(Chest. 2003;123:1355-1360.)
© 2003
American College of Chest Physicians
Transcardiac Release of Soluble Adhesion Molecules During Coronary Artery Bypass Grafting*
Effects of Crystalloid and Blood Cardioplegia
Ryszard Kalawski, MD, PhD;
Marcin Majewski, MD;
Eugeniusz Kaszkowiak, MD;
Henryk Wysocki, MD, PhD and
Tomasz Siminiak, MD, PhD
* From the Department of Cardiac Surgery, J. Strus Hospital Poland, Poznan; and University School of Medical Science, Department of Cardiology, District Hospital, Poznan, Poland.
Correspondence to: Tomasz Siminiak, MD, PhD, Department of Cardiology, University School of Medical Science, District Hospital, 7/19 Juraszow St, 60-479 Poznan, Poland; e-mail: tomasz.siminiak{at}usoms.poznan.pl
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Abstract
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Background: Dysfunction of myocardium as a result of ischemia/reperfusion during coronary artery bypass grafting (CABG) is currently one of the biggest problems in cardiovascular surgery. In previous studies, it has been well established that activated leukocytes and coronary vascular endothelial cells play an important role in the development of cardiac tissue damage during ischemia followed by reperfusion. Interactions between both of these cell types require the expression of adhesion molecules on their surface. In certain conditions, on cell activation, the adhesion proteins may be released from activated cells in soluble form into circulation. The purpose of our study was to establish whether the use of blood cardioplegia modifies plasma levels of soluble intracellular adhesion molecule-1 (sICAM-1), soluble vascular cell adhesion molecule-1 (sVCAM-1), soluble E-selectin (sE-selectin), and soluble L-selectin (sL-selectin) in comparison with crystalloid cardioplegia in patients undergoing CABG.
Methods: Patients undergoing CABG were classified into two groups to receive cold crystalloid cardioplegia (St. Thomas’ Hospital) or cold blood cardioplegia (method of Buckberg), followed by a "warm-shot" of the solution. Coronary sinus and arterial blood samples were obtained from 50 patients (42 men and 8 women; age range, 34 to 73 years) before aortic cross-clamping, at the beginning of reperfusion, and after 30 min of reperfusion. Plasma levels of soluble adhesion molecules were measured using sensitive enzyme-linked immunosorbent assays.
Results: The transcardiac release of sICAM-1 and sVCAM-1 following myocardial ischemia/reperfusion during CABG was evident in both groups of patients. However, the increase of soluble forms of both of these adhesion proteins was more significant in the group of patients receiving crystalloid cardioplegia. Crystalloid cardioplegia resulted in decreased plasma level of sE-selectin in the coronary sinus blood sample after 30 min of reperfusion. No significant changes in plasma levels of sL-selectin in either group were observed.
Conclusion: Cardioplegia may affect the release of soluble forms of adhesion molecules from ischemic myocardium and modify endothelium activation in patients undergoing CABG.
Key Words: blood cardioplegia • coronary artery bypass grafting • crystalloid cardioplegia • E-selectin • L-selectin • soluble intracellular adhesion molecule-1 • vascular cell adhesion molecule-1
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Introduction
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Systolic dysfunction of myocardium following coronary artery bypass grafting (CABG) is currently the most frequent cause of postoperative mortality in cardiac surgery. It is obviously a result of ischemic heart disease itself, but also an effect of cardiac injury subsequent to ischemia and reperfusion during and after the main part of the procedure.
Open-heart surgery induces an inflammatory response in a human heart. There is clinical and experimental evidence indicating that polymorphonuclear neutrophils (PMNs) and endothelial cells (ECs) play a prominent role in the inflammatory component of postischemic injury. Activation of PMNs and ECs initiates a specific cascade of cell-cell interactions leading first to adhesion of PMNs to vascular endothelium, then to coronary capillary plugging (no-reflow) and reduction in coronary blood flow, resulting eventually in myocardial infarction and apoptosis. Moreover, activated PMNs are a rich source of free oxygen radicals and proteolytic enzymes capable of damaging myocardial cell membranes.1
2
The interactions between monocytes, PMNs, and ECs are dependent on adhesion proteins expressed on their surface. These glycoproteins are members of three major families of adhesion molecules: the integrins, the Ig superfamily, and the selectin family. It has been shown that administration of antibodies that block the function of adhesion molecules, its complement fragments, and receptors attenuate PMN-mediated tissue destruction during CABG with cardiopulmonary bypass (CPB).3
Some of them are well-known markers for cell activation, including soluble intracellular adhesion molecule-1 (sICAM-1), soluble vascular cell adhesion molecule-1 (sVCAM-1), and the surface markers L-selectin (expressed by neutrophils) and E-selectin (expressed by endothelium).
In certain pathologic states including attacks of angina pectoris, acute myocardial infarction, and reperfusion during CABG, plasma contains a variety of stimuli capable of PMN and EC activation.2
3
4
In these conditions, adhesion molecules of PMNs and ECs may be shed from both of these cells on their activation into circulating blood and detected there in the soluble forms. They may be involved in the immunologic response to myocardial damage5
and their plasma levels indicate the intensity of activation of PMNs and ECs during reperfusion of ischemic myocardium.4
This study was designed to demonstrate the effects of crystalloid and blood cardioplegia on activation of PMNs and ECs during CABG by evaluation of transcardiac release of the soluble forms of adhesion molecules.
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Materials and Methods
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A group of 50 patients with coronary artery disease scheduled for CABG on routine clinical and angiographic criteria were evaluated in this study. The exclusion study criteria were a history of heart surgery, a history of unstable angina pectoris and myocardial infarction within the last 3 months, diabetes mellitus, renal failure, the use of any pharmacologic inotropic support or intra-aortic balloon pumping during the procedure, CABG with addition of other heart surgery, presence of inflammatory diseases, and other known factors able to affect PMN activation.
The group was composed of 8 women and 42 men between the ages of 34 years and 73 years (mean, 53 years). The ejection fraction estimated by echocardiography a day before the procedure ranged from 28 to 66% (mean, 46%). The bypass circuit period ranged from 51 to 182 min (mean, 86 min), and aortic cross-clamping time was 17 to 64 min. The hematocrit value during the operation was kept at the range of 22 to 25%, and potassium level was from 5.5 to 6.0 mmol/L. The surgical intervention was performed with CPB at moderate hypothermia from 30 to 32°C with hemodilution. All the patients were operated on by the same team to avoid a possible effect of the maneuvers on the results.
The patients were randomly assigned to one of two groups. In one group, the infusion of cold (4°C) crystalloid St. Thomas’ Hospital cardioplegic solution (Plegisol [Polfa; Kutno, Poland], containing NaCL, 110 mmol/L; KCl, 16 mmol/L; MgCl2, 16 mmol/L; CaCl2, 1.2 mmol/L) was used for preservation of myocardium. In the other group, cold (4°C) blood cardioplegia according to Buckberg (blood and crystalloid solution in 4:1 ratio) was used, followed by "warm-shot" of the blood cardioplegia. No statistically significant differences in mean age, sex, left ventricular ejection fraction, CPB time, and severity of angina pectoris (Canadian Cardiovascular Society) were observed.
The blood samples were collected from radial artery and coronary sinus directly before myocardial ischemia (aortic cross-clamping), at the beginning of reperfusion (aortal clamp release), and 30 min after the beginning of reperfusion. The samples in tubes containing heparin (5 IU/mL) were centrifuged and immediately deeply frozen (- 70°C) until the moment of analysis of all plasma samples.
Informed consent to participate in the study was obtained from each subject, and the protocol was accepted by the local ethics committee. The plasma levels of soluble adhesion molecules were measured using sensitive enzyme-linked immunosorbent assays (R&D Systems Europe; Abingdon, England). The data were not normally distributed as assessed by the Kolgomorov-Smirnov test. Statistical analysis was performed by the nonparametric repeated-measures analysis (the Friedmann test) of variance within groups and with the Mann-Whitney test for comparisons between groups. The results are shown as median values (the first quartile and the third quartile).
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Results
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The increase of plasma level of sICAM-1 and sVCAM-1 in the coronary sinus during CABG was observed in both groups of patients (Fig 1 , 2
). However, the increase of concentration of soluble forms of both of these proteins was more evident in the group receiving crystalloid cardioplegic solution. In the group receiving crystalloid cardioplegia, the concentrations of sICAM-1 and sVCAM-1 in the coronary sinus were significantly (p < 0.05) higher 30 min after reperfusion (243 ng/mL [range, 183 to 300 ng/mL] and 1,450 ng/mL [range, 1,199 to 1,881 ng/mL]) than at the aortic clamp release (211 ng/mL [range, 148 to 262 ng/mL] and 815 ng/mL [range, 581 to 1,005 ng/mL]), respectively. They were higher in blood obtained from the coronary sinus 30 min after reperfusion than from peripheral artery at the same time (sICAM-1, 199 ng/mL [range, 140 to 249 ng/mL] and sVCAM-1, 1,100 ng/mL [range, 705 to 1,327 ng/mL]). In the group of patients receiving blood cardioplegia, the significantly lower concentration of both of these proteins in coronary sinus and arterial blood was observed in comparison with the group of crystalloid cardioplegia 30 min after reperfusion (Fig 1
, 2)
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Figure 1. Plasma levels of sICAM-1 in patients undergoing CABG, receiving crystalloid (CR) and blood (BL) cardioplegia. Samples were obtained from the peripheral (periph) artery and the coronary sinus before aortal cross-clamping, at the beginning of reperfusion (reperf), and 30 min thereafter. *p < 0.05 vs artery; #p < 0.05 vs beginning of reperfusion and before ischemia; °p < 0.05 vs crystalloid. The results are shown as median values ± the first and the third quartile.
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Figure 2. Plasma levels of sVCAM-1 in patients undergoing CABG, receiving crystalloid and blood cardioplegia. Samples were obtained from peripheral artery and the coronary sinus before aortal cross-clamping, at the beginning of reperfusion, and 30 min thereafter. *p < 0.05 vs artery; #p < 0.05 vs before ischemia; °p < 0.05 vs crystalloid. The results are shown as median values ± the first and the third quartile. See Figure 1
for expansion of abbreviations.
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In the group of patients receiving crystalloid cardioplegic solution, the plasma level of soluble E-selectin (sE-selectin) in the coronary sinus was markedly (p < 0.05) lower 30 min after reperfusion (30 ng/mL; range, 24.6 to 37.1 ng/mL) than at the beginning of reperfusion (42.2 ng/mL; range, 33.4 to 49.6 ng/mL). Thirty minutes after reperfusion, the concentration of sE-selectin in coronary sinus was decreased in comparison with its plasma level in the peripheral arterial blood sample at the same time. The essential changes of sE-selectin plasma level in arterial blood were observed in the group of crystalloid cardioplegia. The significant increase of its concentration 30 min after reperfusion (44.1 ng/mL; range, 35.6 to 52.3 ng/mL) vs the beginning of reperfusion (32.3 ng/mL; range, 25.7 to 40.7 ng/mL) was noted (Fig 3
).
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Figure 3. Plasma levels of sE-selectin in patients undergoing CABG, receiving crystalloid and blood cardioplegia. Samples were obtained from peripheral artery and the coronary sinus before aortal cross-clamping, at the beginning of reperfusion, and 30 min thereafter. *p < 0.05 vs artery; #p < 0.05 vs beginning of reperfusion. The results are shown as median values ± the first and the third quartile. See Figure 1
for expansion of abbreviations.
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No significant changes in sE-selectin plasma level were found in the group of blood cardioplegia in arterial or coronary sinus blood samples throughout the procedure. No differences in mean concentration of soluble L-selectin (sL-selectin) were observed in both groups of patients with crystalloid and blood cardioplegia at any time during procedure (Fig 4
). In the presented group of patients, a trend toward the increased release of molecules during longer time of CPB in both cardioplegia groups was noted; however, no statistically significant relationship was observed.
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Figure 4. Plasma levels of sL-selectin in patients undergoing CABG, receiving crystalloid and blood cardioplegia. Samples were obtained from peripheral artery and the coronary sinus before aortal cross-clamping, at the beginning of reperfusion, and 30 min thereafter. The results are shown as median values ± the first and the third quartile. See Figure 1
for expansion of abbreviations.
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Discussion
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The involvement of leukocytes in the development of myocardial injury, particularly during ischemia followed by reperfusion, is well established. The activation of PMNs and ECs during coronary angioplasty and open-heart surgery with CPB is a well-known phenomenon.6
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8
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The activation of PMNs during CABG is not only an effect of blood contact with synthetic surfaces of CPB circuit,6
7
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but also a result of their direct interactions with activated ECs as well as an effect of systemic PMN activation by several stimuli appearing in circulating blood during the procedure.2
3
8
Plasma obtained from the coronary sinus at the beginning of reperfusion (at aortic clamp release or a balloon deflation during percutaneous transluminal coronary angioplasty) showed its ability in up-regulation of the neutrophilic ß-integrin family of adhesion molecules and stimulation of PMNs to release of cytotoxic free oxygen radicals.2
11
Activated endothelium, platelets, and PMNs themselves are the source of mediators that may affect the activation of one another.1
Several experiments and clinical studies have shown that different methods resulting in depletion of PMNs, including neutrophil antiserum, leukocyte filters, and pharmacologic interventions as monoclonal antibodies against leukocyte adhesion molecules may reduce the extent of myocardial injury.1
3
4
11
The adhesion molecules expressed on the surface of neutrophils and endothelial cells, on their activation, may be shed and be released into circulating blood and detected there in the soluble forms. Significant changes in plasma levels of sICAM-1, sVCAM-1, and sL-selectin have been confirmed during attacks of angina pectoris and acute myocardial infarction.3
5
In the current study, we estimated the transcardiac changes of sICAM-1, sVCAM-1, sE-selectin, and sL-selectin in plasma in patients subjected to CABG. Moreover, we verified the possibility of modification of the release of soluble forms of adhesion molecules, known as markers of cell activation, by the cardioplegic solution used during CABG. We demonstrate the transcardiac release of sICAM-1 and sVCAM-1 during CABG with CPB in patients with both crystalloid and blood cardioplegia; however, the increase of concentration of the soluble forms of both of these proteins was less significant in the group of blood cardioplegia. The considerable decrease of sE-selectin plasma level in coronary sinus after 30 min of reperfusion was noted in the group of crystalloid cardioplegia, and there were no substantial changes in sE-selectin plasma level in the group of blood cardioplegia. The sL-selectin plasma concentration remain unchanged in both groups of patients.
At this stage of investigation, the exact mechanism responsible for the differences between groups receiving crystalloid and blood cardioplegia is unclear. It has been previously shown that the use of blood cardioplegia may reduce the release of pro-inflammatory cytokines in patients undergoing cardiac surgery12
13
and affect the release of PMN-oriented stimuli from ischemic myocardium during the CABG procedure. PMNs from healthy donors, incubated with plasma obtained from the coronary sinus immediately after reperfusion during CABG in patients receiving crystalloid cardioplegia, showed the increase in superoxide anion production in comparison with PMNs incubated with plasma obtained from patients receiving blood cardioplegia.2
The differences in increase of sICAM-1 and sVCAM-1 plasma concentration in both groups of patients indicate that cardioplegia itself may influence the release of soluble adhesion molecules and thus modify EC activation.
In a previous study, it has been observed that L-selectin expression on the surface of PMNs obtained from healthy donors incubated with patient’s plasma obtained from the coronary sinus after 30 min of reperfusion (after release of aortic cross-clamping in patients undergoing CABG) was decreased in comparison with its expression on PMNs incubated with plasma sample obtained from the coronary sinus at the beginning of reperfusion and from peripheral artery after 30 min of reperfusion.11
This may suggest the release of mediators inducing shedding of neutrophilic L-selectin from ischemic and reperfused myocardium. We speculate that the unchanged level of sL-selectin as well as the reduced plasma level of sE-selectin in coronary sinus during reperfusion in the current study may be associated with the balance between their increased shedding from activated PMNs and ECs, and tissue expression of counter receptors for these selectins appearing on PMNs, monocytes, lymphocytes and/or endothelium following CABG.4
5
14
Any pharmacologic interventions able to reduce EC and PMN activation during ischemia and reperfusion have the potential to exert a beneficial effect on myocardial function after heart surgery. In our current study, we evaluated the typical protocol of blood cardioplegia used routinely in many cardiac surgery centers, including a terminal warm-shot of the solution and the standard crystalloid cardioplegia protocol (ie, not including the warm shot). Although it has been previously reported that the type of cardioplegic solution may affect the inflammatory reactions during reperfusion and the extent of myocardial damage,13
15
16
17
it should be noted that the temperature of cardioplegia itself or a terminal warm-shot may also attenuate the protective effect on myocardium.16
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No clear difference has been observed in the clinical outcome between the both groups in our study, probably as a result of limited number of the patients. The results however support the possible superior cytoprotective effect of blood cardioplegia in coronary bypass surgeries.15
16
17
22
23
The clinical relevance of the finding still requires further investigation.
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Conclusion
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The transcardiac release of soluble forms of adhesion molecules ICAM-1, VCAM-1, and E-selectin during CABG, and thus endothelium activation, may be modified by the cardioplegic solution; however, the exact mechanism responsible for differences between the study groups remains to be elucidated.
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Footnotes
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Abbreviations: CABG = coronary artery bypass grafting; CPB = cardiopulmonary bypass; EC = endothelial cell; PMN = polymorphonuclear neutrophil; sE-selectin = soluble E-selectin; sICAM-1 = soluble intracellular adhesion molecule-1; sL-selectin = soluble L-selectin; sVCAM-1 = soluble vascular cell adhesion molecule-1
The study was supported by a governmental grant from Komitet Badan Naukowych.
Received for publication January 17, 2002.
Accepted for publication September 23, 2002.
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References
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Abstract
Introduction
