Chapter 11B

Extracorporeal Circulation

Thrombosis and Bleeding

L. Henry Edmunds, Jr./ Robert W. Colman

    Contact System
    Intrinsic Coagulation Pathway
    Extrinsic (Tissue Factor) Coagulation Pathway
    Tenase Complexes
    Common Coagulation Pathway
    Thrombin Generation during Extracorporeal Perfusion
    Endothelial Cells

Within the body the endothelial cell, the only surface in contact with circulating blood, simultaneously maintains the fluidity of blood and the integrity of the vascular system. This remarkable cell maintains a dynamic equilibrium by producing anticoagulants to maintain blood in a fluid state and by generating procoagulant substances to enhance gel formation when perturbed. Coagulation proteins circulate as inert zymogens, which convert to active enzymes when stimulated. Likewise, blood cells remain quiescent until activated to express surface receptors and release proteins and enzymes involved in the coagulation equilibrium. The continuous exposure of heparinized blood to the perfusion circuit and to nonendothelial cell tissues of the wound during clinical cardiac surgery produces an intense thrombotic stimulus that involves both the tissue factor pathway (extrinsic coagulation pathway) in the wound and the contact and intrinsic coagulation pathways in the perfusion circuit. Heparin does not block thrombin formation; during extracorporeal perfusion (ECP) heparin partially inhibits thrombin after it is produced. Thrombin is continuously generated and circulated despite massive doses of heparin in all applications of extracorporeal perfusion.365369

When heparinized blood contacts any biomaterial, plasma proteins are instantly adsorbed (<1 second) onto the surface to form a monolayer of selected proteins.370372 For each protein the amount adsorbed depends upon its bulk concentration in plasma and the intrinsic surface activity of the biomaterial. Different biomaterials have different intrinsic surface activities for each plasma protein. The physical and chemical composition of the biomaterial surface determine the intrinsic surface activity of the biomaterial, but intrinsic surface activity is not predictable from knowledge of chemical and physical characteristics. Thus intrinsic surface activity differs among biomaterial surfaces, among plasma proteins, and among different bulk concentrations of plasma proteins. Concentrations of plasma proteins on a given biomaterial differ from concentrations in bulk plasma. Similarly, concentrations of surface-adsorbed proteins from the same plasma differ on different biomaterials. The composition of the protein monolayer is specific for the biomaterial and for various concentrations of proteins in the plasma, but the topography of the adsorbed protein layer may not be uniform across the surface of the biomaterial.373 Thus it is not possible to predict the "thrombogenicity" of any biomaterial except by trial and error.

On most biomaterial surfaces fibrinogen is selectively adsorbed, but the adsorbed concentration of fibrinogen and other proteins may change over time.372 Surface-adsorbed proteins "compete" for space on the biomaterial surface, but are tightly packed, irreversibly bound, and immobile. The density of surface-adsorbed proteins is 100 to 1000 times greater than the density of proteins in bulk plasma.373 The complexity of blood-biomaterial interactions is further compounded by the fact that adsorbed proteins often undergo limited conformational changes374,375 that may expose "receptor" amino acid sequences that are recognized by specific blood cells or bulk plasma proteins. Conformational changes of adsorbed factor XII and fibrinogen initiate activation of the contact pathway and platelet surface adhesion, respectively; similar changes in complement protein 3 participate in activation of the complement system.375 For a given adsorbed protein these conformational changes may vary between biomaterial surfaces and in turn vary the reactivity of the adsorbed protein with cells and blood proteins in the bulk phase.

Thus heparinized blood does not directly contact biomaterial surfaces in extracorporeal perfusion circuits, but contacts monolayers of densely packed, immobile plasma proteins arranged in undefined mosaics that differ between locations and possibly across time. All biomaterial surfaces, including heparin-coated surfaces, are procoagulant367,376; only the endothelial cell is truly nonthrombogenic (Fig. 11-9).

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FIGURE 11-9 Electron micrograph of a rabbit endothelial cell (E), the only known nonthrombogenic surface. Note the overlapping junctions with neighboring endothelial cells. Endothelial cells rest on the internal elastic lamina (I), which abut medial smooth muscle cells. The vessel lumen is at the top. (Reproduced with permission from Stemerman MB: Anatomy of the blood vessel wall, in Colman RW, Hirsh J, Marder VJ, Salzman E (eds): Hemostasis and Thrombosis: Basic Principles and Clinical Practice, 2d ed. Philadelphia, JB Lippincott, 1987; p 775.)


ECP and cardiopulmonary bypass (CPB) are not possible without anticoagulation; the large procoagulant surface quickly overwhelms natural circulating anticoagulants—antithrombin, proteins C and S, tissue factor pathway inhibitor, and plasmin—to produce thrombin and thrombosis within the circuit. Thrombin is produced in extracorporeal perfusion systems with small surface areas and high-velocity flow,368,377,378 but thrombosis may not be apparent if other procoagulants (e.g., addition of blood from wounds) are absent. Generation of thrombin varies widely between applications of extracorporeal technology (see below), but this powerful and potentially dangerous enzyme is produced whenever blood contacts a nonendothelial cell surface (Fig. 11-10).

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FIGURE 11-10 Plasma thrombin-antithrombin (TAT) measurements of thrombin generation during CPB and clinical cardiac surgery of varying duration. (Data from Brister SJ, Ofosu FA, Buchanan MR: Thrombin generation during cardiac surgery: is heparin the ideal anticoagulant? Thromb Haemost 1993; 70:259.)

During CPB and open heart surgery (OHS) high concentrations of heparin (3–4 mg/kg, initial dose) are needed to maintain the fluidity of blood. Heparin has both advantages and disadvantages; the most notable advantages are parenteral use, immediate onset of action, and rapid reversal by protamine or recombinant platelet factor 4.379 Heparin does not directly inhibit coagulation, but acts by accelerating the actions of the natural protease, antithrombin.380 Heparin-catalyzed antithrombin, however, does not inhibit thrombin bound to fibrin381 or factor Xa bound to platelets within clots382; thus heparin only partially inhibits thrombin in vivo. Antithrombin primarily binds thrombin; its action on factors Xa and IXa is much slower. Heparin inhibits coagulation at the end of the cascade after

nearly all other coagulation proteins have been converted to active enzymes. In addition, heparin to varying degrees activates several blood constituents: platelets,383385 factor XII,386 complement, neutrophils, and monocytes.387389 Heparin increases the sensitivity of platelets to soluble agonists,385 inhibits binding to von Willebrand factor,390 and modestly increases template bleeding times.384 Thrombin concentrations cannot be measured in real time and only insensitive, indirect methods are available to regulate heparin anticoagulation in the operating room.391393

Heparin is also associated with some clinical idiosyncrasies. In some patients recent, prolonged parenteral heparin may reduce antithrombin concentrations and produce heparin resistance.380,394,395 Insufficient antithrombin may also occur due to insufficient synthesis or increased consumption in some cyanotic infants, premature babies, cachectic patients, and patients with advanced liver or renal disease. The deficiency in antithrombin prevents heparin from prolonging activated clotting times to therapeutic levels. In these patients fresh frozen plasma is needed to increase plasma antithrombin concentrations to inhibit thrombin. Heparin rebound is a delayed anticoagulant effect after protamine neutralization due to the rapid metabolism of protamine and delayed seepage of heparin into the circulation from lymphatic tissues and other deposits. Heparin is also associated with an allergic response in some patients that produces heparin-induced thrombocytopenia (HIT) with or without thrombosis (see below). Lastly, heparin only partially suppresses thrombin formation during CPB and all applications of extracorporeal perfusion and mechanical circulatory and respiratory assistance despite doses 2 to 3 times those used for other indications (see Fig. 11-10).365368 Thus heparin is far from an ideal anticoagulant.

Potential alternatives for heparin during ECP include low molecular weight heparin, danaparoid (Organan), recombinant hirudin (Lepirudin), and the organic chemical, argatroban (Texas Biotechnology Corp.). All have important drawbacks and are approved for use in heparin-induced thrombocytopenia and in patients with circulating IgG anti-heparin-PF4 complex antibodies (see below). Low molecular weight heparins have long half-lives in plasma (4–8 hours), require antithrombin as a cofactor, primarily inhibit factor Xa, and are not reversible by protamine.396,397 Although less antigenic than standard heparin, low molecular weight heparins can stimulate production of IgG anti-heparin-PF4 complex antibodies.397 Danaparoid is a mixture of heparin sulfate, dermatan sulfate, and chrondroitin sulfate that catalyzes antithrombin to inhibit thrombin and factor Xa. To a lesser extent, Danaparoid also catalyzes inhibition of thrombin by heparin cofactor II. The anticoagulant effect is long lasting (plasma half-life 4.3 hours)398 and is not reversed by protamine.

Recombinant hirudin (Lepirudin) is a direct inhibitor of thrombin, is effective rapidly, does not have an effective antidote, is monitored by the partial thromboplastin time, is cleared by the kidney, and has a relatively short half-life in plasma (40 minutes).399 This drug has been successfully used during CPB and open heart surgery, but in many instances bleeding after bypass has been troublesome and substantial. A newer drug is a semisynthetic bivalent thrombin inhibitor composed of 12 amino acids from hirudin, which binds to exosite 1 of thrombin linked to an active site-directed moiety, D Phe Pro Arg Pro, by four glycines.400 This drug, bivalirudin (Angiomax), has a shorter half-life than hirudin and therefore may be safer. In addition, only a small amount is excreted by the kidney. In coronary angioplasty, bivalirudin was as effective as heparin but there was less bleeding. Argatroban is also a direct thrombin inhibitor401 with rapid onset of action and short plasma half-life (40–50 minutes).402 Argatroban is metabolized in the liver and is without an antidote, but can be monitored with partial thromboplastin times or activated clotting times. At present there is little clinical experience with argatroban or bivalirudin in cardiac surgical patients.

Heparin-associated thrombocytopenia (HAT) is a benign, nonimmune, 5% to 15% decrease in platelet count that occurs within a few hours to 3 days after heparin exposure. The etiology is due to mild platelet stimulation from multifactorial causes; bleeding does not occur; and the condition is clinically inconsequential.403

Heparin-induced thrombocytopenia (HIT) and heparin-induced thrombocytopenia and thrombosis (HITT) are different manifestations of the same immune disease. Heparin binds to platelets in the absence of an antibody and releases small amounts of platelet factor 4 (as occurs in HAT). PF4 avidly binds heparin to form a heparin-PF4 (H-PF4) complex, which is antigenic in some people. In these individuals IgG antibodies to the H-PF4 complex are produced within 5 to 15 days after exposure to heparin and continue to circulate in the absence of more heparin for approximately 3 to 6 months.404 IgG-anti-H-PF4 antibodies plus H-PF4 complexes form HIT complexes, which unite IgG Fc terminals to platelet Fc receptors (Fig. 11-11). This binding strongly stimulates platelets to release more PF4.405 A self-perpetuating, accelerating cascade of platelet activation, release, and aggregation ensues. Since platelet granules contain several procoagulatory proteins (e.g., thrombin, fibronectin, factor V, fibrinogen, von Willebrand factor), release also activates coagulation proteins to generate thrombin.

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FIGURE 11-11 The generation of HIT complexes. Read each horizontal group of three left to right beginning at top left. See text for full explanation.

The intensity of the immune reaction varies between patients, but also varies by the indications for heparin use. Both heparin and PF4 must be available to form the antigenic H-PF4 complex. Patients who do not have conditions that activate platelets have a low incidence of HIT following administration of heparin, because few PF4 molecules are available to form H-PF4 complexes. In medical patients the incidence of thrombocytopenia after heparin is about 0.5%; the incidence of HITT is approximately 0.25%; and only 3% have IgG anti-H-PF4 antibodies by enzyme immunoassay.406 Large doses of heparin are given and huge numbers of platelets are activated during CPB. Thus after CPB, 50% of patients have IgG anti-H-PF4 antibodies; 2% have immune heparin-induced thrombocytopenia; and approximately 1% develop HITT.406 A combination of three ingredients is necessary to produce HIT or HITT: heparin, platelet factor 4, and IgG anti-H-PF4 antibodies. Since IgG antibodies are transient, a second heparin exposure 6 months after HIT is not likely to produce HIT or HITT,404 but will stimulate production of new IgG antibodies to the H-PF4 complex. The danger is a second heparin exposure when IgG anti-H-PF4 antibodies are still circulating.

IgG anti-H-PF4 antibodies are detected in two ways. The serotonin release test detects the release of radioactive serotonin from normal platelets washed by the patient's serum.407 An enzyme immunoassay measures IgG anti-H-PF4 antibodies directly. Both assays are equally sensitive in patients with clinical HIT, but the enzyme immunoassay is more sensitive in detecting IgG anti-H-PF4 antibodies in patients without other evidence of the disease.407

The clinical presentation of HIT may be insidious. If the platelet count was originally normal, the earliest sign is an abrupt decrease of at least 50% in platelet count (to less than 150,000/µL) in a patient who has had exposure to heparin within the past 5 to 15 days.404 This event is a preoperative stop sign for elective cardiac operations. After CPB, platelet counts below 80,000/µL should trigger an order to stop all heparin, including heparin flushes, and to obtain daily platelet counts. The patient should be thoroughly examined for deep vein thrombosis, extremity ischemia, stroke, myocardial infarction, or any evidence of intravascular thrombosis using ultrasound and appropriate radiographic technology. Any evidence of vascular thrombosis should prompt a plasma sample for IgG anti-H-PF4 antibodies. A positive antibody test confirms the diagnosis of HIT in patients with thrombocytopenia and HITT in those with either venous or arterial thrombosis or both. It is important to stress that HIT or HITT is a clinical diagnosis and that a positive antibody test is not required before stopping heparin.

Once the diagnosis of HIT or HITT is suspected, management must focus on prevention of further intravascular thrombosis. Bleeding is rarely the problem; intravascular thrombosis is. Neither heparin nor platelet transfusions should be given; platelet transfusions only add more PF4 if heparin and IgG anti-H-PF4 antibodies are still circulating. If heparin is proven absent from the circulation, platelet transfusions may be used very cautiously if the patient has significant nonsurgical bleeding. Surgical measures to reopen thrombosed large arteries are usually futile because the platelet-rich thrombus (white clot) often extends into small arteries and arterioles. An inferior vena cava filter is recommended if pulmonary embolism is likely or has occurred.

Modern management also includes full anticoagulation with recombinant hirudin (Lepirudin), argatroban, or possibly bivalirudin to prevent further extension of thrombosis or development of clinical intravascular thrombosis. This may occur in 40% to 50% of patients with HIT who are treated only with heparin cessation.408 At present there is little experience with argatroban in cardiac surgical patients with HITT, but the drug is a direct thrombin inhibitor, has attractive pharmokinetics, and is approved for patients with HITT. Full anticoagulation with hirudin in fresh postoperative cardiac surgical patients is recommended, but the safety zone between bleeding and thrombosis is narrow. The patient must be carefully monitored for pericardial tamponade and signs of hidden bleeding. Hirudin is monitored by aPTT and the range used is similar to that with intravenous heparin. The effective blood concentration of hirudin for thrombin inhibition is 0.5 to 1.5 µg/mL.409 To achieve this, 0.2 mg/kg/h infusions are recommended.409 Dose must be reduced in patients with renal failure because the kidney clears the drug. Argatroban is sometimes a better choice, but it should be remembered that it is difficult to manage in the presence of liver disease since it is metabolized in that organ. In most patients oral anticoagulation with warfarin is started at the same time as intravenous hirudin, but warfarin should not be started prior to hirudin.

Emergency or urgent open heart surgery with CPB using hirudin is possible in patients with circulating IgG anti-HPF4 antibodies. The therapeutic level of drug should be between 3.5 and 4.5 µg/mL during CPB.409 Greinacher recommends bolus doses of 0.25 mg/kg IV and 0.2 mg/kg in the priming volume followed by an infusion of 0.5 mg/min until 15 minutes before stopping CPB.409 At that time 5 mg of hirudin is added to the perfusate to prevent clotting within the heart-lung machine.

Patients who require elective cardiac surgery are best deferred until circulating IgG anti-H-PF4 antibodies are absent by enzyme immunoassay. Patients with a history of HIT who require elective cardiac surgery with CPB should have IgG anti-H-PF4 antibodies measured in their serum before surgery is scheduled. If antibodies are absent, elective surgery can be safely carried out using heparin anticoagulation, if the first reexposure to heparin is the bolus dose given just before starting CPB. Since HIT requires the presence of the H-PF4 complex plus IgG anti-H-PF4 antibodies to form the HIT complex, and since it takes about 5 days to produce these antibodies, HIT or HITT will not occur if no further heparin is given after operation.

Generation of thrombin during cardiopulmonary bypass and other applications of extracorporeal circulatory technology is the cause of the thrombotic and bleeding complications associated with ECP. Theoretically, if thrombin formation could be completely inhibited during ECP, the consumptive coagulopathy, which consumes coagulation proteins and platelets and causes bleeding complications, would not occur.

Thrombin generation and the fibrinolytic response primarily involve the extrinsic and intrinsic coagulation pathways, the contact and fibrinolytic plasma protein systems, and platelets, monocytes, and endothelial cells.

Contact System

The contact system includes four primary plasma proteins—factor XII, prekallikrein, high molecular weight kininogen (HMWK), and C-1 inhibitor410—and is activated during CPB and clinical cardiac surgery.411 This system is involved in complement and neutrophil activation and the inflammatory response to ECP, but is not involved in thrombin formation in vivo. However, when blood contacts a negatively charged surface (protein surfaces contain both positive and negative charges) in ECP, small amounts of factor XII are adsorbed and undergo a conformational change to factor XIIa.373,412 Factor XIIa in the presence of HMWK activates factor XI and initiates the intrinsic coagulation pathway (Fig. 11-12). Thrombin also activates factor XI, and is the predominating agonist in vivo in pathologic states.413

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FIGURE 11-12 Steps in the generation of thrombin in the wound and in the perfusion circuit via the extrinsic, intrinsic, and common coagulation pathways. PK, prekallikrein; HMWK, high molecular weight kininogen; Ca++, calcium ion; PL, cellular phospholipid surface; TF, tissue factor; mono, monocyte. Activated coagulation proteins are indicated by the suffix "a."

Intrinsic Coagulation Pathway

The intrinsic coagulation pathway probably does not generate thrombin in vivo, but does initiate thrombin formation when blood contacts nonendothelial cell surfaces such as perfusion circuits.414,415 Factor XIa, produced by activation of the contact system and subsequently thrombin generation, activates factor IX, which forms part of the intrinsic tenase complex.416,417 Factor XI is primarily activated by thrombin (see Fig. 11-12).

Extrinsic (Tissue Factor) Coagulation Pathway

The extrinsic coagulation pathway is the major coagulation pathway in vivo and is a major source of thrombin generation during CPB and clinical cardiac surgery.418,419 Exposure of blood to tissue factor initiates the extrinsic coagulation pathway.417 Tissue factor (TF) is a cell-bound glycoprotein that is constitutively expressed on the cellular surfaces of fat, muscle, bone, epicardium, adventia, injured endothelial cells, and many other cells except pericardium.419421 Plasma TF associated with wound monocytes is a second source of TF and may be an important source during CPB and clinical cardiac surgery (authors' unpublished data; ref. 422). Tissue factor is the cofactor for the activation of factor VII to factor VIIa, which is part of extrinsic tenase (see Fig. 11-12).

Tenase Complexes

Intrinsic and extrinsic tenase catalyze the activation of factor X to factor Xa (see Fig. 11-12). Extrinsic tenase is formed by the combination of tissue factor, factor VIIa, calcium, and a phospholipid surface to cleave a small peptide from factor X to form factor Xa.417 Extrinsic tenase also generates small amounts of factor IXa,423 which greatly accelerates formation of intrinsic tenase and is the major pathway for the formation of factor Xa. Intrinsic tenase is produced by the combination of factor IXa, factor VIIIa, and calcium on the surface of an activated platelet,424 and catalyzes production of factor Xa 50 times faster than extrinsic tenase.417 Factor Xa activates factors V and VII in feedback loops.

Common Coagulation Pathway

Factor Xa is the gateway protein of the common coagulation pathway. Factor Xa slowly cleaves prothrombin to {alpha}-thrombin, the active enzyme, and a fragment, F1.2, but the reaction is 300,000 times faster if catalyzed by the prothrombinase complex.417 The prothrombinase complex is produced when factor Xa, in the presence of Ca2 +, is anchored by factor Va onto a phospholipid surface provided by platelets, monocytes, or endothelial cells.417 Either factor Xa or thrombin activates factor V to factor Va. The prothrombinase complex cleaves prothrombin to {alpha}-thrombin and a fragment, F1.2, and is the major pathway producing thrombin.417 F1.2 is a useful marker of the reaction.


Thrombin is a powerful enzyme that accelerates its own formation by several feedback loops.425 Thrombin is the major activator of factor XI and the exclusive activator of factor VIII in the intrinsic pathway. Thrombin is a secondary activator of factor VII, but once formed may be the most important activator in the wound. Lastly, thrombin is the primary activator of factor V in the formation of the prothrombinase complex (see Fig. 11-12).

Thrombin has both procoagulant and anticoagulant properties.425 Thrombin is the enzyme that cleaves fibrinogen to fibrin and in the process creates two fragments, fibrinopeptides A and B. Thrombin activates platelets via the platelet thrombin receptor and thus may be the major agonist for platelets both in the wound and in the perfusion circuit. Thrombin also activates factor XIII to cross-link fibrin to an insoluble form and to attenuate fibrinolysis. Lastly, thrombin activates TAFI, thrombin-activated fibrinolysis inhibitor, which alters fibrin to reduce lysis.425

Thrombin also stimulates the production of anticoagulants. Surface glycosaminoglycans, such as heparan sulfate, inhibit thrombin and coagulation via antithrombin. Thrombin stimulates endothelial cells to produce tissue plasminogen activator, t-PA, which is the major enzyme that cleaves plasminogen to plasmin. Thrombin also stimulates the production of nitric oxide and prostaglandin by endothelial cells. Thrombin in the presence of thrombomodulin activates protein C, which in the presence of protein S destroys activated factor V and VIII.

Thrombin Generation during Extracorporeal Perfusion

All applications of extracorporeal perfusion and exposure of blood to nonendothelial cell surfaces generate thrombin.365367 F1.2 is a protein fragment that is formed when prothrombin is cleaved to thrombin; thus F1.2 is a measure of thrombin generation but not of thrombin activity. F1.2 and thrombin-antithrombin (TAT) complex increase progressively during clinical cardiac surgery with CPB, during applications of circulatory assist devices368,379 and during extracorporeal life support (ECLS) (see Fig. 11-10). The amount of thrombin produced seems to vary with the intensity of the stimuli for thrombin production and may vary with age, comorbid disease, and clinical health of the patient. The cytokines, IL-1ß and TNF-{alpha}, are procoagulant and inhibit the thrombomodulin/protein C anticoagulant pathway and stimulate production of type I plasminogen activator inhibitor.426 Complex cardiac surgery that requires several hours of CPB produces more F1.2366 than short procedures with minimal exposure of circulating blood to the wound (e.g., first-time myocardial revascularization).427 Thrombin generation varies with the amount and type of anticoagulant used; surface area of the blood-biomaterial interface; duration of exposure to the surface; turbulence, stagnation, and cavitation within perfusion circuits; and to a lesser degree temperature and the "thromboresistant" characteristics of biomaterial surfaces.428 Very high concentrations of heparin, sufficient to increase spontaneous bleeding, reduce F1.2 production probably by interfering with thrombin-activated feedback loops,429 since heparin does not directly inhibit thrombin formation.

For many years blood contact with the biomaterials of the perfusion circuit was thought to be the major stimulus to thrombin formation during CPB and OHS. Increasing evidence indicates that the wound is the major source of thrombin generation during CPB and clinical cardiac surgery.365,366,430 This understanding has encouraged development of strategies to reduce the amounts of circulating thrombin during clinical cardiac surgery by either discarding wound blood431 or by exclusively salvaging red cells by centrifugation in a cell saver. The reduced thrombin formation in the perfusion circuit has also supported misguided strategies for reducing the systemic heparin dose during first-time coronary revascularization procedures using heparin-bonded circuits.427,432 While there is no good evidence that heparin-bonded circuits reduce thrombin generation,367 there is strong evidence that discarding wound plasma or limiting exposure of circulating blood to the wound (e.g., less bleeding in the wound) does reduce the circulating thrombin burden.376,432


Platelets are activated by thrombin, contact with the surface of nonendothelial cells, heparin, and platelet-activating factor produced by a variety of cells during all applications of extracorporeal perfusion and/or recirculation of anticoagulated blood that has been exposed to a wound. Circulating thrombin and platelet contact with surface-adsorbed fibrinogen in the perfusion circuit are probably the earliest and strongest agonists. Circulating thrombin, although rapidly inhibited by antithrombin, is a powerful agonist and binds avidly to two specific thrombin receptors on platelets: PAR-1 and GPIb{alpha}.433 As CPB continues, C5a, C5b-9,434,435 plasmin,436 hypothermia,437 platelet-activating factor (PAF), interleukin-6,438 cathepsin G, serotonin, epinephrine, eicosanoids, and other agonists also activate platelets and contribute to their loss and dysfunction.

The initial platelet reaction to agonists is shape change. Circulating discoid platelets extend pseudopods, centralize granules, express GPIb and GPIIb/IIIa receptors,439 and secrete soluble and bound P selectin receptors from alpha granules.440 GPIIb/IIIa ({alpha}IIbß3) receptors almost instantaneously bind platelets to exposed binding sites on the {alpha}- and {gamma}-chains on surface-adsorbed fibrinogen (Fig. 11-13). 441 The number of adherent platelets is proportional to the amount of surface-adsorbed fibrinogen recognized by fibrinogen antibody,442 but the density of adherent platelets also varies with the chemical and physical composition of the surface biomaterial.443,444 Rough surfaces accumulate more platelets than smooth surfaces.445 Fewer platelets adhere to polyurethane, cuprophane, and PMEA (poly 2-methoxyethylacrylate) than to silicone rubber.374,428 Platelet adhesion and aggregate formation reduce the circulating platelet count, which is already reduced by dilution with pump priming solutions.

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FIGURE 11-13 Adhesion of activated platelets binding to surface-adsorbed fibrinogen via GPIIb/IIIa ({alpha}IIbß3) receptors. The same receptors bind plasma fibrinogen molecules to form platelet aggregates.

Plasma fibrinogen forms bridges between platelets expressing GPIIb/IIIa receptors to produce circulating platelet aggregates. Platelet bound P-selectin binds platelets to monocytes and neutrophils to form aggregates.446 During ECP some adherent platelets detach, leaving membrane fragments behind,447 to produce platelet microparticles and partially fragmented platelets.448,449 Some of these platelet membrane fragments also detach and circulate.448,449

A small percentage of activated platelets synthesize and release a variety of chemicals and proteins from granules that include thromboxane A2,450 platelet factor 4, ß-thromboglobulin,451 P-selectin, and serotonin. Platelet lysosomes release neutral proteases and acid hydrolases.452

During ECP the circulating platelet pool is reduced by dilution, adhesion, aggregation, destruction, and consumption. The platelet mass consists of a reduced number of morphologically normal platelets, platelets with pseudopod formation, new and larger platelets released from megakaryocytes,453 partially and completely degranulated platelets, platelet membrane fragments, platelet microparticles, and resealed platelets that have lost some of their membrane receptors.446,447,452,453 Most of the circulating platelets appear structurally normal,453 but bleeding times increase and remain prolonged for several hours after protamine.454 The functional state of the circulating intact platelet during and early after CPB is reduced, but it is not clear whether this functional defect is intrinsic or extrinsic to the platelet. Flow cytometry studies of circulating intact platelets show little change in platelet membrane receptors.455 In prolonged applications of ECP, platelets are consumed and may or may not be adequately replaced by new platelets from the bone marrow.456


During CPB and clinical cardiac surgery the concentration of plasma tissue factor, which normally is 0.26 to 1.1 pM,422,457 doubles to 2.0 pM.422 During cardiac surgery wound plasma contains 6 to 11 pM.422 In the wound with calcium present, monocytes associate with plasma tissue factor to rapidly accelerate the conversion of factor VII to factor VIIa (authors' unpublished data). This association is specific for monocytes—the reaction is essentially nil for platelets, neutrophils, and lymphocytes—and does not occur if monocytes, plasma tissue factor, or factor VII is not present. Monocytes also synthesize and express tissue factor, but this process, which peaks 3 to 4 hours after monocytes are activated,458 is not a major source of tissue factor during CPB and clinical heart surgery but does occur during prolonged perfusions.459 Plasma microparticles, also present in wound plasma, are procoagulant460 and monocytes may express the procoagulant CD 11b receptor,461 but the clinical importance of these pathways in thrombin generation is not clear and probably minor. The major sources of tissue factor in the wound are the combination of monocytes, plasma tissue factor, and cell-bound tissue factor.

Agonists for activating monocytes during CPB and clinical cardiac surgery include C5a,462 endotoxin, IL-6, IL-1ß, tumor necrosis factor (TNF-{alpha}), and monocyte chemotactic protein-1 (MCP-1). Monocytes and macrophages produce MCP-1, IL-1ß, IL-6, and TNF-{alpha}463,464; express tissue factor,458 Mac-1,461 L-selectin, and monocyte chemotactic protein –1 (MCP-1)465; and form aggregates with platelets.440 For the most part, monocyte reactions are slow and peak concentrations of cytokines occur several hours after CPB ends.466468

Endothelial Cells

Endothelial cells, charged with maintaining the fluidity of circulating blood and the integrity of the vascular system, are activated during CPB and clinical cardiac surgery by thrombin, C5a,469 IL-1, and TNF-{alpha}.470 Endothelial cells produce both procoagulants and anticoagulants. Procoagulant activities of endothelial cells include expression of tissue factor and production of a host of procoagulant proteins, including collagen, elastin, microfibillar protein, laminin, fibronectin, thrombospondin, von Willebrand factor, factor V, platelet-activating factor, and plasminogen activator inhibitor 1, and the vasoconstrictors, endothelin-1 and renin. Endothelial cells also bind von Willebrand factor, vibronectin, and factors IXa and Xa. Anticoagulant activities of endothelial cells include the production of tissue plasminogen activator (t-PA), heparin sulfate, dermatan sulfate, protein S (which accelerates the activation of protein C), tissue factor inhibitor protein, thrombomodulin and protease nexin 1 (which both bind thrombin), prostacyclin,471 nitric oxide, and adenosine. Prostacyclin concentrations increase rapidly at the beginning of CPB and then begin to decrease.472 During clinical cardiac surgery endothelin-1 peaks several hours after CPB ends.473

Except for expression of tissue factor and expression of CD11b/CD18 (Mac-1), which is weakly procoagulant, endothelial cell receptors do not participate heavily in thrombin generation during ECP.


During ECP neutrophils express Mac-1 receptors,474 which bind factor X and fibrinogen and weakly facilitate thrombin formation. Neutrophils secrete elastase, which can destroy protease inhibitors such as antithrombin and coagulation factors such as factor V and may contribute significantly to the equilibrium between the fluid and gel forms of blood.

A vast number of particulate emboli are produced during all applications of ECP and most of these emboli originate from blood constituents (see sections 11A and 11D). These macro- and microemboli contribute to temporary and permanent organ damage and impairment of neurocognitive function. Emboli directly produced by activation of the coagulation system during ECP include fibrin; platelet microparticles, fragments, and aggregates; and platelet-leukocyte aggregates.

Circulating thrombin activates endothelial cells to produce tissue plasminogen activator (t-PA), which binds avidly to fibrin.475477 Endothelial cells are the principal source of t-PA.476 The combination of t-PA, fibrin, and plasminogen cleaves plasminogen to plasmin; plasmin cleaves fibrin.476 This reaction produces the protein fragment, D-dimer, which is a useful marker of fibrinolysis, and a marker of thrombin activity because fibrin is cleaved from fibrinogen by thrombin. Kallikrein, produced by the contact system, cleaves pro-urokinase to urokinase; however, this enzyme is less important in fibrinolysis than t-PA because urokinase binds poorly to fibrin.476 F1.2, D-dimer, and fibrinopeptide A (produced by the conversion of fibrinogen to fibrin) increase during extracorporeal perfusion, indicating ongoing thrombin production, fibrin formation, and fibrinolysis.366,367,478,479 D-dimer and other fibrin degradation products are themselves anticoagulants inhibiting fibrin polymerization.476

Fibrinolysis is controlled by native protease inhibitors, {alpha}2-antiplasmin, {alpha}2-macroglobulin, and plasminogen activator inhibitor-1.477 Plasminogen activator inhibitor-1, produced by endothelial cells, directly inhibits t-PA and urokinase, but little is produced during CPB and open cardiac surgery.480 Alpha 2-antiplasmin rapidly inhibits unbound plasmin, preventing the enzyme from circulating, but poorly inhibits plasmin bound to fibrin. Alpha 2-macroglobulin is a slow inhibitor of plasmin.

Plasmin is both a stimulator and inhibitor of platelets depending upon concentration and temperature.481 High concentrations of plasmin at normothermia and low concentrations during hypothermia cause conformational changes in platelets, centralization of platelet granules, and internalization of platelet GPIb receptors but not GPIIb/IIIa receptors.482

Simultaneous and ongoing thrombin formation and fibrinolysis is by definition a consumptive coagulopathy483 and is present in all applications of ECP. In the normal state the fluidity of blood and the integrity of the vascular system are established and maintained by an equilibrium between procoagulants favoring clot and anticoagulants favoring liquidity (Fig. 11-14A). Blood contact with ECP systems and the wound disrupts this equilibrium to produce a massive procoagulant stimulus that overwhelms natural anticoagulants; therefore, an exogenous anticoagulant, heparin, is required for nearly all applications of ECP (Fig. 11-14B). Exceptions are only possible in applications that produce a relatively weak procoagulant stimulus and a minimal thrombin burden that can be contained by natural anticoagulants. Surgeons must realize that any blood exposure to nonendothelial cell surfaces, including prosthetic heart valves, produces a procoagulant stimulus whether or not clot is produced. Except for the healthy endothelial cell, no nonthrombogenic surface exists.

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FIGURE 11-14 (A) The balance between procoagulant and anticoagulant forces that produces an equilibrium allows blood to circulate. (B) During CPB and OHS (open heart surgery) the normal equilibrium is disturbed by changes in both procoagulants and anticoagulants. Imbalance of procoagulants risks thrombosis; an imbalance of anticoagulants risks bleeding.

This concept of an equilibrium between procoagulants and anticoagulants is helpful in managing the thrombotic and bleeding complications associated with all applications of ECP. During ECP procoagulant stimuli, manifested by thrombin formation that is not measurable in real time, must be balanced by either increased anticoagulation or a reduction in the thrombin burden to maintain equilibrium. After ECP, anticoagulants must be inhibited to avoid excessive bleeding. During consumptive coagulopathy, coagulation proteins and platelets are consumed and may become too deficient to generate thrombin and fibrin-platelet clots. In cardiac surgical patients many additional variables affect the coagulation equilibrium and impact the availability of coagulation proteins and functional platelets. These variables include the quantity of blood in contact with the wound; surface area of the perfusion system; duration of perfusion; circulating anticoagulants; and to lesser degrees temperature and the rheology and biomaterials of the perfusion system. Patient factors also affect the coagulation equilibrium; these include age, infection, history or presence of cardiogenic shock, massive blood losses and transfusions, platelet coagulation deficiencies, fibrinolysis, liver disease, cachexia, reoperation, and hypothermia.

The cornerstone of bleeding management is meticulous surgical hemostasis during all phases of an operation. The surgical techniques, topical agents, and customary drugs used do not need reiteration for trained surgeons. Most cardiac surgical operations involving CPB are accompanied by net blood losses between 200 to 600 mL. Reoperations; complex procedures; prolonged (>3 hours) cardiopulmonary bypass; and patient factors listed above may be associated with excessive and ongoing blood losses. Most surgeons use an antifibrinolytic, such as aprotonin or epsilon amino caproic acid, to reduce fibrinolysis in prolonged or complex operations. Problem patients who bleed excessively after heparin neutralization require an attempt to rebalance pro- and anticoagulants to near normal, pre–CPB concentrations.

The most useful tests in the operating room are an activated clotting time or a protamine titration test to assess the presence of heparin; prothrombin time to uncover deficiency in the extrinsic coagulation pathway; and platelet count. If heparin is neutralized, the partial thromboplastin time may be measured to assess possible deficiency of coagulation proteins. Other tests such as measurements of fibrinogen, template bleeding time, and the thromboelastograph are controversial and/or difficult to obtain. Platelet counts below 80,000 to 100,000/µL should initiate platelet transfusions in bleeding patients, except those with IgG anti-H-PF4 antibodies, to add functioning platelets to the mass of partially dysfunctional platelets.

Measurements of F1.2 and D-dimer are two tests that can be very helpful and probably should be made available on an emergency basis in hospitals that perform complex procedures and offer mechanical circulatory and respiratory assistance. F1.2 measures thrombin formation by factor Xa, and if absent or low, there may be a deficiency in the concentrations of coagulation proteins; fresh frozen plasma is needed. If F1.2 and D-dimer (a measurement of fibrinolytic activity) are both elevated, thrombin is being formed and an antifibrinolytic (aprotinin or epsilon amino caproic acid) is needed to neutralize plasmin. If both markers or F1.2 remain elevated after the antifibrinolytic drug, this indicates continuing thrombin generation and the cause (e.g., infection usually) should be aggressively treated with antibiotics. Some thrombin is needed to stop bleeding, but excessive thrombin production feeds the consumptive coagulopathy. As with diffuse intravascular coagulopathy,483 no guaranteed therapeutic recipe is known; success requires patience, persistence, and judicious use of platelets, antifibrinolytics, fresh frozen plasma, and replacement transfusions to rebalance the coagulation equilibrium at near normal concentrations of the constituents.


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