Hemodynamics Amir Kol DVM, Ph.D, Dip ACVP (Clinical Pathology) UC Davis, School of Veterinary Medicine akol@ucdavis.edu Objectives Understand the mechanisms that promote extravascular fluid accumulation Edema (tissues) Effusion (cavities) Hyperemia vs congestion Hemostasis - understand the cascade of events that lead to blood clotting Primary hemostasis Secondary hemostasis Tertiary hemostasis Cell based vs. cascade models Define hemorrhagic and thrombotic disorders Disseminated intravascular coagulopathy (DIC) Shock definition and pathogenesis Filtration across the endothelium K[(P cap - P int )-σ(π cap π int )] Robins & Cotran. Pathologic Basis of Disease. Fig 4.1, 9 th edition. 1
Filtration across the endothelium K[(P cap - P int )-σ(π cap π int )] Cleaver & Melton. Nature Medicine. 2013 Filtration across the endothelium Increased microvascular permeability Inflammation Histamine, bradykinin, leukotrienes, substance P IL1, TNFα, IFNγ Inflammatory exudate Infectious agents Immune mediated Toxins Filtration across the endothelium Increased hydrostatic pressure Usually passive (i.e. congestion) Systemic Heart (right or left side) failure, fluid overload Local Hepatic fibrosis Venous obstruction Thrombosis, organ volvulus and torsion, compressive mass 2
Filtration across the endothelium Decreased osmotic pressure Protein losing nephropathy Protein loosing enteropathy Decreased hepatic synthesis Severe burns Malnutrition Water intoxication Hemostasis Arteriolar vasoconstriction Primary hemostasis Secondary hemostasis Clot stabilization and resorption Robins & Cotran. Pathologic Basis of Disease. Fig 4.4, 9 th edition. 3
Hemostasis Arteriolar vasoconstriction Primary hemostasis Secondary hemostasis Clot stabilization and resorption Robins & Cotran. Pathologic Basis of Disease. Fig 4.4, 9 th edition. Contributions of Vessels Vessels are anticoagulant in health Anticoagulant activities of normal endothelium. NO, nitric oxide; PGI2, prostacyclin; t-pa, tissue plasminogen activator; vwf, von Willebrand factor. The thrombin receptor is also called a protease-activated receptor (PAR). Robins & Cotran. Pathologic Basis of Disease. Fig 4.10, 9 th edition. Vessel response to injury Neuromechanical transient vasoconstriction Biochemical endothelial cells synthesize and metabolize many factors involved in hemostasis (pro- and anti-coagulant) and inflammation (See Box 2-1 in Zachary & McGavin, 5 th edition page 63) subendothelial collagen plays a key role in activating platelets and the coagulation system 4
Pro-coagulant Endothelial Function Endothelial Injury Endothelial Injury Endothelial Injury Russell JA. N Engl J Med, 2006. Primary hemostasis The final outcome of primary hemostasis is a platelet plug Platelet adhesion Rapid shape change Secretion Recruitment Platelet aggregation Primary hemostasis http://www.eclinpath.com/hemostasis/physiology/primary-hemostasis/print-11/ 5
Primary hemostasis Robins & Cotran. Pathologic Basis of Disease. Fig 4.5, 9 th edition. Primary hemostasis Video 2dry Hemostasis - Coagulation Multiple component backup systems fail safe (redundancy) Amplification coagulation systems anticoagulation systems Checks and balances positive amplification negative feedback loops 6
Key Aspects of Coagulation (Con t) Localized on a surface, does not occur in solution Occurs on phospholipid membranes (mostly platelet surfaces) Confines reactions to selected sites/wounds Requires Calcium Coagulation Factors Small Mostly produced by the liver Activity requires Vitamin K (II, VII, IX, X, protein C) Inactive precursors are made functional by vitamin K-dependent post-translational carboxylation of glutamic acid residues on the molecules Circulate as inactive pro-enzymes until activated Complexed with inhibitors prior to clearance by the liver Most are proteases (think of coagulation as a series of protein cleavages to make inactive enzymes into active enzymes) Primary exception is tissue factor (aka thromboplastin, factor III) Factors V and VIII and fibrinogen are also not proteases Coagulation Overview An amplification system to rapidly form a fibrin clot Rapid production of thrombin, which converts fibrinogen to fibrin: fibrinogen fibrin (soluble) thrombin (insoluble) Thrombin also has other important functions 7
2 schemes to classify coagulation pathways: 1 st Scheme Cascade (waterfall) 2 nd Scheme Cell-Based Model Coagulation System Cascade View Comprised of 3 inter-related pathways Intrinsic Pathway Extrinsic Pathway Common Pathway Common Pathway This scheme is a simplification for ease of understanding and clarification of laboratory testing Intrinsic Pathway Numerous substances XII XIIa XI IIa XIa IX IXa IIa VIII VIIIa Coagulation Cascade (IXa + PF-3 + VIIIa + Ca +2 ) VIIa Extrinsic Pathway Tissue factor (III) and other substances VII (VIIa + Tissue factor + Ca +2 + PF-3 ) Platelet phospholipid (PF-3) is a cofactor for several reactions Common Pathway X IIa V Va Prothrombin (II) Xa (Xa + Va + PF-3 + Ca +2 ) Fibrinogen Thrombin (IIa) Fibrin Cross linked fibrin Calcium (Ca +2 ) is a vital cofactor for several reactions IIa XIIIa XIII 8
Cascade scheme Implies 2 pathways of activation Exposure to tissue factor (TF) Tissue factor is found on exposed fibroblasts, activated endothelial cells, other cells Contact with basement membrane Sees coagulation as sequential activation Limits the role of the surface as passive In fact, cells (endothelial cells, platelets and leukocytes) provide a dynamic and regulatory function in coagulation Represents well how we measure coagulation in the laboratory, less adequate for definition of process in vivo Cell-Based Model of Coagulation Acknowledges central role of platelet surface, tissue factor and Thrombin Suggests that most coagulation is activated by TF which generates a small amount of thrombin which is followed and amplified by loop activation of intrinsic, extrinsic and common pathways Lessens role of contact activation and implies that the intrinsic pathway is secondary after initial activation through TF and thrombin. The intrinsic pathway is essentially amplification after extrinsic pathway generates a small amount thrombin via TF More real, less reflective of how we measure coagulation And now, A video Cell Based Model of Coagulation: 9
Cell-Based Model of Coagulation Initiation: plasma in contact with TF bearing cells due to break in vessel wall, small thrombin signal Amplification: small thrombin signal binds platelets that have adhered to collagen via vwf, enhances platelet activation activates the tenase complex Propagation: assembly of the prothrombinase complex and generation of lots of thrombin Hoffman & Monroe. Thromb Haemost, 2001. Cell-Based Model of Coagulation Initiation: plasma in contact with TF bearing cells due to break in vessel wall, small thrombin signal Amplification: small thrombin signal binds platelets that have adhered to collagen via vwf, enhances platelet activation activates the tenase complex Propagation: assembly of the prothrombinase complex and generation of lots of thrombin James W. Wisler & Richard C. Becker. Nature Reviews Cardiology, 2012. Functions of Thrombin Promotes coagulation Converts fibrinogen to fibrin Activates Factors V, VIII, XI, positive feedback (amplification) Activates Factor XIII strengthens clot by cross-linking fibrin Potent platelet agonist (via PAR receptors) 10
Functions of Thrombin Regulates coagulation: Converts plasminogen to plasmin, which breaks down fibrin clot. Activates Protein C (APC), an inhibitor of Va and VIIIa Stimulates the production of prostacyclin (PGI 2 ) a platelet inhibitor by endothelial cells J. Borrisoff et al. Cardiovascular Research, 2009. Clot Stabilization So, we have platelets Adhesion, aggregation, granule release Coagulation proteins Series of proteolytic cleavages with the end result of cleaving fibrinogen to fibrin Clot stabilization Fibrin monomers are polymerized into long, crosslinked chains through the action of factor XIIIa. 11
Regulators of Hemostasis Inhibitors of Coagulation - Natural Anticoagulants Antithrombin III (ATIII) Protein C TFPI Thrombomodulin Robins & Cotran. Pathologic Basis of Disease. Fig 4.10, 9 th edition. Regulators of Hemostasis ATIII Produced by hepatocytes Inactivates most coagulation enzymes, especially thrombin, IXa, Xa Loss is associated with thrombosis. Loss most commonly occurs through glomerular disease or consumption in DIC Acts at endothelial surface, needs heparin, binds thrombin, inactivates thrombin Heparin and heparan sulfate ENHANCE ATIII activity This is why heparin is used as an anticoagulant! Regulators of Hemostasis Protein C/ Protein S Produced by hepatocytes Vitamin K dependent Along with factors II, VII, IX, X Protein C is activated by thrombin and Protein S to activated protein C (APC) Part of Protein C / Protein S / Thrombomodulin complex Complex inactivates VIIIa and Va Thrombomodulin + thrombin markedly enhances activation of Protein S 12
Protein C and Protein S anticoagulant System: Vitamin K dependent proteins http://themedicalbiochemistrypage.org/blood-coagulation.php The Protein C Anticoagulant Pathway: Thrombin leaves a site of vascular injury binds to its receptor thrombomodulin (TM) on the intact cell surface. As a result, thrombin loses its procoagulant properties and instead becomes a potent activator of protein C. Activated protein C (APC) functions as a circulating anticoagulant, which specifically degrades and inactivates the phospholipid-bound factors Va and VIIIa. This effectively down-regulates the coagulation cascade and limits clot formation to sites of vascular injury. Regulators of Hemostasis Tissue Factor Pathway Inhibitor (TFPI) Produced by endothelial cells, monocytes, macrophages, hepatocytes and platelets Requires Ca +2 TFPI inhibits TF-factor VIIa complex. Therefore inhibits generation of IXa and Xa via TF-VIIa Regulators of Hemostasis Hepatocytes eliminate activated clotting factors and factor / inhibitor complexes Macrophage Phagocytic System removes factors Endothelium degrades activated factors Blood Flow removes factors 13
Major Regulators of Hemostasis XIIa + XIa AT III + Heparin VIIa + Tissue Factor IXa VIIIa Tissue Factor Pathway Inhibitor IIa APC-Protein S Protein C Xa Va IIa (Thrombin) Fibrin AT III + Heparin AT III + Heparin Where we are going Primary hemostasis: formation of platelet plug Secondary hemostasis: stabilization of the platelet plug with cross-linked fibrin Coagulation proteins Inhibitors of Coagulation Tertiary hemostasis: resorption of the clot by fibrinolysis Fibrinolysis Occurs within the developing clots where plasminogen activators and plasminogen bind to fibrin Factor XII and Tissueplasminogen activator 14
Control of fibrinolysis There are also checks & balances on fibrinolysis Robins & Cotran. Pathologic Basis of Disease. Fig 4.9, 9 th edition. Hemorrhagic disorders - Vasculitides Petechial and ecchymotic pattern Underlying etiologies (see Box 10-7 in Zachary & McGavin, 5 th edition pp.562): Viral Bacterial Mycotic Parasitic Immune mediated Hemorrhagic disorders Primary hemostasis Petechial and ecchymotic pattern Thrombocytopenia Decreased production Consumption Destruction Sequestration Thrombocytopathy Primary (vwd, Glanzmann Throbasthenia, Thrombopathy) Secondary (NSAIDs, uremia, E. canis) 15
Hemorrhagic disorders Secondary hemostasis Hereditary Coagulopathies Hemophilia A factor VIII deficiency Hemophilia B factor IX deficiency Other specific factor deficiencies less common and variable bleeding (See Box 2-5 in Zachary & McGavin, 5th edition page 77) Acquired Coagulopathies Rodenticides (Vitamin K antagonism) Liver Dz. PLN and PLE Drugs and toxins Acquired Coagulopathies Vitamin K antagonists (and, rarely, decreased vitamin K availability) In the absence of vitamin K, coagulation factors are secreted without sufficient carboxylation and have inadequate function Rodenticides, coumarin (moldy sweet clover) Malabsorption, others Drugs Liver disease Decreased production of coagulation factors Liver also responsible for removal of activated factors Envenomation Thrombosis Virchow s Triad THROMBOSIS Hypercoagulability 16
Endothelial Injury Viruses CAD1, equine Morbilivirus, herpes virus and Arterivirus, ovine orbivirus, bovine and procine pestvirus Bacteria Salmonella typhimurium, Manhemia haemolytica, Eryspelothrix rhusiopathiae, Haemophilus somnus Fungi Aspergilus, Mucor, Absidia, Rhizopus Nematodes Strongylus vulgaris, Dirofilaria, Spirocerca, Aelorostrongylus, Angiostrongylosis Endothelial Injury Immune mediated vasculitis FIP, Ehrlichia canis, Leismania Toxins Endotoxin, Claviceps Vitamin E/selenium deficiency Faulty intravenous injection Alterations in Blood Flow Local stasis GDV, intestinal torsion and volvulus, varicocele Cardiac disease DCM, HCM etc. Aneurysm Spirocerca lupi, Strongyulus vulgaris, Copper deficiency in pigs Hypovolemia Shock, diarrhea, burns 17
Hypercoagulability Inflammation Platelets hyper-reactivity Diabetes mellitus, nephrotic syndrome, malignant neoplasia, heartworm disease, uremia Antithrombin deficiency PLN, PLE, hepatic disease Metabolic abnormalities Hyperadrenocorticism Infarction Area of ischemic necrosis caused by occlusion of either the arterial supply or the venous drainage. Zachary and McGavin. Pathologic Basis of Veterinary Disease. Infarction Factors That Influence Development of an Infarct: Anatomy of the vascular supply Rate of occlusion Tissue vulnerability to hypoxia Hypoxemia 18
Fate of the Thrombus Zachary and McGavin. Pathologic Basis of Veterinary Disease. Disseminated Intravascular Coagulation - DIC When things go out-of-control Disseminated Intravascular Coagulation - DIC Excessive activation of coagulation, diffuse thrombosis in microcirculation results in consumption of coagulation factors and platelets leads to bleeding Secondary to other diseases Widespread tissue damage, IMHA, pancreatitis, gram negative sepsis, heat stroke, neoplasia, infections (Heartworm, Rickettsia) 19
Shock A state of reduced effective circulating blood volume which results in cellular hypoxia Cardiogenic Hypovolemic Septic (inflammatory) Anaphylactic Neurogenic Pathogenesis of Septic Shock Inflammatory and counter-inflammatory responses Endothelial activation and injury Induction of a procoagulant state Metabolic abnormalities Organ dysfunction Pathogenesis of Septic Shock Robins & Cotran. Pathologic Basis of Disease. Fig 4.20, 9 th edition. 20
Stages of Shock Non-progressive phase Reflex compensation (baroreceptor reflex, catecholamine release, RAS activation, ADH release and sympatheic stimulation Progressive phase Cellular hypoxia Lactic acidosis Blood pooling Irreversible stage Wide spread cellular necrosis Shock Zachary and McGavin. Pathologic Basis of Veterinary Disease. Questions??? 21