Secondary Hemostasis
Factor I – fibrinogen
Describes the activation of a series of coagulation proteins in the plasma, mostly serine proteases, to form a fibrin clot.
Factor II – prothrombin
Factor III – tissue thromboplastin, a crude mixture of tissue factor and phospholipid
Final event is hemostasis is fibrinolysis, the gradual digestion and removal of the fibrin clot as healing occurs.
Factor IV – Ionic Calcium
The numeral VI was asigned to procoagulant that later was determined to be activated factor V
Primary Hemostasis Activated by desquamation and small injuries Involves vascular intima and platelets Rapid, short-lived response Procoagulant substances expose or released by damaged or activated endothelial cells
Secondary Hemostasis Activated by large injuries to blood vessels and surrounding tissues Involves platelets and coagulation system Delayed, long-term response The activator, tissue factor, is exposed on cell membranes
Coagulation System Nomenclature of Procoagulants
Plasma transports at least 16 procoagulants
Phosphatidylserine – platelet phospolipid
Six others are cofactors that bind and stabilize their respective enzymes
Seven additional plasma glycoproteins are controls that regulate the coagulation process
1958 – International Committee for the Standardization of the Nomenclature of Blood Clotting Factors named the plasma procoagulants using Roman Numerals
When procoagulants become activated, a lower case “a” appears behind the numeral
no Roman numeral; collectively called platelet factor 3
Prekallikrein (pre-K) and HMWK have never received Roman numerals because they belong to the kallikrein and kinin system
Physiologic Function of Procoagulants a. Serine proteases
Proteolytic enzymes of the trypsin family
Includes thrombin (IIa); factors VIIa, IXa, Xa, XIa, XIIa; and pre-K
Has reactive seryl amino acid residue in its active site and acts on its substrate by hydrolyzing peptide bonds
Synthesized as inactive zymogen
Activation occurs when the zymogen is cleaved at one or more specific site by the action of another protease
Also called coagulation factor or clotting factors
Eight are enzymes that circulate in an inactive form called zymogens
VI was withdrawn from the naming system
a. Limited in the site of injury b. Cofactor
Tissue factor – a noncirculating constitutive protein of the subendothelium
Bind to its serine protease
When bound, serine proteases gain stability and increased reactivity
c. Remaining components:
Factor XIIIa – transglutaminase that catalyzes the transfer of amino acids among the gamma chains of fibrin polymers; essential to wound healing and tissue integrity
Fibrinogen – primary substrate of thrombin; synthesized in the liver
VWF – participates in platelet adhesion and transport procoagulant factor VIII
Thrombin catalyze the conversion of fibrinogen to fibrin.
Vitamin K-Dependent Prothrombin Group
Prothrombin, Factors VII, IX, X, and the regulatory proteins protein C, S, and Z
Vitamin K catalyzes an essential posttranslational modification of the prothrombin group proteins; gammacarboxylation of amino-terminal glutamic acid
Two short fibrinopeptides are cleaved from the ends of the fibrinogen Aα and Bβ chains, respectively.
Factors V and VIII Some of the thrombin generated during the early stages of coagulation enters a feedback activation loop.
Formation of coagulation complexes
Once activated, these factors enhance the function of FXa and Ixa.
Factor XIII Thrombin proteolytically activates FXIII in a procoagulant reaction that is enhanced by fibrin. Protease activated receptors The majority of thrombin generation occurs on the activated platelet surface.
THROMBIN
The principal enzyme of hemostasis.
It plays a pivotal role in clot promotion and inhibition, and cell signaling, as well as additional processes that influence fibrinolysis and inflammation. These functions involve numerous cleavage reactions, which must be tightly coordinated.
HAEMOSTATIC SUBSTRATES OF THROMBIN
Fibrinogen
Glycoprotein V Thrombin involves proteolysis of glycoprotein GpV, part of the GpIbIX-V complex on circulating platelet surface. This in turn yields hyper-responsive platelets.
Protein C thrombin also participates in interactions/reactions that elicit anticoagulant function.
Thrombin-activated Fibrinolysis Inhibitor the thrombin–thrombomodulin complex also activates the carboxypeptidase, thrombinactivated fibrinolysis inhibitor. Anti-thrombin
by cleaving R393 of the reactive bait loop of antithrombin, thrombin becomes locked in an irreversible complex and then cleared from circulation.
EXTRINSIC PATHWAY
The extrinsic pathway is activated by external trauma that causes blood to escape from the vascular system. This pathway is quicker than the intrinsic pathway. It involves factor VII.
INTRINSIC PATHWAY
The intrinsic pathway is activated by trauma inside the vascular system, and is activated by platelets, exposed endothelium, chemicals, or collagen. This pathway is slower than the extrinsic pathway, but more important. It involves factors XII, XI, IX, VIII.
COMMON PATHWAY
Both pathways meet and finish the pathway of clot production in what is known as the common pathway. The common pathway involves factors I, II, V, and X.
Fibrinolysis System
PLASMIN’S ROLE Activates Fibrinolysis
Activates Intrinsic Coagulation Pathway Interferes with Intrinsic and common pathways. Block Thrombin conversion of fibrinogen to fibrin.
Fibrinolysis, the final stage of coagulation, begins a few hours after fibrin polymerization and cross-linking. Two activators of fibrinolysis, TPA and UPA, are released in response to inflammation and coagulation. There are checks and balances in the formation of clot. Activation of Plasminogen to Plasmin A zymogen known as Plasminogen which is normally present in plasma is converted to Plasmin by the action of specific enzyme called Plasminogen activators (TPA and UPA) Plasminogen is a 92,000 D single-chain glycoprotein possessing five glycosylated loops called kringles. Synthesized in the liver. Increased concentrations are found in association with inflammation. Fibrinolysis is dependent to the enzyme Plasmin, normally not present in blood in an active form. Kringles, along with TPA and UPA, to bind fibrin lysine molecules during polymerization, Fibrin-bound plasminogen becomes converted into a two-chain active plasmin molecule. Plasmin is a serine protease that systematically digest COMMENTS fibrin polymer. Cleaves Fibrin and Bound plasmin Fibrinogen digests clots and degradation products restores blood vessel X, Y, D-E. patency. Factors XII----XIIa is amplified indirectly by plasmin Activation of Plasminogen can occur to: Destroy factors V and Intrinsic Plasminogen VIII Activation FDP interfere with thrombin influence on fibrinogen
Extrinsic Plasminogen Activation
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The principle inhibitors of fibrinolysis by binding with the plasmin that is free in the plasma (neutralizing plasmin)
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Inhibits the clot-promoting activities of plasma kallikrein
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Inhibits the serine proteases XIIa, XIa, IIa, and Xa
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Hereditary deficiencies have been associated with: Excessive clotting and excessive fibrinolysis
Exogenous Plasminogen Activation Plasminogen Activation in Secretory Ducts. SUMMARY -
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Fibrinolysis is dependent on the enzyme PLASMIN Tissue plasminogen activators, which convert plasminogen to plasmin are released from Injured vessel walls. Plasminogen which is normally present in plasma is converted to plasmin by the action of specific enzyme called “Plasminogen activators.”
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Large naturally occuring plasma GP
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Inhibits component in both the fibrinolysiS and coagulation systems
Activation of Plasminogen can occurs due to: Intrinsic, Extrinsic, Exogenous Plasminogen Activation, and Activation in secretory ducts.
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Inhibits plasmins after alpha 2 antiplasmin depletion
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Plasminogen is a part of any clot because of the tendency of Fibrin to absorb plasminogen from the plasma in normal circumstances.
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When plasminogen activators performed their function, Plasmin is formed within the clot, which gradually dissolves the clot while leaving time for tissue repair
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2. Alpha 2 macroglobulin
Free plasmin is released to the plasma, however, anti-plasmin is there immediately destroy any plasma released from the clot.
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When pathologic coagulation processes are involved, excessive free plasmin is released to the plasma.
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In these situation, the available antiplasmin is depleted and plasmin begins destroying components other than fibrin, including fibrinogen, factor V and VIII and other factors.
FIBRINOLYTIC INHIBITORS 1. alpha-2-anti-plasmin (a2 antiplasmin) -
an (a2) glyco-protein
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Most important naturally occurring inhibitor
3. Alpha 1 antitrypsin -
the third most important naturally occuring inhibitors of fibrinolytic system. Inactivates plasmin slowly and does not bind plasmin until both alpha 2 antiplasmin and alpha 2 macroglobilin are saturated
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inhibits coagulation by its potent inhibitory effects on factor XIa.
Other Inhibitors -
anti-thrombin III, Inhibits fibrinolysis by inhibiting plasmin and kallikrein
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The C1 inactivator also inhibits plasmin
Factor) and GpVI receptor (connects with the subendothelial collagen). Secondary hemostasis •
Progression of the coagulation cascade resulting in the formation of an insoluble fibrin mesh to strengthen the platelet plug.
Primary hemostasis
NORMAL HEMOSTASIS AND COAGULATION
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The formation of the primary platelet plug.
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Platelet adhesion: Platelets adhere to the exposed subendothelial matrix (directly or indirectly via vWf).
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Platelet activation: Once platelets adhere, they then become activated and recruit (and activate) additional platelets to the injured site.
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Platelet plug formation: Fibrinogen forms bridges between activated platelets to form the platelet plug
Endothelial cells • •
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Line the vessels. Are thromboresistant in nature. They express thrombomodulin and heparin sulfate to keep inappropriate thrombi from forming.
Primary Hemostasis Mechanism •
They also release tissue plasminogen activator and urokinase in the presence of thrombin shut off the coagulation cascade in the presence of IIa (thombin).
Platelet adhesion –
GPVI and α2β1 (an integrin): These bind directly to collagen.
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GP1b-IX-V: This binds to vWf, which acts as a bridge or glue between extracellular matrix proteins and the receptor complex of on the platelet surface. vWf is produced by endothelial cells and is secreted into the subendothelial matrix and plasma.
Subendothelium •
Beneath the endothelium. Are thrombogenic in nature.
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Express von Willebrand Factor (vWF)
Hemostasis •
Normal response to stop blood using platelets and the coagulation cascade.
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Platelet activation –
Shape change: Platelets change from their normal discoid shape to elongated cells with cytoplasmic extensions, which increases their surface area.
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The release reaction (degranulation): Platelets release the contents of preformed cytoplasmic granules (α-granules and dense or δ-bodies).
Thrombosis •
Pathologic formation of clots (e.g. deep vein thrombosis)
Primary hemostasis •
Formation of a platelet plug through the platelet’s GpIb-V-IX receptor (connects with the subendothelial von Willebrand
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α-granules contain vWf and coagulation factors, including fibrinogen, Factor V (FV) and FXIII.
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Dense granules are rich in ADP and serotonin
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Enzymatic coagulation factors: Factors XI, X, IX, VII, and II. These are usually in an inactive form and must be activated before they can exert their enzymatic (cleavage) activity.
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Non-enzymatic coagulation factors – cofactors: Tissue factor (TF), Factors V and VIII. Both Factor V and Factor VIII, but not tissue factor, require activation (by an enzymatic coagulation factor, usually thrombin or Factor IIa).
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Non-enzymatic coagulation factor – substrate: Fibrinogen
Platelet aggregation –
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This is mediated primarily by fibrinogen, which binds to the activated fibrinogen receptor (GPIIb/IIIa) on platelets. This links platelets together forming the primary plug. The platelet plug is sufficient to stop bleeding from most small blood vessels in response to every day trauma or venipuncture
Secondary Hemostasis Mechanism •
Initiation of thrombin generation: –
This occurs through tissue factor (extrinsic pathway), which is expressed on fibroblasts.
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This generates small amounts of thrombin and Factor IXa.
Primary Hemostasis Inhibitors •
Physiologic: –
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They also form a physical barrier, preventing platelets from being exposed to thrombogenic subendothelial matrix proteins.
High concentrations of fibrin(ogen) degradation products can inhibit platelet aggregation as can paraproteins
Pharmacologic: –
Arachidonic acid metabolism: Aspirin (irreversibly) and non-steroidal antiinflammatory drugs (reversibly) inhibit cyclo-oxgenase, which is needed for thromboxane A2 production.
Secondary Hemostasis •
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Amplification of thrombin generation: –
This occurs through thrombin, which activates Factor XI of the intrinsic pathway (with platelet polyphosphates as a cofactor) and the cofactors Factor VIII (intrinsic) and Factor V (common).
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This generates a large amount of Factor Xa on the platelet surface.
Pathologic: –
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Endothelial cells release prostacyclin and nitric oxide, both of which inhibit platelet activation.
Secondary hemostasis is usually initiated simultaneously with primary hemostasis upon endothelial damage, although this depends on the degree and location of injury.
Secondary Hemostasis Mechanism •
Propagation of thrombin generation: –
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This occurs through Factor Xa on the platelet surface, with the help of the cofactor, Factor Va, and generates an explosive thrombin burst.
Fibrin formation: –
The explosive thrombin burst cleaves fibrinogen to fibrin, which then forms a soluble fibrin polymer and network of polymers.
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This network is stabilized by Factor XIII, which crosslinks the fibrin.
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C is enhanced by free protein S, which is also vitamin K-dependent and produced in the liver.
Thrombin is responsible for activating Factor XIII and simultaneously inhibiting fibrinolysis, by activating thrombinactivatable fibrinolytic inhibitor. Polyphosphates released from dense granules of activated platelets also inhibit fibrinolysis by helping form a dense fibrin network.
Protein Z: This is a
• factor X inhibitor. •
Pathologic: –
Physiologic
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– •
Extrinsic pathway:
Tissue factor pathway inhibitor (TFPI) is the main inhibitor of this pathway. TFPI inhibits the TF-FVIIa complex and the TF-FVIIa-FXa complex.
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TFPI rapidly down regulates the initiation phase of coagulation, hence thrombin has to amplify its own production. Activity of TFPI is promoted by protein S, which is also a cofactor for an inhibitor of thrombin, protein C.
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Antithrombin also weakly inhibits FVIIa, with stronger inhibition occurring in the presence of heparin-like glycosaminoglycans. –
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Antithrombin: It is an important endogenous anticoagulant, inhibiting the activity of most activated coagulation factors (FIIa, IXa, Xa, XIa and XIIa), although its greatest inhibitory effect is against thrombin (FIIa). Protein C: This is a thrombin inhibitor and vitamin Kdependent enzyme that is produced in the liver. Thus, it is antithrombotic and profibrinolytic. Protein C is activated when thrombin binds to thrombomodulin on endothelial and works by inhibiting the intrinsic tenase and prothrombinase cofactors, FVIIIa and FVa, respectively. Activity of protein
Pharmacologic: –
This includes heparin (which potentiates the action of antithrombin)
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and warfarin, which inhibits recycling of vitamin K, resulting in a relative deficiency of vitamin Kdependent factors (FII, VII, IX, X) – these are not truly deficient (they are being produced); they just cannot bind to PS and become activated.
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Heparin:
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This is frequently administered as an anticoagulant in patients with hypercoagulability (e.g. immunemediated hemolytic anemia).
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unfractionated heparin inhibit both FXa and thrombin.
Intrinsic pathway:
The main inhibitors are antithrombin (AT) and protein C.
High concentrations of fibrin degradation products can inhibit fibrin polymerization as can paraproteins (high concentrations of monoclonal immunoglobulins with multiple myeloma).
Coagulation Pathways Extrinsic pathway This is composed of: •
Tissue factor
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FVII (pro-enzyme; FVIIa is the enzyme)
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Calcium (Ca2+)
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The TF-FVIIa complex the common pathway –
activates FX of
it is thus called the “extrinsic tenase” (extrinsic pathway Factor X activator).
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The TF-FVIIa complex can also activate FIX of the intrinsic pathway (in the so-called “alternate pathway”)
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The FXa is generated by the intrinsic tenase on the platelet surface.
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Thus both intrinsic and extrinsic pathways converge at the activation of FX, however the site of activation of FX differs (fibroblast for the extrinsic pathway and platelet for the intrinsic pathway).
Extrinsic Pathway Mechanism: •
It begins with trauma to blood vessel or tissues outside the blood vessel.
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It releases F-VII and tissue phosholipids.
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F-VII comes in contact with F-III (TF or Thromboplastin) expressed on TF- bearing cells (stromal fibroblasts & leukocytes) forming an activated complex (TF-VIIa)
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Coagulation PathwaysCommon pathway This is composed of: •
TF-VIIa activates F-IX,F-X in presence of Ca++ and tissue phosholipids •
Activation of F-X by TF-VIIa is almost immediately inhibited by TFPI(Tissue factor pathway inhibitor)
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F-Xa acts on F-V and activates it
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F-Xa complexes with tissue phospholipids,F-Va,Ca++ and forms a complex called prothrombinase complex or prothrombin activator
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FXII
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FXI
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and FIX
Ca2+
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PS (Phosphatidylserine )
Intrinsic pathway •
The ultimate product of the intrinsic pathway is activated FIX, which (with the aid of activated cofactor FVIIIa), activates FX.
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In fact, the FIX-FVIIIa-Ca2+-PS complex is called the “intrinsic tenase” (intrinsic pathway Factor X activator).
prothrombin (FII)
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and FXIII (crosslinker)
The cofactor FV
Protein substrate: Fibrinogen
Ca2+
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PS (Phosphatidylserine )
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FXa, with the aid of its cofactor FVa, activates prothrombin to thrombin.
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Thrombin activates FXIII and cleaves fibrinogen to soluble fibrin, which is then crosslinked to insoluble fibrin by FXIIIa.
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X is activated, either by VIIa or tenase , to form Xa – aka prothrombinase
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Xa, with the help of calcium ions, and Va will turn prothrombin into thrombin(FIIa) –
FVIII
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The cofactor (non-enzyme) –
FX
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Enzymatic coagulation factors –
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Intrinsic pathway This is composed of:
Enzymatic coagulation factors
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Factor V is not activated until it has come into contact with thrombin itself. Thus V is not required for this step, but when present will increase the rate.
Thrombin will then activate fibrinogen to fibrin. Fibrin strands will begin to join together, and with the help of XIIIa this will cause the cross-linking of fibrin strands. –
XIII is also activated by thrombin. XIII is also known as fibrin stabilising factor.
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Fibrinolysis involves the dissolution of the fibrin clot by the protease, plasmin
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Constituents: •
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tissue plasminogen activator (tPA)
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contact pathway factors (FXII, prekallikrein, high molecular weight kininogen [HMWK])
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and plasminogen.
Cells: •
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PT (prothombin time) •
Measures the function of the intrinsic pathway and the common pathway.
Heparin •
Increases the activity of ATIII.
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Will increase the aPTT in vitro.
Synthesis of TXA2 is initiated by activated platelets.
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TXA2 increases platelet activation and aggregation.
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plasminogen (proenzyme, plasmin is the enzyme)
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contact pathway factors (FXII, prekallikrein and high molecular weight kininogen)
Cofactor: Fibrin polyphosphates
Facilitators: Bradykinin (releases tPA)
Fibrinolysis Mechanism: This involves the following: •
Release of plasminogen activators
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Plasmin production from plasminogen
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Clot lysis releasing degradation products
Release of plasminogen activators •
Injured endothelial cells release tPA, which is the main plasminogen activator.
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Factor XII is also activated on exposure to subendothelial matrix proteins and forms a complex with high molecular weight kininogen (HMWK) and prekallikrein.
Its synthesis is inhibited by aspirin
Fibrinolysis
tPA
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Thromboxane A2 (TXA2) •
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Activated endothelial cells (source of tPA)
Enzymes:
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Measures the function of the extrinsic pathway and the common pathway.
aPTT (partial thomboplastin time) •
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fibrin clot
Plasmin production •
Plasminogen binds to fibrin through lysine residues.
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Plasminogen activators (tPA, FXIIIa and kallikrein) cleave this bound plasminogen to plasmin, which also remains bound to the clot.
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The binding of plasminogen to the clot amplifies its conversion to plasmin (fibrin acts like a cofactor for tPA).
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Plasmin is a potent lytic enzyme, which can cleave proteins other than fibrin.
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Binding of both plasminogen and its active enzymatic form, plasmin, to fibrin serves to localize fibrinolysis to the clot and prevents plasmin from nonspecifically lysing other proteins.
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Pathologic: This includes polyphosphates ( these can be released by cancer cells and are also found in bacteria) The DNA can also help PAI-1 inhibit the action of tPA on generating plasmin from plasminogen. Histones and DNA also make thicker fibrin strands, which are more resistant to fibrinolysis
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Pharmacologic: Tranexamic acid and εaminocaproic acid. These both act like lysine and are competitive inhibitors for plasminogen (which binds to lysine residues in the fibrin clot).
Clinical Significances of Hemostasis The Bleeding Disorders •
Clot lysis •
Plasmin degrades crosslinked fibrin, releasing variably sized degraded fragments of these proteins, called crosslinked degradation products or Xoligomers.
Hemophilia A •
Hemophilia A is classic hemophilia (a disease referring to the inability to clot blood).
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It is an X-linked disorder resulting from a deficiency in factor VIII, a key component of the coagulation cascade.
Fibrinolytic Inhibitors •
Physiologic: –
Thrombin-activatable fibrinolytic inhibitor (TAFI, also known as carboxypeptidase B): This cleaves lysine residues in fibrin preventing plasminogen binding. It is activated by the thrombin burst.
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Plasminogen activator inhibitor-1 (PAI-1): This is the main inhibitor of tPA and is produced in the liver.
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Antiplasmin: This is the main inhibitor of plasmin and forms complexes with plasmin, targeting it for degradation.
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Polyphosphates: These are released from dense granules in platelets during the release reaction and make a more rigid dense fibrin clot, that resists fibrinolysis.
Defects in the process of hemostasis, leading to bleeding disorders, have been identified at the level of the proteins of the clotting cascades, platelet activation and function, contact activation and antithrombin function.
Hemophilia B •
Hemophilia B results from deficiencies in factor IX.
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The prevalence of hemophilia B is approximately one-tenth that of hemophilia A.
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All patients with hemophilia B have prolonged coagulation time and decreased factor IX clotting activity.
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Like hemophilia A, there are severe, moderate and mild forms of hemophilia B and reflect the factor IX activity in plasma.
Disorders of Fibrinogen and Factor XIII
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Several cardivascular risk factors are associated with abnormalities in fibrinogen.
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As a result of the acute-phase response or through other poorly understood mechanisms, elevated plasma fibrinogen levels have been observed in patients with coronary artery disease, diabetes, hypertension, peripheral artery disease, hyperlipoproteinemia and hypertriglyceridemia.
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In addition, pregnancy, menopause, hypercholesterolemia, use of oral contraceptives and smoking lead to increased plasma fibrinogen levels.
laboratory tests of binding to platelets. –
Factor XI and Contact Activation •
When blood makes contact with negatively charged surfaces it triggers a series of interactions that involve factor XI, prekallikrein and high molecular weight kininogen leading to blood coagulation.
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This process is referred to as contact activation. Deficiency in factor XI confers an injury-related bleeding tendency.
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This deficiency was identified in 1953 and originally termed hemophilia C.
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Factor XI deficiency is very common in Ashkenazic Jews and is inherited as an autosomal disorder with either homozygosity or compound heterozygosity.
Although rare, there are inherited disorders in fibrinogen. –
afibrinogenemia (a complete lack of fibrinogen)
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hypofibrinogenemia (reduced levels of fibrinogen)
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dysfibrinogenemia (presence of dysfunctional fibrinogen
von Willebrand Disease •
Due to inherited deficiency in von Willebrand factor (vWF).
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vWD is the most common inherited bleeding disorder of humans.
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Deficiency of vWF results in defective platelet adhesion and causes a secondary deficiency in factor VIII.
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The result is that vWF deficiency can cause bleeding that appears similar to that caused by platelet dysfunction or hemophilia. –
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Type I vWD is the most common and is inherited as an autosomal dominant trait. This variant is due to simple quantitative deficiency of all vWF multimers Type 2 vWD is also subdivided further dependent upon whether the dysfunctional protein has decreased or paradoxically increased function in certain
Type 3 vWD is clinically severe and is characterized by recessive inheritance and virtual absence of vWF.
Antithrombin Deficiency •
Antithrombin functions to inhibit several activated coagulation factors including thrombin, factor IXa and factor Xa, by forming a stable complex with the various factors.
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Heparin and heparan sulfates increase the activity of antithrombin at least 1000 fold.
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Seen in approximately 2% of patients with venous thromboembolic disease.
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Deficiencies results from mutations that affect synthesis or stability of antithrombin or from mutations that affect the protease and/or heparin binding sites of antithrombin.
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Clinical manifestations of antithrombin deficiency include deep vein thrombosis and pulmonary embolism.
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Arterial thrombosis is rare in anththrombin deficiency.
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Thrombosis may occur spontaneously or in association with surgery, trauma or pregnancy. Treatment of acute episodes of thrombosis is by infusion of heparin (for 5-7 days) followed by oral anticoagulant therapy.
PLATELET DISORDERS 1. Neonatal Alloimmune Thrombocytopenia 2. Immune Thrombocytopenic Purpura 1. Acute ITP 2. Chronic ITP 3. Thrombotic Thrombocytopenic Purpura NEONATAL ALLOIMMUNE THROMBOCYTOPENIA •
It develops when the mother lacks platelet specific antigen that the fetus has inherited from the father.
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HPA – 1A is the most often involved 80% of cases and HPA – 5b accounts for another 10% to 15% of cases.
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Fetal platelet antigens may pass from the fetal to the maternal circulation as early as fourteenth week of gestation.
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If the mother is exposed to a fetal antigen she lacks, she make antibodies to that fetal antigen .
patients between the ages of 20 and 50 years. •
Females with this disorder outnumber males (2:1 to 3:1), with the highest incidence in women between 20 to 40 years of age..
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Platelet destruction in chronic ITP is the result of an immunologic process.
Antibodies cross the placenta Attach to the antigen – bearing fetal platelets Thrombocytopenia in fetus NAIT = HDN •
Clinical significant thrombocytopenia develops in an estimated 1 in 1000 to 2000 newborns.
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With the first pregnancy, about 50% of neonates born to mothers lacking a specific platelet antigen are affected, whereas with subsequent pregnancies the risk is 75% to 97%
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The diagnosis of NAIT is one of the exclusion of other causes of neonatal thrombocytopenia, including maternal ITP and maternal ingestion of drugs known to be associated with drug induced thrombocytopenia.
IMMUNE THROMBOCYTOPENIC PURPURA •
The term was used previously to describe cases of thrombocytopenia arising without apparent cause or underlying disease state.
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IDIOPATHIC = IMMUNE
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Acute and Chronic ITP are immunologically mediated.
THROMBOTIC THROMBOCYTOPENIC PURPURA •
Referred to as Moschowitz syndrome.
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Characterized by the triad of microangiopathic hemolytic anemia, thrombocytopenia and neurologic abnormalities.
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Becomes pentad due to fever and renal dysfunction.
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SYMPTOMS
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Diarrhea
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Anorexia
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Nausea
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Weakness
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Fatigue
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TTP is uncommon but not rare, and its incidence may be increasing.
ACUTE ITP •
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This is primarily a disorder of children, although a similar condition is seen occasionally in adults. The disorder is characterized by the abrupt onset of bruising, petechiae, and sometimes mucosal bleeding (e.g. epistaxis) ina previously healthy child. The primary hematologic feature is thrombocytopenia, which frequently occurs 1 to 3 weeks after an infection.
CHRONIC ITP •
This disorder can be found in ppatients of any age, although most cases occur in
Heparin Induced Thrombocytopenia Also called “Heparin Induced Thrombocytopenia with thrombosis.” Is an adverse effect of treatment with unfractionated heparin 1% to 5% of patients developing this complication In patient who develop HIT, heparin therapy should be stopped as soon as the diagnosis is made, because continued heparin therapy can lead to significant
morbidity and mortality, including gangrene of the extremities, amputation and death The immune system is involved in the development of HIT The clinical signs of HIT are not seen until 7 to 14 days after the initiation of heparin therapy If the patient has been exposed to heparin previously symptoms of HIT may be seen in 1 to 3 days It is recommended that platelet counts be measured daily in patients receiving unfractionated heparin therapy. Immune HIT/HIT-T Also known as: HIT type II, white clot syndrome Heparin-dependent antibodies are usually detectable Non-Immune HAT Also known as HIT type I, HAT Denotes absence of heparindependent antibodies and the potential role for other factors in causing thrombocytopenia Heparin Resistance is a sign of the impending HIT in some patient in which a patient experienced adequate anticoagulation at a certain heparin dosage suddenly requires increasing amount of heparin to maintain same level of anticoagulation. Release of PF4 and B-thromboglobulin from alpha granules both of these substance neutralize heparin, which leads to a normalization of results on the partial thromboplastin time test that is used to monitor heparin therapy The binding of heparin & related compounds depends on polysaccharide chain length, composition and degree of sulfation
Short chain heparin polysaccharides (low MW) have lower affinity to PF4 and are less prone to cause type II HIT The detection of clinically significant HIT occurs by laboratory testing using Immunoassay & Platelet function test All test lack sensitivity Commonly used, but all depend on the presence of free heparin induced antiplatelet antibodies in the patient’s serum or plasma in sufficient quantity to cause a positive test result HIT can be detected by Platelet Aggregation Technique Platelet Aggregation Technique In this method, serum from the patient is added to platelet rich plasma from normal donors, heparin is added to the mixture, and typically monitored for 20 minutes Specificity of the method is excellent but the sensitivity is quite low Sensitivity of the test can be improved, but this requires the use of several heparin concentrations & plate rich plasma from two or more blood donors, preferably of the same ABO blood type as the patient Individuals donating blood for PRP must not have taken aspirin for 10 to 14 days before platelet donation because platelets from donor who have ingested aspirin will produce a false negative result for HIT Serotonin Release Assay (SRA) (Functional Test) In which washed normal platelets from a donor are incubated with radioactive serotonin. Radioactive Serotonin is taken up rapidly and stored in the dense granules of the donor platelets which are washed and resuspended. In the presence of heparin dependent antiplatelet antibody and heparin, the
donor donor platelets become activated and release the contents of their dense granules when the concentration of heparin in the test suspension is near the therapeutic range. The reappearance of radioactive serotonin in the plasma indicates the presence of a heparin dependent antiplatelet antibody The gold standard for the detection of HIT Enzyme Linked Immunosorbent Assay (Antigen Test)
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Hemophilias occur in 1 in 10,000 individuals.
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Also known as classical haemophilia or haemophilia A.
Genetics •
Found in the X chromosome.
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X- linked recessive.
Clinical Manifestations •
Causes anatomic bleeds with deep muscle and joint hemorrhages, hematomas, wound oozing after trauma or surgery and bleeding into the CNS, peritoneum, GI tract, and kidneys.
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Chronic joint bleeds causes inflammation and permanent loss of mobility.
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Bleeding into muscles may cause nerve compression injury.
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Cranial bleeds may lead to severe, debilitating and durable neurologic symptoms such as loss of memory, paralysis, seizures, and coma.
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The diagnosis of haemophilia A begins with the laboratory testing after the birth of an infant to a mother who has a family history of haemophilia.
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In the absence of a family history, abnormal bleeding in the neonatal period, which may appear as early bruising, bleeding from the umbilical stump, postcircumcision bleeding, hematuria, or intracranial bleeding is considered suspicious for haemophilia
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Severe haemophilia is diagnosed in the first year oof life
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Mild haemophilia may not become apparent until a triggering event occurs in late childhood, adolescence, or adulthood
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Severity of haemophilia A is inversely proportional to factor VIII activity.
“ ELISA “ Have been developed based on the knowledge that the antigenic target of the heparin dependent antiplatelet antibody is a PF4-heparin complex In this assay PF4 and Heparin are coated to the surfaces of microplate wells. The serum or plasma from the patient suspected to have HIT is added to wells of microtiter plate If the antibody is present, it adheres to the PF4 heparin complex The plate are washed and enzyme labeled monoclonal antibodies are against human IgG and IgM are added after incubation period the plate is washed and a chromogenic substance for enzyme is added. Color development in the assay indicates the presence of Heparin dependent antiplatelet antibody in the px specimen.
Disorders of the Secondary Hemostasis Hemophilia A (Factor VII deficiency) •
Congenital single factor deficiencies marked by anatomic soft tissue bleeding.
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Second to VWD in prevalence
Complications •
Musculoskeletal lesions and deformities
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Neurologic deficiencies after intracranial hemorrhages
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Other effects:limited productivity, low self-esteem, poverty, drug dependency and depression.
Laboratory Diagnosis •
Records a history of patient’s hemorrhagic symptoms.
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The frequency and severity of bleeding episodes do not correlate with Factor XI deficiency and laboratory monitoring of treatment serves little purpose after the diagnosis is established.
Laboratory Diagnosis •
Increased- PTT
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N- PT
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N- PT and thrombin time
Other congenital Single Factor Deficiencies
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Increased- PTT
Factor V Deficiency -
Treatment •
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The goal of demand haemophilia A treatment is o raise patient’s factor VIII activity to hemostatic levels whenever he or she experiences or suspects a bleeding episode or anticipates a hemostatic challenge such as a surgical procedure. Recombinant factor VIII concentrateprophylactic approach
HEMOPHILIA B (factor IX deFiciency) •
Also called Christmas Disease
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Approximately 14% of haemophilia cases in the United States.
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Reduces thrombin production and causes soft tissue bleeding that is indistinguishable from Hemophilia A.
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Sex-Linked Trait
Laboratory Diagnosis •
Increased- PTT
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Normal- PT
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Factor IX deficiency- if the clinical symptoms suggests haemophilia B.
HEMOPHILIA C (Rosenthal SyndrOME, Factor XI Deficiency) •
Autosomal dominant haemophilia with mild to moderate bleeding symptoms.
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More than half of the cases have been described in Ashkenazi Jews.
The platelet function may be diminished
Factor VII Deficiency -
causes moderate to severe anatomic bleeding.
Factor X Deficiency -
Has been described in amyloidosis, in paraproteinemia and in association with antifungal drug therapy.
Disseminated Intravascular Coagulation (DIC) •
refers to a deposition of large amounts of fibrin throughout the microcirculation that results in a pathologic activation of coagulation pathways.
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is classified as a consumption coagulopathy, because it results in a depletion of platelets as well as plasma coagulation factors.
Can Occur in …
Disorders of Fibrinolysis
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Extrinsic system activation occurs by large amounts of tissue factor entering the circulation.
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Intrinsic system activation results from events that damage or alter thevascular endothelium, thereby exposing collagen
Disorders of Fibrinogen: Hereditary Afibrinogenemia • •
Quantitative deficiency of fibrinogen caused by a lack of synthesis by the liver. Severe hemorrhages predominate in the umbilical, mucosal, gastrointestinal (GI), and intracranial regions.
Dysfibrinogenemia •
Congenital disorders with clinically significant functional abnormalities of fibrinogen
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Qualitative disorder of fibrinogen
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Treatment – Cryoprecipate
Factor XIII Deficiency •
can be clinically severe with moderate-tosevere bleeding.
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Delayed bleeding and wound healing is often observed after trauma.
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Low factor XIII concentrations are detected by incubation of a fibrin clot
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in a 5 M urea solution. A normal fibrin clot will not be dissolved in 5 M urea after 24 hours, but a clot deficient in factor XIII will be dissolved after 24 hours.
Disorders of Fibrinogen: Acquired
Clinical Conditions Associated •
Systemic Hyperfibrinolysis
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HELLP Syndrome
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Hemolytic Uremic Syndrome (HUS)
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Pancreatitis
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Sepsis
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Severe toxic or immunological reactions.
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snake bites
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recreational drugs
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transfusion reactions
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transplant rejection
Disseminated Intravascular Coagulation Treatment: •
Treat underlying disorder
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Heparin therapy (controversial)
Primary fibrinolysis •
symptomatically similar to DIC, but results from increased levels of plasmin.
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Common causes include cirrhosis, shock, metastatic carcinoma of the prostate,
injury to the genitourinary tract, and leaking of urokinase from the urine into tissues.
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can cause a life-threatening clot known as a pulmonary embolism (PE)
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infants develop a life-threatening blood clotting disorder called purpura fulminans soon after birth
Liver disease • •
Majority of the coagulation factors are synthesized in the liver.
Protein C deficiency
Severe liver disease often causes decreased fibrinogen production (hypofibrinogenemia) or an abnormal fibrinogen molecule (dysfibrinogemia).
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are one of the most common causes of hypercoagulability disorders and can be either inherited or acquired.
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Inherited deficiencies are genetically heterozygous in 6% to 12% of cases.
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Acquired deficiencies are caused by chronic oral anticoagulant therapy with coumarin (vitamin K blocker).
Disorders leading to THROMBOSIS -
hypercoagulable state (thrombophilia)
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Patients may present with laboratory abnormalities or with clinical conditions that have been associated with an increased incidence of thrombosis or thromboembolic conditions.
Antithrombin III Deficiency • • •
Principal inhibitor of thrombin in plasma FXa, FXIIa, FXIa, FIXa, TF-FVIIa patient suffers recurrent venous thrombosis and pulmonary embolism
Three natural anticoagulant mechanisms that control thrombus formation •
Antithrombin-III – neutralizes thrombin and the other activateds erine proteases in the coagulation cascade
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Protein C pathway – Protein C :: converted by thrombin to activated protein C (pCa)
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Fibrinolytic System – activates plasmin, which lyses the fibrin clot into fibrin degradation products
Two Types •
Primary Disorders
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Secondary Disorders
Fibrinolytic System Disorders •
Disorders leading to THROMBOSIS: Acquired Homocystinuria
Primary Disorders •
Congenital conditions
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Single component of the hemostatic system
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Patients suffer recurrent thrombosis
Protein S Deficiency •
at risk of a type of clot called a deep vein thrombosis (DVT)
Congenital and acquired disorders involving the fibrinolytic system can impair the breakdown of fibrin, resulting in the build up of fibrin and thus thrombosis
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Disorder of methionine metabolism
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Rare autosomal recessive disorder
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Usually affects infants during the first few years of life
Secondary Disorders -
Lupus Anticoagulant
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Hemostatic Protein Abnormalities
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Platelet abnormalities
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Blood vessel and flow abnormalities
Lupus Anticoagulant (LA) •
Acquired immunoglobulin directed against phospholipids (autoantibodies)
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Associated with an increased risk of thrombosis
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Immune system attacks its own body
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white blood cells mistakenly make anti bodies to mark some of your own body cells and circulation chemicals for destruction.
Hemostatic Protein Abnormalities •
There may be increases in one or more coagulation factors (I, II, V, VII, VIII, IX, and X) or decreases in AT-III, protein C, or fibrinolytic activity.
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down-regulates fibrinolysis in the circulation
Warfarin (coumadin/coumarin) •
used to prevent thrombosis
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inhibits synthesis of Vitamin K dependent factors - impaired fibrin formation
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inadequate level can cause increase risk of thrombosis
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excessive level can cause risk of bleeding
Warfarin Disease Interactions •
Bleeding
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Diabetes
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Hypertension
Platelet abnormalities
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Liver Disease
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Elevated numbers
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Protein C Deficiency
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Increased aggregation response to ADP, collagen, and epinephrine
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Renal Dysfunction
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Increased thrombin generation
Blood Vessel and Flow Abnormalities •
Artificial Surfaces
Other Disorders Related to Fibrinolysis
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Abnormal Vascular Surfaces
Systemic Hyperfibrinolysis
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Abnormal Blood Flow
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Spontaneous bleeding
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From acute states such as heatstroke, hypoxia, hypotension, thoracic surgery, administration of thrombolytics, and neoplasms.
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Also found in Extracorporeal bypass, congenital alpha2-antiplasmin deficiency, and Liver transplant.
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Lab’s: decreased plasminogen and fibrinogen, elevated D-Dimer
Disorders due to Pathologic and Pharmacologic Inhibitors of Coagulation
Plasminogen Activator Inhibitor type 1 Deficiency •
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rare inherited autosomal recessive bleeding disorder characterized by excessive clot lysis leading to a moderate lifelong bleeding diathesis
HELLP Syndrome
may result in increased fibrinolysis and an associated bleeding diathesis
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Pregnant women
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Severe preeclampsia
important component of the coagulation system
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Microangiopathic hemolytic anemia
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Elevated liver enzymes
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Low platelets (Thrombocytopenia)
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Elevated D-Dimer
Thrombotic Thrombocytopenic Purpura (TTP) •
Appears abruptly in previously completely normal healthy individuals.
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Pentad: Thrombocytopenia, Microangiopathic hemolytic anemia, Neurologic abnormalities, Abnormal Renal function, and Fever.
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Auto-immune mediated diseases
Hemolytic Uremic Syndrome (HUS) •
Most common in children
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Associated with E. coli & Campylobacter jejuni.
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Similar to TTP, but we see more kidney damage.
Treatment of TTP / HUS
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Most of cases are women, white race.
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Corticosteroids
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LDH is very high.
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Plasmapheresis / exchange
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The 2 organs relatively spared by the vascular thrombi are the lungs and liver.
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Tx of the underlying disease
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Most of the cases: Unknown
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The goal is to achieve:
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Obstetric emergencies
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Infectious diseases
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Drugs and Toxins
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Neoplastic diseases receiving cytotoxic therapy
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LDH < 500 u/dl
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Normal Platelets
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Normal Hgb / Hct
