Inflammatory response to cardiopulmonary bypass

Inflammatory Response to Cardiopulmonary Bypass L. Henry Edmunds, Jr, MD Division of Cardiothoracic Surgery, Hospital of the University of Pennsylvania, Philadelphia, Pennsylvania

This article reviews the roles of the contact and complement systems and of neutrophils and monocytes in the inflammatory response to cardiopulmonary bypass and open heart operation. These blood proteins and cells, together with other blood elements, produce the vasoac-

tive and cytotoxic substances and microemboli that cause the morbidity associated with cardiopulmonary bypass and open heart operation. (Ann Thorac Surg 1998;66:S12– 6) © 1998 by The Society of Thoracic Surgeons

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blood elements: the contact activation system, the complement system, neutrophils, and monocytes. Many nonblood variables affect the magnitude of the inflammatory response, including biomaterials in contact with blood, surface coatings, activation of some proteins by the operation itself, temperature, aortic cross-clamp time, myocardial reperfusion, steroids, antioxidants, and the use of various protease inhibitors that may attenuate the response. With respect to biomaterials, it should be noted that there is no such thing as a nonthrombogenic biomaterial; a biomaterial described as “thromboresistant” may cause clotting more slowly than other materials but does not prevent it. Like thromboresistance, “biocompatibility” is also an imprecise term that suggests some undefined advantage of one material over another.

allikrein is produced by the contact system of plasma proteins and directly mediates neutrophil activation; complement is separately activated by both the classic and alternative pathways. Complement activation produces three anaphylatoxins, powerful vasoactive substances, and the terminal complement complex that lyses cells. Monocytes also are activated during cardiopulmonary bypass (CPB) and open heart operation, in both the wound and the perfusion circuit. Monocytes are very strongly procoagulant and express tissue factor (TF), which initiates the extrinsic coagulation pathway. They also produce numerous proinflammatory and antiinflammatory cytokines, with plasma concentrations of nearly all peaking several hours after CPB. A very large number of vasoactive substances are produced during CPB and open heart operation; these substances cause edema, decreased myocardial contractility, and changes in vascular resistance in various vascular beds. Preventing or attenuating activation of these blood elements during open heart operation offers a means of substantially reducing the morbidity associated with this technology. Cardiopulmonary bypass is not possible without heparin, but it is not an ideal anticoagulant. The coagulation cascade is an amplification system in which one protease begets thousands of copies of the next protease, which then begets thousands more of the next enzyme in the chain. Heparin is not an ideal anticoagulant because it inhibits coagulation at the end of the coagulation cascade rather than at the beginning. Thus many powerful proteases are produced before heparin inhibits clot formation. Cardiopulmonary bypass activates five plasma protein systems: contact, intrinsic coagulation, extrinsic coagulation, complement, and fibrinolytic (Table 1). Blood cells activated by CPB are platelets, neutrophils, monocytes, endothelial cells, and lymphocytes. Activation of these blood elements mediates the principal complications of CPB: bleeding, thromboembolism, fluid retention, and temporary organ dysfunction [1]. The inflammatory reaction principally involves four Presented at “Risk Management in CABG: Significant Surgical Considerations,” New Orleans, LA, Jan 24, 1998. Address reprint requests to Dr Edmunds, Division of Cardiothoracic Surgery, Hospital of the University of Pennsylvania, Philadelphia, PA 19104.

© 1998 by The Society of Thoracic Surgeons Published by Elsevier Science Inc

The Contact System The contact system of plasma proteins consists of factor XII (Hageman factor), prekallikrein, high-molecularweight kininogen, and factor XI. On negatively charged surfaces, factor XII is autoactivated and cleaves into two serine proteases, factor XIIa and factor XIIf. Factor XIIa activates factor XIa and initiates the intrinsic coagulation pathway. Factor XIIa also activates prekallikrein to form kallikrein and high-molecular-weight kininogen to form bradykinin [2]. Kallikrein is a major mediator that directly activates neutrophils and facilitates cleavage of factor XII in a feedback loop (Fig 1). Kallikrein cannot be measured directly, but is measured by measuring the kallikrein–C1 inhibitor complex. In vitro recirculation of heparinized fresh human blood demonstrates sharp increases in the kallikrein–C1 inhibitor complex and the

Table 1. Plasma Protein Systems and Blood Cells Activated by Cardiopulmonary Bypass Plasma Protein Systems Contact Intrinsic coagulation Extrinsic coagulation Fibrinolytic Complement

Blood Cells Platelet Endothelial cell Neutrophil Monocyte Lymphocyte

0003-4975/98/$19.00 PII S0003-4975(98)00967-9

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reason for hypotension coincidental with protamine in approximately 50% of patients.

Neutrophil Activation

Fig 1. Factor XII (FXII) in the presence of prekallikrein and highmolecular-weight kininogen (HMWK) is cleaved into active fragments, FXIIa and FXIIf (fragments, not shown). FXIIa acting on FXI initiates the intrinsic coagulation cascade. Cleavage of HMWK produces bradykinin. Neutrophils are activated by kallikrein, complement, and other agonists; fibrinolysis is initiated by generation of thrombin, which stimulates production of tissue plasminogen activator by endothelial cells. Complement is not activated by contact system proteins. (HLE 5 human leukocyte elastage; TF 5 tissue factor.)

C1–C1 inhibitor complex. C1 is the first protein in the complement system.

Complement Complement is activated by both the classic and alternative pathways during CPB. The initial activation of blood in the nonendothelial cell perfusion circuit probably occurs by the classic pathway. However, a small amount of C3a formed by the classic pathway initiates activation of the alternative pathway that has a feedback loop. The classic pathway is also activated by the heparin–protamine complex when protamine is given. This causes a big spike in complement activation and is the most likely

Complement has three anaphylatoxins, C3a, C4a, and C5a, which are powerful vasoactive substances. After C5a is produced, C5b forms the terminal complement complex (C5b through C9). Years ago, Craddock and coworkers [3] photographed the response of neutrophils when exposed to activated complement; within 20 seconds of exposure, docile neutrophils turn into very angry cells, expressing pseudopods and releasing granule contents (Fig 2). The neutrophil is the killer cell of the defense reaction and contains many cytotoxic enzymes: neutrophil elastase, cathepsin G, lysozymes, and myeloperoxidase. Neutrophil elastase is a powerful enzyme that, among other actions, has an important role in the development of emphysema, especially in smokers. Neutrophil elastase is released during open heart operation; levels of elastase increase during CPB and return toward baseline over 24 hours (Fig 3) [4]. Neutrophils also produce oxygen free radicals, hydrogen peroxide, and hypobromous acid; all are extremely cytotoxic. Activated complement and neutrophils both increase perivascular edema by directly attacking vein walls. Although both complement and neutrophil activation are important, the neutrophil is the main target in efforts to control the inflammatory response associated with CPB and open heart operation. As noted above, there are many factors that can affect complement, and it does not remain in the circulation long after CPB ends. Neutrophils pose a major challenge and, unfortunately, we do not have a feasible way to inhibit neutrophil activation during open heart operation.

Monocytes Monocytes also are activated in the inflammatory response. In an in vitro perfusion circuit monocytes appear to be activated more slowly than contact and complement proteins and neutrophils. When fresh, heparinized

Fig 2. Activation of neutrophils by activated complement. (Reproduced from [3].)

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activity in the wound promptly and without the delay observed in the in vitro perfusion circuit (Fig 5). Simultaneous blood samples obtained from blood around the heart and from the perfusion circuit showed that pericardial samples contained increased amounts of F1.2 (a marker of thrombin formation), activated factor VII, and monocytes that expressed TF as compared with perfusate samples [6]. Monocytes express TF both in the wound and in the perfusion circuit and are strongly procoagulant. Expression of TF initiates the extrinsic coagulation pathway. Monocytes also produce numerous proinflammatory and antiinflammatory cytokines; the literature documents increases in interleukins IL-1, IL-2, IL-4, IL-6, IL-8, IL-10, and even IL-12, which acts as an inhibitor, during or after CPB. The literature is ambivalent about tumor necrosis factor, with nearly as many studies showing that it is not increased as there are showing that it is increased. Plasma concentrations of nearly all cytokines peak several hours after CPB [7]; thus, the impact of circulating cytokines is largely in the early postoperative period.

Vasoactive Substances Associated With CPB A vasoactive substance may be defined as any substance that causes vascular smooth muscle to contract or relax, that causes an endothelial cell to contract or relax, or that influences myocyte contractility [8]. Large numbers of vasoactive substances are produced or affected by CPB and open heart operation, and the list in Table 2 is only a partial one. These substances produce edema, decrease myocardial contractility, and change vascular resistance in various vascular beds. The net result is that Claude Bernard’s postulated homeostasis is supplanted by chaos. Additionally, the coagulation pathways, destruc-

Fig 3. Neutrophil counts and plasma levels of neutrophil lactoferrin and human neutrophil elastase during clinical cardiopulmonary bypass (CPB) are shown. Each point is the mean and standard error of the mean of 10 patients (a 5 before induction of anesthesia; b 5 after anesthesia, before heparin; c 5 after heparin, before CPB; d 5 5 minutes after start of CPB; e 5 45 minutes after start of CPB; f 5 just before CPB ended; g 5 60 minutes after CPB; h 5 24 hours after CPB.) (Reproduced from [4] with permission of Lippincott Williams & Wilkins.)

human blood is recirculated at 37° or 28°C in an in vitro system for up to 6 hours, monocyte expression of TF and the Mac-1 receptor is delayed (Fig 4) [5]. Monocytes attached to the tubing of the circuit are highly activated when studied at the end of 6 hours of recirculation. In a patient study, monocytes expressed TF and procoagulant

Fig 4. Bar graph showing monocyte and lymphocyte surface Mac-1 fluorescence during simulated extracorporeal circulation. The expression of monocyte Mac-1 was measured by reactivity with monoclonal antibody LM2 to the (a chain of the leukocyte integrin Mac-1). Lymphocytes were negative. (R 5 recovered cells; SC 5 standing control cells.) (Reproduced from [5] with permission of Lippincott Williams & Wilkins.)

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Interestingly, no one has demonstrated an increase in intracellular fluid associated with CPB; the fluid increase is all extravascular or intravascular, and most of it is extravascular. This edema and multiple emboli produce dysfunction of virtually all organs, including the heart, lungs, kidneys, central nervous system, pancreas, and liver. Fortunately, the dysfunction is usually temporary, without detectable permanent effects. However, sophisticated neuropsychologic tests done 6 weeks, 6 months, and even 1 year postoperatively consistently show a detectable decrease in function in up to 60% of patients who have had an open heart operation [9].

Inhibiting the Inflammatory Response

Fig 5. Tissue factor (TF) expressed by monocytes during and after cardiopulmonary bypass (CPB) and open heart operation. Values are the mean and standard error of the mean and represent the percentage of cells that express TF compared with monocytes maximally stimulated by lipopolysaccharide. Samples were taken after heparin infusion, before CPB; 30 – 45 minutes after start of CPB from the perfusate simultaneously with the pericardial blood sample; within 5 minutes after CPB; and 15 minutes after protamine infusion. (*p , 0.05 for paired t test statistic versus the heparin sample; †, p , 0.05 for paired t test statistic versus CPB perfusate sample.) (Reproduced from [6] with permission of Mosby, Inc.)

tion of blood cells, the wound, and the heart-lung machine produce numerous emboli of fibrin, fat, platelets, leukocytes, aggregates of platelets and white cells, red cell debris, gas, foreign material, and spallated particles. Although emboli larger than 40 mm are removed by the arterial filter, the patient’s body is showered by many emboli smaller than 40 mm. As capillaries are only about 8 to 10 mm in diameter, these emboli obstruct small, diffusely distributed vascular beds that supply small numbers of cells. The damage is largely unnoticed: a few cells die in hundreds of locations throughout the entire body. Widely distributed cell necrosis and the increase in capillary permeability increase interstitial fluid. Fluid balance also is disturbed by an increase in systemic venous pressure and decreased colloid osmotic pressure produced by hemodilution. Consequently, interstitial fluid is vastly increased in proportion to the magnitude and duration of the operation. Whereas a first-time revascularization procedure with a relatively short pump run and small wound does not elicit much of an inflammatory response, a fourth-time redo with a bad heart that involves multiple grafts or multiple valve procedures elicits a large inflammatory response.

A number of studies have shown that a heparin-bonded surface attenuates terminal complement complex formation during CPB [10]. Other studies have shown attenuation of myeloperoxidase and lactoferrin release by neutrophils [11]. In a clinical study we observed attenuation of neutrophil elastase release [12]. To some degree, heparin-coated surfaces of the perfusion circuit attenuate the inflammatory response to CPB but do not obliterate it. Whether this attenuation is sufficient to justify the considerable expense remains to be resolved. In a study at Boston University, Aldea and associates [12] used a heparin-coated perfusion circuit and halfdose systemic heparin in a large number of first-time revascularization patients. Half-dose systemic heparin with activated clotting times less than 300 seconds is dangerous because increased thrombin is produced and circulated. Thrombin is a dangerous enzyme to circulate; it has many actions. These patients did well, however, because the surgeons did not return blood aspirated from the wound to the perfusate. All blood aspirated from the wound was put into the cell-saving device, washed, and returned as packed red cells to the circulation. This substantially reduced the amount of thrombin added to

Table 2. Vasoactive Substances Altered During Cardiopulmonary Bypass Epinephrine

Thyroid: T3, T4

Norepinephrine

Electrolytes: Ca21, Mg21, K1 Complement: C3a, C4a, C5a Membrane attack complex, C5b9 Oxygen free radicals Lysosomal enzymes Proteases, cathepsins

Renin Angiotensin II Vasopressin Aldosterone Atrial natriuretic factors Glucagon

Leukotrienes LTB4, LTC4, LTD4 Interleukins IL-1, IL-2, IL-4, IL-6, IL-8, IL-10, IL-12

Platelet-activating factor Prostacyclin Thromboxane A2 Prostaglandin E2 Nitric oxide Endothelin-1 Serotonin Histamine

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the perfusate from the wound. The study convincingly showed a reduction in the inflammatory response using these clinical perfusion circuits. Aldea and colleagues [12] observed reductions in ventilatory requirements, hospital stays, and other benefits. It has been suggested that low-dose aprotinin reduces the inflammatory response. In my view, any effect of aprotinin on the inflammatory response is probably very small. Aprotinin inhibits both plasmin and kallikrein, but the inhibitor dose for kallikrein is 100 times greater than that for plasmin. Thus, 100 times more aprotinin is required to inhibit kallikrein than to inhibit fibrinolysis. High doses of aprotinin may attenuate some of the inflammatory response, but the drug is not potent enough to have much impact. Considerable research is under way on the antiinflammatory effects of protease inhibitors during CPB. At present nafamostat is the most promising among the six or seven protease inhibitors that we have evaluated [13]. During in vitro recirculation of fresh heparinized human blood, nafamostat completely inhibited kallikrein, factor XIIa formation, and neutrophil elastase release, but did not inhibit complement activation. Although complement activates neutrophils, the fact that elastase release was inhibited underscores the fact that neutrophils are activated by many substances other than complement. These include kallikrein, platelets, neutrophil-activating proteins 1 and 2, and cathepsin. Eventually, we will probably be more successful in inhibiting the inflammatory response of CPB with protease inhibitors than with circuit-surface coatings. Protease inhibitors are abundant and are likely to be more effective, more specific, and cheaper than surface coatings. As increasing numbers of investigators address the effects of protease inhibitors on the inflammatory response, drugs that temporarily shut down activation of the four main blood components involved will be found. This will make open heart operations essentially comparable to surgical procedures done without the heart-lung machine.

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