Ann Thorac Surg 1997;64:1360-1367
© 1997 The Society of Thoracic Surgeons
Original Articles: Cardiovascular
Comparative Effects of Continuous Warm Blood and Intermittent Cold Blood Cardioplegia on Coronary Reactivity
Motohisa Tofukuji, MD, PhD,
Alon Stamler, MD,
Jianyi Li, MB, MS,
Mukesh D. Hariawala, MD,
Alvin Franklin, MS,
Frank W. Sellke, MD
Division of Cardiothoracic Surgery, Department of Surgery, Beth Israel–Deaconess Medical Center and Harvard Medical School, Boston, Massachusetts
Accepted for publication June 2, 1997.
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Abstract
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Background. Cardioplegia is known to affect coronary vascular reactivity. We examined the effects of intermittent cold and continuous warm blood cardioplegia on ß-adrenoceptor–mediated, adenosine triphosphate–sensitive K+ (K+ATP)–channel–mediated, and endothelium-dependent relaxation and on the myogenic tone of coronary arterioles.
Methods. Pigs were placed on cardiopulmonary bypass. Hearts were arrested for 1 hour with a cold blood cardioplegic solution administered intermittently (n = 12; iCB-CP) or with a warm blood cardioplegic solution delivered continuously (n = 12; cWB-CP). Selected hearts (n = 6 in each group) were then reperfused for 1 hour. In vitro relaxation responses of precontracted microvessels (50 to 160 µm) were studied in a pressurized no-flow state.
Results. Relaxation in response to isoproterenol (ß-adrenergic agonist) was similar after iCB-CP and cWB-CP, whereas forskolin (adenylate cyclase activator)–induced relaxation was impaired more after iCB-CP than after cWB-CP. After reperfusion the respective responses were similar. Both iCB-CP and cWB-CP preserved receptor-mediated, endothelium-dependent relaxation in response to adenosine, 5`-diphosphate; non–receptor-mediated endothelium-dependent relaxation in response to A23187; endothelium-independent cyclic guanosine monophosphate–mediated relaxation in response to sodium nitroprusside, and K+ATP-channel–mediated relaxation. Relaxations in response to 8-bromo-cyclic guanosine monophosphate (a cyclic guanosine monophosphate–dependent protein kinase activator) and to 8-bromo-cyclic adenosine monophosphate (a cyclic adenosine monophosphate–dependent protein kinase activator) were impaired after iCB-CP alone and after reperfusion, whereas the respective responses were not affected after cWB-CP. Myogenic tone was decreased similarly after iCB-CP and cWB-CP but was not further altered after reperfusion. Cardiac function was similar after iCB-CP and cWB-CP.
Conclusions. These results suggest that cWB-CP is similar to iCB-CP in its ability to preserve endothelium-dependent relaxation and K+ATP-channel function. The superior preservation of ß-adrenergic-cyclic adenosine monophosphate–mediated coronary responses after cWB-CP is brief and associated with minimal improvement of myocardial function and myogenic tone.
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Introduction
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Recent studies have shown that the endothelial and vascular smooth muscle regulation of coronary perfusion may be altered after cardioplegia and subsequent reperfusion. This may result in vascular spasm or contraction and impair optimal recovery of myocardial function after cardiac operations. A variety of cardioplegic solutions have been developed and are in clinical use. However, it is unclear which cardioplegic solution is optimal for the protection of both the coronary circulation and myocardium.
Previous studies from our laboratory and those of others have shown that reduced ß-adrenergic–mediated relaxation [1], impaired endothelium-dependent relaxation [2, 3], and reduced myogenic tone [1, 4] occur in the coronary circulation after hyperkalemic crystalloid cardioplegia. The addition of blood [1, 3, 5, 6] or magnesium [7] to a cold crystalloid cardioplegic solution can improve these indices of vascular recovery after ischemic arrest. Several investigators [8, 9] have reported that cold and warm blood cardioplegia may preserve coronary endo-thelium–dependent relaxation. The effects of intermittent cold blood (iCB-CP) and continuous warm blood (cWB-CP) cardioplegia on ß-adrenergic–mediated relaxation, myogenic tone, and other indices of vascular function need to be compared.
Thus the objective of this study was to compare the effects of cWB-CP and iCB-CP on ß-adrenergic–mediated, endothelium-dependent, adenosine triphosphate–sensitive K+ (K+ATP)–channel–mediated, and endothelium-independent relaxation and on myogenic reactivity and to correlate these effects on vascular function with left ventricular functional recovery.
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Material and Methods
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Animal Preparation
Yorkshire pigs (weight, 20 to 25 kg) of either sex were premedicated with ketamine (10 mg/kg intramuscularly) and anesthetized with 
-chloralose and urethane (60 and 300 mg/kg intravenously initially and 15 and 60 mg/kg every 60 minutes as needed, respectively). Pigs were intubated and mechanically ventilated. In the control group (n = 6), a sternotomy was performed and the pig was heparinized (500 units/kg). The heart was rapidly excised and immediately placed in a cold (5° to 10°C) Krebs' buffer solution of the following composition (in millimoles per liter): NaCl, 118.3; KCl, 4.7; CaCl2, 2.5; MgSO4, 1.2; NaH2PO4, 1.2; NaHCO3, 25; and glucose, 11.1.
In 12 pigs, after the induction of anesthesia and tracheal intubation, a fluid-filled catheter was introduced from the femoral artery for the measurement of peripheral arterial pressure. After a sternotomy was performed, pigs were heparinized (500 units/kg) and cannulated through the distal ascending aorta and the right atrium. A left ventricular vent was placed through the left ventricular apex for decompression. A cannula was inserted into the ascending aorta through a pursestring suture for the infusion of a cardioplegia solution. Total cardiopulmonary bypass (CPB) was instituted using a bubble oxygenator (Bentley Bio-2; Baxter Healthcare Corp, Irvine, CA) and a standard roller pump. An arterial filter (Bentley Bio-1025; Baxter Healthcare Corp) was inserted into the circuit distal to the roller pump. Blood flow was maintained at a rate of 2.0 to 3.0 L/min (2.6 to 4.2 L • min-1 • m-2) to maintain a mean perfusion pressure of 50 to 70 mm Hg. The systemic blood temperature was maintained at 37°C throughout the experiment. Arterial blood gas values were determined before the start of CPB and at approximately 20-minute intervals thereafter. The ventilatory rate, tidal volume, and the fraction of inspired oxygen were adjusted to maintain the arterial oxygen tension at more than 100 mm Hg, the pH at between 7.35 and 7.45, and the arterial carbon dioxide tension at more than 30 and less than 45 mm Hg.
Cold Cardioplegia Group
In 6 pigs, after stabilization of the preparation for CPB, an aortic cross-clamp was placed and 150 mL of a high-potassium cold (0° to 4°C) blood cardioplegic solution (4:1 mixture of oxygenated blood with a hyperkalemic crystalloid cardioplegic solution resulting in 25 mmol/L KCl) was infused into the aortic root at a perfusion pressure of 60 mm Hg, followed by an intermittent infusion of 150 mL of a low-potassium cold blood cardioplegic solution (4:1 mixture of oxygenated blood with a hyperkalemic crystalloid cardioplegic solution resulting in 12 mmol/L KCl) every 20 minutes (iCB-CP group). The composition of the high- and low-potassium crystalloid cardioplegic solution used in this study was 25 or 12 mmol/L KCl, respectively, together with 5 g/L mannitol, 20 mL/L citrate phosphate dextrose solution, 4 mmol/L THAM (trihydroxy methylamino methane) in 5% dextrose, and 0.225% saline solution. Saline slush was placed on the surface of the heart to provide topical hypothermia during the cross-clamp period.
Warm Cardioplegia Group
In 6 pigs, after stabilization of the preparation for CPB, an aortic cross-clamp was placed and 150 mL of a high-potassium warm (35° to 37°C) blood cardioplegic solution (4:1 mixture of oxygenated blood with a hyperkalemic crystalloid cardioplegic solution resulting in 25 mmol/L KCl) was infused into the aortic root at a perfusion pressure of 60 mm Hg, followed by a continuous infusion of warm, low-potassium cold blood cardioplegia (4:1 mixture of oxygenated blood with a hyperkalemic crystalloid cardioplegic solution resulting in 12 mmol/L KCl) at a flow rate of 45 mL/min (cWB-CP group).
Rapid arrest of the hearts was obtained in all cases. At no time did the hearts fibrillate during the ischemic cardioplegia period. After 60 minutes of cardioplegic arrest, the heart was excised and immediately placed in a cold Krebs' buffer solution.
Cold or Warm Cardioplegia–Reperfusion Group
In 6 pigs from both groups, the aortic cross-clamp was removed after 60 minutes of cardioplegic arrest and the heart was reperfused with normothermic blood in the bypass circuit. The heart was kept decompressed with a left ventricular vent until a stable rhythm was obtained. A mean perfusion pressure of 50 to 70 mm Hg was maintained. In the event of ventricular fibrillation, lidocaine (10 mg) was infused intravenously and the heart was defibrillated with 10 joules after the myocardial temperature increased to greater than 30°C. Pigs were weaned off of CPB shortly after the release of the aortic cross-clamp by increasing the left atrial pressure, and then decannulated. After 60 minutes of reperfusion, the heart was rapidly excised and immediately placed in a cold Krebs' buffer solution.
All animals received humane care in compliance with the Beth Israel Hospital Committee on Animal Research and with the "Guide for the Care and Use of Laboratory Animals" (NIH publication 85-23, revised 1985).
Coronary Microvessel Studies
Coronary arterial microvessels (50 to 160 µm in internal diameter) were dissected from the left anterior descending artery–dependent subepicardial region in the left ventricle using a x10 to x60 dissecting microscope (Olympus Optical, Tokyo, Japan). Microvessels were placed in an isolated microvessel chamber, cannulated with dual glass micropipettes measuring 40 to 80 µm in diameter, and secured with 10-0 nylon monofilament suture (Ethicon, Somerville, NJ). Oxygenated (95% oxygen and 5% carbon dioxide) Krebs' buffer solution warmed to 37°C was continuously circulated through the microvessel chamber and reservoir containing a total of 100 mL of the solution. The vessels were pressurized to 40 mm Hg in a no-flow state using a burette manometer filled with a Krebs' buffer solution. With an inverted microscope (x40 to x200; Olympus CK2, Olympus Optical) connected to a video camera, the vessel image was projected onto a black-and-white television monitor. An electronic dimension analyzer (Living System Instrumentation, Burlington, VT) was used to measure the internal lumen diameter. Measurements were recorded with a strip-chart recorder (Graphtec, Irvine, CA). Vessels were allowed to bathe in the microvessel chamber for at least 30 minutes before an intervention.
Microvessel Study Protocols
In all experimental groups, the relaxation responses of coronary microvessels were examined after precontraction of the microvessels with acetylcholine chloride by 20% to 70% of the baseline diameter at a distending pressure of 40 mm Hg. Once the steady-state tone was reached, the dose responses to isoproterenol (10-12 to 10-4 mol/L), forskolin (10-9 to 10-5 mol/L), 8-bromo-cyclic adenosine monophosphate (AMP) (10-10 to 10-4 mol/L), adenosine 5`-diphosphate (10-9 to 10-4 mol/L), calcium ionophore A23187 (10-9 to 10-5 mol/L), 8-bromo-cyclic guanosine monophosphate (GMP) (10-9 to 10-4 mol/L), or sodium nitroprusside (10-9 to 10-4 mol/L) were examined. The order of drug administration was random, except that A23187 was always tested last. The dose response to isoproterenol was examined only once in each vessel to avoid tachyphylaxis. All drugs were applied extraluminally. Measurements were always made 2 to 3 minutes after the drug was administered, when the response had stabilized. One to four interventions were performed on each vessel. The vessels were washed with a Krebs' buffer solution and allowed to equilibrate in a drug-free Krebs' buffer solution for 15 to 30 minutes between interventions.
Myogenic reactivity was examined after microvessels had equilibrated for at least 30 minutes at a transmural pressure of 50 mm Hg. The active pressure–diameter was then recorded as follows: the pressure was reduced to 10 mm Hg to stabilize for 10 minutes and then the pressure was increased in increments of 10 mm Hg up to 100 mm Hg. At each pressure increment, changes in internal diameter were measured after the microvessel diameter had stabilized for 2 to 3 minutes. On completion of the determination of the active pressure–diameter relationship, the pressure was returned to 50 mm Hg and then 0.1 mmol/L papaverine was applied. The passive pressure–diameter relationship was then examined according to the protocol already described. Vessel diameters were normalized to diameters at a pressure of 50 mm Hg after the application of 0.1 mmol/L papaverine.
Drugs
Isoproterenol, acetylcholine chloride, calcium ionophore A23187, adenosine 5`-diphosphate, and sodium nitroprusside were obtained from Sigma Chemical (St. Louis, MO). Forskolin, 8-bromo-cyclic AMP, and 8-bromo-cyclic GMP were obtained from RBI (Natick, MA). Forskolin, calcium ionophore A23187, 8-bromo-cyclic AMP, and 8-bromo-cyclic GMP were dissolved in dimethyl sulfoxide to make a stock solution and were stored at -20°C. Other drugs were dissolved in ultrapure distilled water. All solutions were prepared on the day of the study.
Statistical Analysis
The microvessel relaxation response to each agent was examined only once in each animal. Therefore each animal served as one sample. The data were pooled from each dose response in each experimental group and an average was calculated. The relaxation responses were expressed as the percentage of relaxation of the acetylcholine-preconstricted diameter (mean ± standard error of mean) of the microvessels. The dose-response curves of all experimental groups were compared using two-way analysis of variance with a repeated-measures design (followed by Fisher's test). Student's t test was used to compare changes in hemodynamic variables after reperfusion. Statistical significance was taken at a p value of less than 0.05.
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Results
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Vessel Characteristics
Coronary microvessels ranged from 50 to 160 µm in internal diameter, averaging 123 ± 5 µm in the control group, 107 ± 3 µm in the iCB-CP group, and 112 ± 4 µm in the iCB-CP-Rep group. These values are comparable to those in the cWB-CP and cWB-CP-Rep groups of 104 ± 3 µm and 95 ± 3 µm, respectively. The percentage of precontraction after the application of acetylcholine was 46% ± 2% in the control group, 45% ± 2% in the iCB-CP group, and 44% ± 2% in the iCB-CP-Rep group. These values are comparable to those in cWB-CP and cWB-CP-Rep groups of 42% ± 1% and 34% ± 6%, respectively. The mean concentrations of acetylcholine required to obtain these percentages of contractions were 5 x 10-7 mol/L in the control group, 4 x 10-7 mol/L in the iCB-CP group, and 6 x 10-7 mol/L in the iCB-CP-Rep group. These values are comparable to those in the cWB-CP and cWB-CP-Rep groups of 9 x 10-7 and 8 x 10-7 mol/L, respectively.
In Vitro Response to Isoproterenol
Isoproterenol (a ß-adrenoceptor agonist) induced a potent relaxation in control microvessels. The relaxation response to isoproterenol was reduced after cardioplegic arrest in both the iCB-CP and cWB-CP groups (both p < 0.01 versus control group) (Fig 1
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Fig 1. . In vitro responses of precontracted porcine coronary microvessels to isoproterenol from control hearts, hearts after 1 hour of ischemic cardioplegic arrest using intermittent cold blood (iCB-CP) or continuous warm blood (cWB-CP) cardioplegic solution, or after ischemic arrest with either cardioplegia, followed by 1 hour of reperfusion (iCB-CP-Rep or cWB-CP-Rep). Responses are expressed as the percentage of relaxation of the acetylcholine-induced vascular contraction. (**p < 0.01 versus control group.)
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In Vitro Response to Forskolin
Forskolin (an adenylate cyclase activator) also induced a potent relaxation in control microvessels. The relaxation response to forskolin in the iCB-CP group was reduced after cardioplegic arrest (p < 0.01 versus control group), whereas the response of vessels in the cWB-CP group was preserved (p < 0.01 versus iCB-CP group) after cardioplegic arrest. The response of vessels to forskolin in the cWB-CP group was greater than that of vessels in the iCB-CP group (p < 0.05) (Fig 2). After 1 hour of reperfusion the responses in both groups were similar to those in the control group.
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Fig 2. . In vitro responses of precontracted porcine coronary microvessels to forskolin from control hearts, hearts after 1 hour of ischemic cardioplegic arrest using intermittent cold blood (iCB-CP) or continuous warm blood (cWB-CP) cardioplegic solution, or after ischemic arrest with either cardioplegia, followed by 1 hour of reperfusion (iCB-CP-Rep or cWB-CP-Rep). Responses are expressed as the percentage of the relaxation of the acetylcholine-induced vascular contraction. (*p < 0.05 versus iCB-CP; **p < 0.01 versus control and cWB-CP groups.)
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In Vitro Response to 8-Bromo-Cyclic AMP
The relaxation response to 8-bromo-cyclic AMP (a membrane-permeable cyclic AMP–dependent protein kinase activator) was not significantly reduced after iCB-CP or after cWB-CP, although the response of the vessels in the iCB-CP-Rep group was significantly less than that of the cWB-CP-Rep group (p < 0.05; iCB-CP-Rep versus cWB-CP-Rep groups). The responses in the cWB-CP and cWB-CP-Rep groups were not further affected after 1 hour of reperfusion (Fig 3).
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Fig 3. . In vitro responses of precontracted porcine coronary microvessels to 8-bromo-cyclic adenosine monophosphate (AMP) from control hearts, hearts after 1 hour of ischemic cardioplegic arrest using intermittent cold blood (iCB-CP) or continuous warm blood (cWB-CP) cardioplegic solution, or after ischemic arrest with either cardioplegia, followed by 1 hour of reperfusion (iCB-CP-Rep or cWB-CP-Rep). Responses are expressed as the percentage of the relaxation of the acetylcholine-induced vascular contraction. (*p < 0.05 versus iCB-CP-Rep group.)
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Endothelium-Dependent Relaxation
Relaxation responses to the receptor-mediated vasodilator adenosine 5`-diphosphate (Fig 4) and the non–receptor-mediated endothelium-dependent agent A23187 (Fig 5) were not statistically different between groups.
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Fig 4. . In vitro responses of precontracted porcine coronary microvessels to adenosine 5`-diphosphate (ADP) from control hearts, hearts after 1 hour of ischemic cardioplegic arrest using intermittent cold blood (iCB-CP) or continuous warm blood (cWB-CP) cardioplegic solution, or after ischemic arrest with either cardioplegia, followed by 1 hour of reperfusion (iCB-CP-Rep or cWB-CP-Rep). Responses are expressed as the percentage of the relaxation of the acetylcholine-induced vascular contraction
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Fig 5. . In vitro responses of precontracted porcine coronary microvessels to the calcium ionophore A23187 from control hearts, hearts after 1 hour of ischemic cardioplegic arrest using intermittent cold blood (iCB-CP) or continuous warm blood (cWB-CP) cardioplegic solution, or after ischemic arrest with either cardioplegia, followed by 1 hour of reperfusion (iCB-CP-Rep or cWB-CP-Rep). Responses are expressed as the percentage of the relaxation of the acetylcholine-induced vascular contraction
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In Vitro Responses to 8-Bromo-Cyclic GMP
Relaxation responses to 8-bromo-cyclic GMP (a membrane-permeable cyclic GMP–dependent protein kinase activator) were significantly reduced in the iCB-CP group (p < 0.05 versus control and cWB-CP groups) and remained reduced after reperfusion (p < 0.05 versus cWB-CP-Rep group) (Fig 6). The responses in the cWB-CP and cWB-CP-Rep groups were preserved.
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Fig 6. . In vitro responses of precontracted porcine coronary microvessels to 8-bromo-cyclic guanosine monophosphate (GMP) from control hearts, hearts after 1 hour of ischemic cardioplegic arrest using intermittent cold blood (iCB-CP) or continuous warm blood (cWB-CP) cardioplegic solution, or after ischemic arrest with either cardioplegia, followed by 1 hour of reperfusion (iCB-CP-Rep or cWB-CP-Rep). Responses are expressed as the percentage of the relaxation of the acetylcholine-induced vascular contraction. (*p < 0.05 versus control and cWB-CP groups; #p < 0.05 versus cWB-CP-Rep group.)
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Endothelium-Independent Cyclic GMP–Mediated Relaxation
Relaxation responses to sodium nitroprusside were similar in all groups, indicating no alteration had occurred in the ability of the vascular smooth muscle to relax through the activation of guanylate cyclase (Fig 7).
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Fig 7. . In vitro responses of precontracted porcine coronary microvessels to sodium nitroprusside (SNP) from control hearts, hearts after 1 hour of ischemic cardioplegic arrest using intermittent cold blood (iCB-CP) or continuous warm blood (cWB-CP) cardioplegic solution, or after ischemic arrest with either cardioplegia, followed by 1 hour of reperfusion (iCB-CP-Rep or cWB-CP-Rep). Responses are expressed as the percentage of the relaxation of the acetylcholine-induced vascular contraction
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K+ATP-Channel–Mediated Relaxation
Relaxation responses to pinacidil (a K+ATP-channel opener) were similar and unchanged in all experimental groups, suggesting that channel activity was preserved (Fig 8).
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Fig 8. . In vitro responses of precontracted porcine coronary microvessels to pinacidil from control hearts, hearts after 1 hour of ischemic cardioplegic arrest using intermittent cold blood (iCB-CP) or continuous warm blood (cWB-CP) cardioplegic solution, or after ischemic arrest with either cardioplegia, followed by 1 hour of reperfusion (iCB-CP-Rep or cWB-CP-Rep). Responses are expressed as the percentage of the relaxation of the acetylcholine-induced vascular contraction.
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Myogenic Activity and Passive Vessel Characteristics
In control vessels, myogenic contractions were seen when the stepwise increase in the pressure reached more than 40 mm Hg (Fig 9). However, both iCB-CP and cWB-CP caused a decrease in the myogenic contraction of vessels in response to elevations in the pressure. No additional changes were observed after reperfusion (p < 0.01; all groups versus control group). The decreased myogenic tone was comparable in all groups (Fig 9A).
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Fig 9. . Plot showing (A) active and (B) passive pressure–diameter relationships in porcine coronary microvessels from control hearts, hearts after 1 hour of ischemic cardioplegic arrest using intermittent cold blood (iCB-CP) or continuous warm blood (cWB-CP) cardioplegic solution, or after ischemic arrest with either cardioplegia, followed by 1 hour of reperfusion (iCB-CP-Rep or cWB-CP-Rep). Passive pressure–diameter relationships were obtained after pretreatment of the vessels with 0.1 mmol/L papaverine (Pap). Vessel diameters were normalized to diameters at a pressure of 50 mm Hg after the application of 0.1 mmol/L papaverine. (**p < 0.01 control versus iCB-CP, iCB-CP-Rep, cWB-CP, and cWB-CP-Rep groups.)
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After the vessels were pretreated with papaverine, the vessels behaved passively in response to the stepwise elevation of the pressure. In control vessels, passive pressure–diameter relationships obtained after the application of 0.1 mmol/L papaverine were significantly different from the active pressure–diameter relationships. However, there were no statistical differences between the experimental groups (Fig 9B
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Postischemic Cardiac Function and Myocardial Blood Flow
The hematocrit decreased from a mean of 30% ± 3% at the time of initiation of CPB to 23% ± 2% and 23% ± 2% in the cWB-CP-Rep and iCB-CP-Rep groups, respectively, after the period of cardioplegia and to 21% ± 2% and 20% ± 2%, respectively, after 1 hour of reperfusion. All indices of cardiac function, including left ventricular systolic pressure and the maximum rate of increase of left ventricular pressure, were similar in both groups. The left ventricular systolic pressure decreased from 102 ± 3 mm Hg to 86 ± 4 mm Hg and 78 ± 8 mm Hg at 30 and 60 minutes, respectively, in the iCB-CP-Rep group and from 100 ± 3 mm Hg to 85 ± 10 mm Hg and 85 ± 7 mm Hg at 30 and 60 minutes, respectively, in the cWB-CP-Rep group. The maximum rate of increase of left ventricular pressure decreased slightly from 1,741 ± 209 mm Hg/s before CPB to 1,746 ± 256 mm Hg/s and 1,554 ± 211 mm Hg/s at 30 and 60 minutes, respectively, in the cWB-CP-Rep group and to 1,674 ± 162 mm Hg/s and 1,410 ± 207 mm Hg/s at 30 and 60 minutes, respectively, in the iCB-CP-Rep group. Heart rate (Fig 10A) and the mean systemic arterial pressure (Fig 10B) were similar in both groups, whereas the postcardioplegia blood flow of the left anterior descending coronary artery (Fig 10C) was significantly greater in the cWB-CP-Rep group than that in the iCB-CP-Rep group (p < 0.05).
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Fig 10. . Plot showing (A) heart rate (HR), (B) mean systemic arterial pressure (MAP) and (C) blood flow of left anterior descending artery (LAD flow) at baseline and at 10, 20, 30, and 60 minutes after reperfusion using intermittent cold blood or continuous warm blood cardioplegic solution (iCB-CP-Rep or cWB-CP-Rep). (*p < 0.05 versus iCB-CP-Rep group.)
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Comment
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Interpretation of Results
The major findings of this study are (1) that cWB-CP is marginally better than iCB-CP in preserving adenylate cyclase– and cyclic AMP–mediated responses of the coronary microcirculation, but the receptor-mediated responses to isoproterenol are similar for the two methods of protection; (2) that both receptor-mediated and non–receptor-mediated endothelium-dependent relaxation responses are preserved well after iCB-CP and cWB-CP; and (3) that active myogenic responses are reduced similarly after cWB-CP or iCB-CP, whereas passive microvascular characteristics are unchanged.
Decreased ß-adrenergic activity has been observed in cardiac myocytes, lymphocytes [10, 11], and, more recently, the coronary circulation [1] after CPB and cardioplegia. Both iCB-CP and cWB-CP caused ß-adrenergic–mediated relaxation to decrease. After 1 hour of reperfusion the responses were restored completely in the cWB-CP group, whereas those in the iCB-CP group remained slightly although not significantly reduced. The adenylate cyclase–mediated relaxation to forskolin and the response to 8-bromo-cyclic AMP were impaired more after cold than after warm blood cardioplegia, suggesting greater impairment in function after cold cardioplegia. The cause of the differential effect of warm blood cardioplegia as opposed to cold blood cardioplegia in preserving cyclic AMP–mediated relaxation may be related to an effect of hypothermia [11] on prolonged intracellular calcium ion influx or to other changes occurring in the vascular smooth muscle contractile mechanism during hyperkalemic cardioplegia.
The receptor-mediated endothelium-dependent relaxation in response to adenosine 5`-diphosphate, the non–receptor-mediated endothelium-dependent relaxation in response to A23187; and the endothelium-independent relaxation in response to sodium nitroprusside were not affected by either warm or cold blood cardioplegia. This is not surprising, because cold blood cardioplegia has previously been reported to preserve endothelium-dependent relaxation [3]. However, the relaxation in response to an analog of cyclic GMP was significantly reduced after cold blood cardioplegia, suggesting a compensatory increase in the activity of guanylate cyclase occurs in association with a decreased activity of the cyclic GMP–dependent protein kinase. However, these findings will need to be verified by direct examination of enzyme activities or through the use of other pharmacologic probes. The response to 8-bromo-cyclic GMP was preserved after warm blood cardioplegia. Cyclic AMP–mediated relaxation showed the same trend, although the differences were minimal. There are several possible explanations for these findings: (1) the concentration of intracellular ATP and GTP (substrates of cyclic AMP and cyclic GTP, respectively) may be impaired more after cold than after warm blood cardioplegia [12]; (2) the greater activation of the respective phosphodiesterases after cold blood cardioplegia could lead to a more rapid degradation of cyclic GMP and cyclic AMP; and (3) the activity of the cyclic GMP– and cyclic AMP–dependent protein kinases may be decreased more after cold than after warm blood cardioplegia.
Cardiopulmonary bypass, and especially normothermic CPB, is associated with the activation of complement [15, 16], the generation of oxygen-derived free radicals [13, 14], increased cytokine (tumor necrosis factor, interleukin-1, interleukin-6) release [17–19], and increased expression of adhesion molecules (CD11b, CD11c, ICAM-1) [17]. Therefore vascular damage could be exaggerated in the setting of warm as opposed to hypothermic heart operations. However, hypothermia tends to only delay some of these detrimental effects of CPB [17]. Furthermore, in our iCB-CP group the systemic temperature during CPB was maintained at 37°C. Thus the systemic release of cytokines probably is not the cause of the slight differential vascular effects observed in this study. However, the myocardial release of cytokines may be increased more after cold than after warm blood cardioplegia, accounting for the slight advantage of warm blood cardioplegia in the preservation of vascular function.
Results of studies of the differential effects of cold and warm blood cardioplegia on myocardial protection are conflicting [9, 20–22]. In this study, no difference was noted between the two groups in any of the indices of cardiac function. However, baseline coronary blood flow was increased more after warm than after cold blood cardioplegia. This may be due to greater activation of K+ATP channels in vivo after warm blood cardioplegia, because K+ATP channels mediate postcardioplegia hyperemia [23].
Clinical Implications
The impaired coronary circulation that occurs after reperfusion may be attributed to the inability of endothelium to maintain the basal and stimulated release of endothelium-derived nitric oxide and to the changes in vascular smooth muscle reactivity mediated by the ß-adrenoceptor, protein kinase C, and other mechanisms. Although the basal release of nitric oxide was not assessed in either group in this study, it is likely that it is increased as a result of the increased expression of the inducible form of nitric oxide synthase. Myogenic reactivity did not recover even after warm blood cardioplegia. This is consistent with the increased expression of the inducible form of nitric oxide synthase in the myocardium and vascular tissue that occurs after cardiac extracorporeal circulation and cardioplegia.
It is well known that blood cardioplegia may have potential advantages. For example, it has rheologic beneficial effects in the microvasculature, it promotes the endogenous oxygen-derived free radical scavenging of superoxide dismutase, and it enhances the natural buffering capacity of blood resulting from histidine. Furthermore, the drawbacks of hypothermia are avoided with warm cardioplegia and the benefits of preserving postischemic cardiac function and the vascular integrity of coronary circulation are ensured [20, 21]. This study revealed that warm blood cardioplegia can preserve ß-adrenoceptor– and cyclic AMP–mediated relaxations in the coronary microcirculation marginally better than cold blood cardioplegia can. In addition, the slight beneficial effect of warm blood cardioplegia is no longer evident after a short period of reperfusion. Endothelium-dependent relaxation and K+ATP-channel–mediated relaxation were preserved in both experimental groups, and no difference was noted in myocardial functional recovery. In the present study, a nonischemic heart model was used. Findings from a study conducted by Horsley and colleagues [12] suggested that cWB-CP preserves myocardial function better than iCB-CP does in the acutely ischemic setting. Thus the advantageous effects of warm blood cardioplegia on vascular function may become apparent in the acutely ischemic setting.
Conclusion
Continuous warm blood cardioplegia preserves ß-adrenoceptor–mediated relaxation of the coronary microcirculation better than iCB-CP does, but this potential benefit disappeared after 1 hour of reperfusion. Endothelium-dependent relaxation responses were equally well preserved by cold and warm blood cardioplegia. Surprisingly, cyclic GMP– and cyclic AMP–dependent protein kinase activities were reduced more after iCB-CP than after cWB-CP. Whereas warm blood cardioplegia had a slight beneficial effect in the preservation of vascular reactivity as opposed to the effect of cold blood cardioplegia, it did not have any beneficial effects on myogenic reactivity or cardiac functional recovery.
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Acknowledgments
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Supported by grant HL 46716 from the National Heart, Lung and Blood Institute.
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Footnotes
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Address reprint requests to Dr Sellke, Division of Cardiothoracic Surgery, Beth Israel–Deaconess Medical Center, East Campus, Dana 905, 330 Brookline Ave, Boston, MA 02215.
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References
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Abstract
Introduction
