Haemostasis: A Question of Balance

Daniel Bernhard Breuninger 2019 Haemostasis: A Question of Balance (Daniel Bernhard Breuninger) Haemostasis is a fundamental phenomenon that balances between coagulation and anti-coagulation factors in the blood and so maintains both blood flow and clot formation. The haemostatic process involves a complex interaction between primary haemostatic mechanisms of the vasculature and platelets and the secondary haemostatic response of the extrinsic and intrinsic coagulation cascades. Disruptions to the haemostatic balance result in haemostatic disorders, which manifest themselves in anatomical or mucocutaneous bleeding. These have severe consequences for the affected person. Common disease states that are often found in children include haemophilia A and B (anatomical bleeding which causes haemoarthrosis by bleeding into surrounding joints) and Von Willebrand disease (easy bruising and epistaxis). Both these disease states, especially haemophilia, are extremely painful. Yet they are usually the result of deficiencies in just one factor of the haemostatic system. Therefore, an evaluation of the different mechanisms involved in haemostasis is essential so that adequate treatment of these disorders can be achieved. Introduction Haemostasis is a collaboration between various processes which, especially in response to external factors such as blood vessel damage, ensure the balance of the haemostatic system. These processes include the vascular response, the platelet response, coagulation, and fibrinolysis, which are all highly interconnected. Their collective action allows the haemostatic system to respond effectively to an innumerable variety of disturbances. In response to blood vessel damage, the first-response mechanism of the haemostatic system is primary haemostasis (Golebiewska and Poole, 2015), while in cases of major bleeding it is reinforced by secondary haemostasis though the formation of a fibrin clot (Undas, 2014). Because dysfunction in one of these mechanisms can lead to a state of disease (Peyvandi, Garagiola, and Young, 2016), it is essential to have a thorough understanding of the normal haemostatic processes in order to effectively treat haemolytic disorders. Discussion Overview of the haemostatic balance between pro-coagulation and anti-coagulation Daniel Bernhard Breuninger 2019 Haemostasis is the phenomenon in which the balance between coagulation and anticoagulation factors in the blood is maintained at a dynamic equilibrium, that is, at a level that is of greatest effectiveness in preventing the two extremes of either bleeding or thrombosis. The vascular intima normally prevents coagulation via the secretion of anticoagulants that collectively maintain blood flow through the vessel; these include nitric oxide, prostacyclin, thrombomodulin and heparin sulphate, tissue plasminogen activator as well as tissue factor (TF) pathway inhibitor. Pro-coagulation is assisted by processes which occur when blood cells are damaged. These involve primary haemostasis (vasoconstriction and platelet activation) and secondary haemostasis (extrinsic pathway and intrinsic coagulation pathway for fibrin clot formation). In response to blood vessel damage, the first-response mechanism of the haemostatic system is primary haemostasis, while in cases of major bleeding is reinforced by secondary haemostasis though the formation of a fibrin clot. Primary haemostasis aims to plug the damaged site of the blood vessel and involves the first two processes, that is the vascular and platelet response. When damaged, the immediate response of the blood vessel is constriction via the release of endothelin and platelets adhere to the exposed subendothelial tissue. Meanwhile, nitric oxide counteracts vasoconstriction in undamaged blood vessels, while prostacyclin prevents any unnecessary activation of platelets in blood vessels that are intact. The extrinsic coagulation cascade of secondary haemostasis is activated, as a result of blood vessel damage, by the exposure of tissue factor-expressing, sub-endothelial cells. Conversely, TF pathway inhibitor acts as an anti-coagulant, controlling the extrinsic coagulation pathway. Both the extrinsic and intrinsic coagulation cascades converge onto the common pathway to form thrombin. Thrombin formation in turn is inhibited by thrombomodulin and heparin sulphate. The coagulation cascades result in the formation of a fibrin clot. This is haemostatically counteracted by tissue plasminogen activator which promotes fibrinolysis. The processes of coagulation will be discussed in more detail in the following sections. Primary haemostasis and the laboratory assessment of primary haemostasis The first-aid response mechanism to blood vessel damage is primary haemostasis that involves its two major processes of the vascular vasoconstrictor response and that of the platelet response. The primary objective of the haemostatic response is to plug the site of vessel damage as quickly and efficiently as possible to prevent haemorrhage. To slow Daniel Bernhard Breuninger 2019 haemorrhage, the initial response of the damaged vasculature is constriction through the release of the polypeptide endothelin from the damaged endothelium. Constriction of the vessel reduces the vessel radius and so is capable of significantly reducing blood flow to a fraction of its original rate, thereby reducing the pressure exerted on the damaged site and limiting bleeding as the initial first-aid measure. The second step in the primary haemostatic response is that of the platelet response in its binding to the exposed collagen of the damaged site and the formation of a plug. However, the binding of the circulating platelets to the damaged endothelium requires a reduction of the shear forces exerted on them, thus, the vasoconstrictor is essential in its role in the reduction of pressure to allow the platelet response to be activated, highlighting their interdependency. The activation of platelet response is aided by the release of von Willebrand Factor (vWF) from the surrounding endothelial cells which accommodates the firm adherence of the platelets to the collagen. The released vWF is a multimeric chain that is cleaved into smaller fragments by the proteolytic enzyme ADAMTS13 that is present in the plasma. Under the high shearing-forces of blood vessels where vWF is essential, the tightly folded protein elongates and exposes itself to the cleavage of the proteolytic enzyme. The cleaved fragments of vWF bind to the exposed collagen of the damaged site and act as an anchoring point for platelets to bind. Once bound, the once discoid shaped platelets, become spherical with pseudopod extensions that make them sticky. The platelets aggregate and are bound to each other via intercellular bridges of fibrinogen. Furthermore, platelets once adhered, secrete their granular contents of fibrinogen, factor five, vWF, and thromboxane, which activate and recruit more platelets to the site to seal the break in the endothelium. The assessment of the primary haemostatic response is not straightforward. While simple measurements of platelet concentrations are easily achieved via the use of a full blood count (FBC), this only gives an indication of the number of platelets in the blood yet gives no insight into their function. A blood smear test can give great insight into this. As a defect in primary haemostasis is often associated with thrombocytopenia (low platelet count) or von Willebrand Disease (vWD), a more complex assessment can be done via the vWF antigen test that measures the amount of vWF protein present in the blood, while vWF activity (Ristocetin Cofactor test) determines the protein function (Geisen et al, 2014). Finally, bleeding time shows whether there is an adequate clotting response to vessel injury. In the past, an incision was made prior to surgery and time taken for bleeding to stop evaluated. However, this is an outdated practice. Daniel Bernhard Breuninger 2019 Secondary haemostasis and the laboratory assessment of secondary haemostasis The primary haemostatic response, in and of itself, is insufficient to control major haemorrhage and so must be reinforced by the formation of a fibrin blood clot. Circulating protein-cleaving serine proteases generally circulate as inactive zymogens. However, when these are activated, they can initiate a cascade of events, called the coagulation cascade, that converts the fibrinogen of the plasma into a localised fibrin clot. There are two separate coagulation cascades, the extrinsic and intrinsic, that converge onto the same common pathway. Subendothelial cells, expressing tissue factor, are exposed to plasma upon blood vessel damage. Tissue factor (TF) or thromboplastin, when exposed to plasma coagulants due to a haemorrhaging vessel, is bound to factor seven in the presence of phospholipid and calcium ion (Ca2+). This activates factor seven which in turn activates the factor ten of the common pathway. The laboratory screening test for the extrinsic coagulation cascade is called the prothrombin time. The test functions by measuring the time taken for clot formation in a specimen of anti-coagulated plasma after the addition of phospholipids and thromboplastin. Hageman factor (factor twelve), the protein-cleaving yet inactive zymogen that circulates the plasma, is changed to its active form when it comes into contact with activated platelets and when in the presence of negatively charged phospholipid. The contact activation system (CAS) are the three proteins of prekallikrein (PK), high molecular weight kininogen (HMWK) and the already mentioned factor twelve. The CAS is activated upon the binding of factor twelve, resulting in the reciprocal activation of factor twelve and PK. The latter facilitates inflammation through its cleavage of HK, to release bradykinin, while the former, more importantly, initiates the coagulation cascade through its cleavage and resulting activation of factor eleven. This in turn, acts on factor nine. Factor nine requires the cofactor factor eight, to form a complex that can convert factor ten to its activated state. Inactivated factor eight is bound to vWF and circulates the bloodstream until, in response to injury, it separates into its active state to form the before-mentioned complex. Once factor ten is activated, the common pathway is initiated. The laboratory screening test for this pathway is the activated partial thromboplastin time (APTT). The test involves the addition of a contact activator to a plasma specimen in the presence of Ca2+ and phospholipid. The time taken for clot formation is the measure for the effectiveness of the intrinsic pathway. Daniel Bernhard Breuninger 2019 The activated factor ten, forms a complex with cofactor five; the prothrombin activator complex. Thus, prothrombin is activated to form thrombin. The circulating soluble plasma glycoprotein, fibrinogen, is then cleaved by thrombin to form fibrin. Fibrin is essential in clot formation due to the monomer’s high affinity for neighbouring fibrin that results in the spontaneous polymerisation of fibrin to form a clot. Activated factor thirteen serves as a catalyst in the formation of covalent bonds between adjacent fibrin polymers to form a fibrin network that is structurally sound and insoluble. Fibronectin and plasminogen are also incorporated into the structure. The first serves as an adhesive molecule, while the latter, when converted into plasmin, is able to degrade the clot in fibrinolysis. Fibrinolysis and the laboratory assessment of fibrinolysis Finally, the degradation of the fibrin clot, fibrinolysis, being part of the healing process, occurs only hours after the initial polymerisation of fibrin. Tissue plasminogen activator is released by endothelial cells during states of hypoxia. Fibrinolysis is triggered by tissue plasminogen activator (TPA) that is integrated initially, together with plasminogen, into the clot upon its formation. TPA binds to plasminogen and converts it into the active enzyme of plasmin. This enzyme degrades fibrin and so the clot is degraded from the inside out. Evaluating the fibrinolytic pathway is important in testing for fibrinolysis. As yet, there are no straightforward measurements of these components and further development is needed. To date, the most common fibrinolytic activity test, is the D-dimer test that measures the amount of degradation product of fibrinogen (Crawford et al, 2016). General overview of haemorrhagic disorders and a table illustrating specific examples of the different types of haemorrhagic disorders. Haemorrhagic Disorder Haemophilia A Aetiology Clinical Consequences Testing Treatment Inherited, Factor VIII deficiency (intrinsic pathway), X-chromosome linked disease Prolonged APTT, PT is normal, Factor VII assay Factor VIII supplementati on, desmopressin Haemophilia B Inherited, Factor IX deficiency Prolonged APTT, PT is normal, Factor XI assay necessary Factor XI administratio n Haemophilia (Acquired) Acquired via development of antibody to factor VIII Anatomical bleeding into joints (haemoarthrosis) and muscles leads to severe inflammation and pain Anatomical bleeding, symptoms as haemophilia A, reduced thrombin production, soft-tissue bleeding Anatomical bleeding, symptoms as haemophilia A Prolonged APTT, PT is normal, Factor VIII assay Immunosuppr ession Daniel Bernhard Breuninger 2019 Von Willebrand Disease (Type 1) Inherited, quantitative vWF deficiency, most common type Mucocutaneous bleeding, epistaxis, menorrhagia, haematemesis, increased bleeding tendency Mucocutaneous bleeding, symptoms as VWD (Type 1) Von Willebrand Disease Type 2 Von Willebrand Disease Type 3 Inherited, qualitative abnormality Inherited, vWF and factor VIII are reduced or absent Mucocutaneous and anatomical bleeding, soft-tissue bleeding, internal bleeding Von Willebrand Disease Acquired Acquired, various genetic mutations, strong association with other disease states (e.g. hyperthyroidism, congenital heart disease) Mucocutaneous bleeding, symptoms as VWD (Type 1) vWF assay, FBC, normal PT, may have prolonged APTT Biostate, antifibrinolyti cs, DDAVP and oestrogen vWF assay, FBC, normal PT, may have prolonged APTT vWF assay, factor VIII assay Biostate, antifibrinolyti cs, DDAVP and oestrogen Biostate, antifibrinolyti cs, VIII/vWF concentrate infusion Biostate, antifibrinolyti cs, factor VIII/vWF concentrate infusion vWF assay, assessment of underlying disease and bleeding history Bleeding can be either localised, usually as result of trauma, or generalised and may either be mucocutaneous or anatomical. Bleeding that is slow or continuous indicates an underlying coagulation disorder. Mucocutaneous bleeding involves bleeding into areas where skin transitions into mucous membranes. Bleeding into the skin may be made manifest as disseminated bruising or purpura presenting as petechiae, that is small point haemorrhages or ecchymoses. Mucocutaneous bleeding suggests a defect in primary haemostasis, often due to thrombocytopenia or otherwise vWD that can be acquired or inherited. vWD type one, is the most common and is characterised by a quantitative deficiency of vWF. vWD type two is rare and is due to a qualitative deficiency of vWF. vWD type one and two are associated with increased bleeding tendencies, epistaxis, menorrhagia as well as haematemesis. There is a very rare and serious form of vWD, that is type three vWD. In this inherited disorder both vWF and factor eight are either absent or reduced. Symptoms include mucocutaneous bleedings as is typical for vWD, including anatomical bleeding and internal bleeding. vWD type one may also be acquired and there is a strong link to its association with other disorders such as congenital heart disease and hyperthyroidism. Treatment of these is done via factor eight/vWF concentrate infusion. Anatomic bleeding is bleeding into soft tissues and is often seen through recurrent or excessive bleeding during minor trauma in younger individuals. Anatomic haemorrhage is seen in congenital and some acquired defects in secondary haemostasis, such as haemophilia. Younger persons that are diagnosed with haemorrhagia are most likely to have an inherited bleeding disorder, while if seen in older people this can generally be attributed to an acquired Daniel Bernhard Breuninger 2019 defect. The most common inherited anatomical bleeding disorder is haemophilia A, that is most commonly seen in males as it is an X-linked disease. Haemophilia involves bleeding into soft tissues, such as muscles and joints, causing haemoarthrosis and resulting in severe inflammation and pain. Haemophilia A is associated with an abnormality of the intrinsic pathway as it involves a factor eight deficiency. In contrast, the less common haemophilia B, that manifests with the same symptoms, is a factor nine deficiency. Testing for haemophilia can be done by a prothrombin (PT) and an APTT, where the PT should be normal and the APTT prolonged. To distinguish between the types of haemophilia, specific factor eight and nine assays need to be carried out. Furthermore, haemophilia may be also be acquired through the development of factor eight antibodies. This is treated by immunosuppression. Conclusion The haemostatic balance is maintained through the highly interconnected mechanisms of the vascular response, the platelet response, coagulation and fibrinolysis. There is a constant interplay between pro-coagulation and anti-coagulation factors, ensuring that there is steady blood flow while an adequate damage response is upheld. The primary haemostatic response is the first response mechanism to blood vessel damage and involves initial blood vessel vasoconstriction and concomitant platelet blood clot formation. In response to major haemorrhage, the secondary haemostatic mechanism reinforces primary haemostasis through the formation of a fibrin clot via the extrinsic and intrinsic coagulation cascades. Healing finally involves the degradation of the formed fibrin clot through fibrinolysis. There are a variety of haemolytic disorders that significantly disturb the quality of life of the individuals affected. Therefore, a comprehensive understanding of the normal processes underlying haemostasis is essential for effective treatment of the disorders. Daniel Bernhard Breuninger 2019 References Crawford, F., Andras, A., Welch, K., Sheares, K., Keeling, D. and Chappell, F. (2016). Ddimer test for excluding the diagnosis of pulmonary embolism. Cochrane Database of Systematic Reviews, [online] (8). Available at: https://www.researchgate.net/profile/Fay_Crawford/publication/305909808_Ddimer_test_for_excluding_the_diagnosis_of_pulmonary_embolism_Reviews/links/5b9114a2 a6fdcce8a4ca670b/D-dimer-test-for-excluding-the-diagnosis-of-pulmonary-embolismReviews.pdf [Accessed 30 Apr. 2019]. Geisen, U., Zieger, B., Nakamura, L., Weis, A., Heinz, J., Michiels, J. and Heilmann, C. (2014). Comparison of Von Willebrand factor (VWF) activity VWF:Ac with VWF ristocetin cofactor activity VWF:RCo. Thrombosis Research, [online] 134(2), pp.246-250. 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