contain granules: – Alpha granules – contain coagulation factors, pro-coagulants (e.g. factor V, Megakaryocyte vWF), platelet-derived growth factor, and maturation adhesion molecules – Dense granules – contain ADP, ATP, Ca2+, and serotonin H&S Fig 24-2 4  Hemostasis Overview The normal hemostatic response acts to arrest bleeding following injury to vascular tissue 1. Vasoconstriction 2. Platelet Plug 3. Clot Formation Reaction of blood Primary Secondary H&S Fig 24-1 vessels hemostasis hemostasis 5  Hemostatic Vasoconstriction Vasoconstriction is the immediate response to vascular injury There are two components to this response 1. Local contractile response (vasoconstriction & increased tissue pressure) – Neurogenic spasm A rapid, but short lasting (~60 seconds) response – Myogenic spasm Longer lasting response (20-30 minutes) 2. Release of humoral substances – Serotonin – Released from activated platelets – Endothelins – Released from injured endothelium – Clotting factors – Blood coagulation cascade reactions Both serotonin and endothelins promote vasoconstriction 6  Platelet Plug Formation In response to injury of the vessel wall, platelets adhere to: von Willebrand factor (vWF), mediated by glycoproteins on the platelet surface (GpIb) Exposed collagen fibers in the vascular wall Activated platelets undergo conformational change (pseudopodia formation) and release alpha granules (clotting factor V, fibrinogen, vWF) and dense granules (ADP, ATP, serotonin, Ca2+) ADP and fibrinogen are responsible for the initiation of platelet aggregation ADP stimulates release of thromboxane A2 (from activated platelets), which acts as a potent vasoconstrictor and potentiates platelet aggregation (positive feedback) Results in the formation of an unstable hemostatic (platelet) plug Plug may be sufficient to stop bleeding in small vascular injuries Plug formation is localized due to ADP-induced prostacyclin & NO release G&H Fig 37-1 7  Platelet Activation Resting Platelets: Maintain an active Ca2+ efflux (i.e., low cytosolic Ca2+) Activated Platelets: Reduced Ca2+ efflux due to low cytosolic cAMP resulting from increased platelet TXA2 production (i.e., high cytosolic Ca2+) TXA2 acts both to potentiate platelet aggregation and as a potent vasoconstrictor Martin Fig 1 8  Role of Prostaglandins Prostacyclin & Thromboxane A2 Prostacyclin (PGI2): Inhibition of platelet aggregation in intact vessels COX-1 & COX-2 COX-1 Thromboxane A2 (TXA2): Activation of platelet aggregation in damaged vessels High cAMP levels reduce the free cytosolic Ca2+, preventing aggregation and adhesion Intact vascular endothelial cells H&S Fig 24-8 9  The Coagulation Cascade The overall aim of the coagulation cascade is to create a stable fibrin formation (clot) to strengthen the platelet plug and complete the seal This requires the production of thrombin Thrombin acts on fibrinogen (factor I) to promote fibrin clot formation It is important that coagulation is localized, and this is achieved by an intricate network of amplification and negative feedback loops 2. Intrinsic Xase 1. Extrinsic Xase Thrombin production is dependent 3. Prothrombinase on three enzymes complexes 1. Extrinsic Xase 2. Intrinsic Xase 3. Prothrombinase B&B Fig 18-12 10  Blood Coagulation Extrinsic Pathway Initial response to vascular tissue injury Tissue factor (TF) is exposed following injury to endothelial cells Plasma factor VII binds to TF and is converted to VIIa The TF-VIIa complex then binds Ca2+ and acts to convert X to Xa The TF-VIIa-Ca2+ complex (Xase) also converts a small amount of prothrombin to thrombin, this ensures propagation of the cascading reactions This pathway clots blood that has escaped the vessel into surrounding tissue 11 B&B Fig 18-12  Blood Coagulation Intrinsic Pathway The extrinsic Xase complex (previous slide) also activates factor IX and factor XI The small amount of thrombin activated by the extrinsic pathway acts to convert V and VIII to Va and VIIIa respectively The intrinsic Xase complex (IXa-VIIIa-Ca2+) also acts to convert X to Xa and this, in combination with Va and Ca2+, leads to the activation of prothrombinase (Xa-Va-Ca2+) The prothrombinase complex converts a massive amount of prothrombin to thrombin This pathway clots blood within the injured vessel 12 B&B Fig 18-12  Blood Coagulation Common Pathway Fibrinogen is hydrolyzed by thrombin into fibrin monomers Fibrin monomers spontaneously polymerize into fibrin polymers Thrombin also converts factor XIII to factor XIIIa, which then catalyzes the formation of a cross-linked lattice with covalent cross-bridging (stable fibrin) 13 B&B Fig 18-12, G&H 37-4  Fibrinolysis Clot Breakdown Tissue plasminogen activator (t-PA) is released from endothelial tissues and binds to fibrin t-PA converts clot-bound plasminogen into plasmin to facilitate the break down of stable fibrin into soluble fragments 14 B&B Fig 18-14  Thrombin in Hemostasis Majority of thrombin activation is mediated by prothrombinase (common pathway) Minor amounts of thrombin activation are mediated by – Thromboplastin (tissue factor) from exposed subendothelial extravascular tissues and activated platelets – Extrinsic pathway Xase activation (TF-VIIa-Ca2+ complex) Role of thrombin: Enhancement of platelet aggregation Conversion of fibrinogen to fibrin monomers & polymers Activation of XIIIa to stabilize fibrin mesh Enhancement of VIIIa activation (intrinsic Xase activation) and Va (prothrombinase activation) – feedback enhancement 15  Anticoagulation In intact vessels, coagulation/clotting can result in impaired blood flow Physical factors – Smooth lining and negative charge of intact vessels minimizes platelet adhesion Vasodilators – NO (increases blood flow preventing platelet activation), PGI2 (reduces platelet adhesion and aggregation) Endogenous anticoagulants – Antithrombin III (AT III, inhibits IXa and Xa), heparin/heparan suphate (stimulates AT III), tissue factor pathway inhibitor (TFPI, inhibits extrinsic Xase), thrombomodulin (inhibits thrombin), α-protease inhibitor (inhibits IXa), protein C and its cofactor protein S (inhibit Va and VIIIa) Exogenous anticoagulants – Heparin (clinical use: prevention of coagulation), Ca2+ chelators (EDTA, oxalate, citrate) (clinical use: blood collection) 16  Disorders of Hemostasis Platelet abnormalities – Thrombocytopenia – Platelet deficiency – Thrombocytosis – Excess platelets Hyperfibrinolysis – PAI-1 deficiency – Rare, congenital or acquired Coagulation defects – Vitamin K deficiency – Vitamin K is required for synthesis of many clotting factors such as prothrombin, VII, IX, and X, and deficiency of vitamin K results in inadequate production of these factors – Hemophilia – Inherited deficiency of specific coagulation factors, injury can result in uncontrolled bleeding Type A – Deficiency of factor VIII Type B – Deficiency of factor IX – von Willebrand Disease – Inherited deficiency of platelet adhesion (deficiency in factor VIII and vWF) 17  Sliced Finger Example You are chopping vegetables to prepare dinner and you accidentally slice your finger. Your finger is still attached but is bleeding. Upon examination you are able to determine that you do not require stitches. Consider the following questions: What steps can you take to minimize your blood loss? Why are they helpful? – Apply pressure – helps to minimize blood flow to the injured region – Raise your hand above your heart – lower blood pressure in the injured region – Cold water or compress – promotes local vasoconstriction After ten minutes or so you realize you are no longer bleeding. How did your body respond to prevent continued blood loss? – Initial vasoconstriction/increased tissue pressure response – Formation of plug by platelet aggregation – Formation of stable clot (stable fibrin deposition ~60 seconds, fully formed stable clot ~3-10 minutes) 18  Concept Check Practice Question Hemophilia Example Jacob suffers from type A (classic) hemophilia, which is characterized by a deficiency of factor VIII. He accidentally slices his finger while chopping vegetables to prepare dinner. His finger is still attached but is bleeding. Upon examination he is able to determine that he does not require stitches. Consider the following questions: What steps can he take to minimize his blood loss? Why are/aren’t they helpful? What outcome would you predict ten minutes following his finger slicing? What about an hour following his finger slicing? Outline the key difference between his response compared to a normal healthy adult. 19  Concept Check Practice Question Hemophilia Example Jacob suffers from type A (classic) hemophilia, which is characterized by a deficiency of factor VIII. He accidentally slices his finger while chopping vegetables to prepare dinner. His finger is still attached but is bleeding. Upon examination he is able to determine that he does not require stitches. Consider the following questions: What steps can he take to minimize his blood loss? Why are/aren’t they helpful? – The same steps can be taken (given the cut is minor) as the previous example (normal healthy adult) – Vasoconstriction and reduced blood pressure will aid in platelet plug formation What outcome would you predict ten minutes following his finger slicing? – With minimal/no factor VIII the function of the intrinsic tenase will be seriously impaired – The extrinsic pathway remains intact, and will be activated following the vessel injury, but will rapidly be inactivated by tissue factor pathway inhibitor (TFPI), so only a small amount of thrombin will be produced – There will be insufficient thrombin to allow stable fibrin clot formation, so if pressure is removed and/or blood pressure increases the platelet plug can easily be dislodge and bleeding will continue 20  Concept Check Practice Question Hemophilia Example Consider the following questions: What about an hour following his finger slicing? – If this is a minor cut (doesn’t require clot formation), the bleeding will most likely have stopped, but caution is still important as to not reopen the injury – If the cut is more severe, factor replacement therapy (treatment with clotting factor) may be required Outline the key difference between his response compared to a normal healthy adult. – Both the local vasoconstriction and platelet plug formation will be the same in this example – The extrinsic response will occur normally, but the intrinsic pathway is impaired and because of the rapid inactivation of the extrinsic pathway, the common pathway activation will be greatly reduced – Clot formation will be greatly reduced/absent, and this will lead to a prolonged bleeding time Many severe hemophilia patients undergo prophylactic treatment to increase factor VIII availability. There are also alternative therapies available (Emicizumab) that bypass factor VIII and increase activation of factor X directly. Clinical trials are currently underway for gene-based therapy to maintain factor VIII levels. Minor cuts are not the major concern, rather spontaneous bleeds, joint bleeds/damage/arthritis, muscle bleeds, post-trauma bleeds, intracranial hemorrhage 21  Figures B&B: Boron & Boulpaep 3rd Ed. 2017 G&H: Guyton & Hall 14th Ed. 2021 H&S: Hoffbrand & Steensma 8th Ed. 2020 Martin: Martin JF, Kristensen SD, Mathur A, Grove EL, Choudry FA. (2012) The causal role of megakaryocyte – platelet hyperactivity in acute coronary syndromes. Nature Reviews Cardiology 9: 658-670 doi:10.1038/nrcardio.2012.131 22