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Anesth Analg 1999;89:42
© 1999 International Anesthesia Research Society

CARDIOVASCULAR ANESTHESIA

Attenuation of Endothelium-Dependent Dilation of Pig Pulmonary Arterioles After Cardiopulmonary Bypass Is Prevented by Monoclonal Antibody to Complement C5a

Kyung W. Park, MD*, Motohisa Tofukuji, MD, PhD{dagger}, Caroline Metais, MD{dagger}, Mark E. Comunale, MD*, Hai B. Dai, MD{dagger}, Michael Simons, MD{ddagger}, Gregory L. Stahl, PhD§, Azin Agah, PhD§, and Frank W. Sellke, MD{dagger}

Departments of *Anesthesia and Critical Care, {dagger}Surgery, and {ddagger}Medicine, Beth Israel Deaconess Medical Center and Center for Experimental Therapeutics and Reperfusion Injury; and §Department of Anesthesiology, Brigham & Women's Hospital, Harvard Medical School, Boston, Massachusetts

Address correspondence and reprint requests to Kyung W. Park, MD, Department of Anesthesia & Critical Care, 330 Brookline Ave., Boston, MA 02215. Address e-mail to kpark{at}caregroup.harvard.edu


   Abstract
Top
 Abstract
Introduction
 Methods
 Results
 Discussion
 References
 
We examined whether pulmonary endothelial dysfunction associated with cardiopulmonary bypass (CPB) may be mediated by complement C5a in pigs. Pigs were placed on normothermic CPB for 1 h with or without a previous administration of 1.6 mg/kg anti-C5a monoclonal antibody (MAb), then reperfused for 2 h. Pulmonary tissue myeloperoxidase activity was measured. Expression of nitric oxide synthase (NOS) was measured by reverse transcriptase polymerase chain reaction and Western blotting. Pulmonary arterioles approximately 100 µm in diameter were preconstricted with the thromboxane analog U46619 1 µM, and relaxation responses to adenosine diphosphate 10-9–10-4 M, substance P 10-12–10-6 M, and sodium nitroprusside 10-9–10-4 M were examined in vitro by videomicroscopy. Relaxation to the endothelium-dependent dilators adenosine diphosphate and substance P was attenuated after CPB; this attenuation was prevented by the previous administration of MAb. Relaxation to sodium nitroprusside was not affected by CPB. Neutrophil sequestration, as measured by MPO activity, increased after CPB, either with or without MAb. Transcription of NOS was unchanged by CPB, but translation of constitutive NOS was decreased after CPB, and this decrease was prevented by a previous administration of MAb. We conclude that pig pulmonary endothelial dysfunction associated with CPB may be mediated by C5a. The mechanism may involve changes in NOS translation.

Implications: In pigs, pulmonary endothelial dysfunction may occur after cardiopulmonary bypass due to product(s) of complement activation.


   Introduction
Top
 Abstract
 Introduction
Methods
 Results
 Discussion
 References
 
Cardiopulmonary bypass (CPB) is associated with endothelial dysfunction of ovine pulmonary microvessels (1). Several factors may contribute to pulmonary endothelial dysfunction after CPB. First, CPB results in nearly complete diversion of pulmonary blood flow, except for bronchial circulation, and there may be a state of reduced pulmonary perfusion. Ischemia and reperfusion may then lead to endothelial dysfunction. However, microvessels from systemic vascular beds, such as cerebral arterioles, which are not bypassed during CPB, also display endothelial dysfunction (2). Thus, diversion of pulmonary blood flow is unlikely to be the sole reason for pulmonary endothelial dysfunction.

Second, blood contact with the foreign surface of the CPB circuit leads to a systemic inflammatory response (3). The most prominent early response is significant complement activation, with a rapid increase of the anaphylatoxins C5a and C3a in the blood exiting the extracorporeal circuit (4–6). C5a is an important mediator of immediate and delayed proinflammatory endothelial activation, leukosequestration, and monocyte, macrophage, and platelet activation and release of cytokines (3). C5a-initiated inflammation may therefore be responsible for CPB-induced pulmonary endothelial dysfunction. In addition, the C5b-9 membrane attack complex (MAC) may contribute to endothelial dysfunction, as suggested with ovine pulmonary vessels after CPB (7) and demonstrated with coronary arteries after ischemia-reperfusion (8).

In this study, we examined whether attenuation of endothelium-dependent dilation of pulmonary arterioles after CPB is also observed in pigs and whether we could prevent or ameliorate the attenuation by blocking the action of C5a with a monoclonal antibody (MAb). We further examined whether any observed endothelial dysfunction is associated with changes in expression of either the constitutive or inducible nitric oxide synthase (cNOS or iNOS, respectively).


   Methods
Top
 Abstract
 Introduction
 Methods
Results
 Discussion
 References
 
In accordance with institutional animal care committee standards, Yorkshire pigs of either sex, weighing 20–25 kg, were premedicated with ketamine 10 mg/kg IM, and IV access was established in an ear vein. Each animal was anesthetized with {alpha}-chloralose and urethane (60 and 300 mg/kg IV initially, then 15 and 60 mg/kg every 60 min as needed, respectively). The trachea was intubated, and the pig was ventilated with a Harvard ventilator (Harvard Apparatus, Cambridge, MA). The right femoral artery was dissected and cannulated with a fluid-filled catheter for continuous blood pressure monitoring and blood sampling. The right femoral vein was cannulated for venous access. After midline sternotomy, pigs were heparinized (500 U/kg IV initially and 300 U/kg every 90 min) and cannulated via the distal ascending aorta and the right atrium. Activated clotting time (ACT) was measured at regular intervals and was maintained >300 s. Total CPB was instituted using a bubble oxygenator (Bentley Bio-2; Baxter Healthcare Corp., Irvine, CA) and a standard roller pump (Cardiovascular Instrument Corp., Wakefield, MA). An arterial filter (Bentley Bio-1025; Baxter Healthcare Corp.) was inserted into the circuit distal to the roller pump. Arterial blood gas tensions were obtained before CPB and at approximately 20-min intervals thereafter. Arterial blood gas tensions were maintained at PO2 >100 mm Hg, pH 7.40 ± 0.05, and PCO2 40 ± 5 mm Hg by adjusting ventilatory rate, tidal volume, and fraction of inspired oxygen. The systemic temperature was maintained at 37°C. Blood flow was maintained at 80–100 mL · kg-1 · min-1 to maintain a mean perfusion pressure of 50–80 mm Hg. After 1 h of CPB, the animal was separated from CPB by increasing cardiac filling. No pharmacological support was required for separation from CPB. Mean pulmonary artery pressures and total pulmonary flows were recorded before and after CPB.

For the MAb group, 1.6 mg/kg murine antiporcine C5a MAb was administered iv 20 min before CPB. To produce the MAb, female Balb-C mice were immunized monthly with 25 µg of purified porcine C5a (pC5a) for 3 mo. The mice were killed 5 days after an iv injection of 1 µg of pC5a. Splenocytes were fused with the P3/NSI/1-Ag4-1 mouse myeloma cell line by using standard techniques. Hybridomas were screened by using an antibody capture enzyme-linked immunosorbent assay using 96-well plates coated with pC5a. Limited dilution techniques were used to isolate a single positive clone. GS17F1C4 was isolated as a murine IgG 1{kappa} isotype. Antibodies were produced in tissue culture and as ascites. Purification of the antibody was achieved by protein G column chromatography using glycine (100 mmol/L, pH 3.0) as the elutant. The specificity of our antibodies has been previously demonstrated (9).

For the vehicle control (VC) group, animals were placed on and separated from CPB in a fashion similar to the MAb group, but saline was injected without MAb before CPB.

After separation from CPB, protamine was administered to reverse heparinization and to normalize ACT. After 2 h of reperfusion, the lungs were removed. A portion of lung tissue was rapidly excised and immediately placed in cold (4°C) modified Krebs buffer solution (NaCl 120 mM, KCl 5.9 mM, dextrose 11.1 mM, NaHCO3 25 mM, NaH2PO4 1.2 mM, MgSO4 1.2 mM, CaCl2 2.5 mM).

Control animals were not placed on CPB, and their lungs were similarly harvested.

Lung tissue samples were obtained from the MAb, VC, and control animals, and each tissue sample was frozen in liquid nitrogen at -80°C. To measure neutrophil sequestration, myeloperoxidase (MPO) activity was measured as previously described (10). Briefly, ice-cold minced sections of the tissue were homogenized (10% wt/vol) with a Polytron homogenizer (Brinkmann Instruments, Westbury, NY) in 50 mM phosphate buffer (pH 6.0) containing 0.5% hexadecyltrimethylammonium bromide for 15 s (x2). The homogenates were then sonicated for 15 s, and the supernatant containing the MPO enzyme was separated from the cellular debris by centrifugation at 20,000 g for 15 min at 2°C. The appearance of a colored product from the MPO-dependent reaction of o-dianisidine and hydrogen peroxide (0.0005%) was spectrophotometrically detected using 100 µL of supernatant and 1.9 mL of hydrogen peroxide and o-dianisidine in 50 mM phosphate buffer (pH 6.0). Supernatant MPO activity was kinetically quantified on a temperature-controlled (25°C) spectrophotometer (Lambda 3B; Perkin-Elmer, Ridgefield, CT) with a chart recorder. The absorbance at 460 nm was recorded for several minutes, and the change in absorbance over 1 min was measured during the linear portion (0.5–1.5 min) of the tracing. One unit of MPO activity was defined as that which degraded 1 mol of hydrogen peroxide per minute at 25°C. We previously (10) demonstrated that 1 U of MPO activity correlated with 2.9 x 106 neutrophils.

Lung tissue samples were frozen in liquid nitrogen and stored at -80°C until study. cNOS and iNOS fragments were amplified by using reverse transcription polymerase chain reaction (RT-PCR). Primers were designed based on the published cNOS (11) and iNOS (12) sequences. The primers of the sense 5'-AGACCCCTGGAAAGGGAG-3' corresponding to bases 1443–1460, and of the antisense 5'-TGTGTTACTGGATTCCTTCC-3' corresponding to bases 1901–1920 were used to amplify a 486-basepair fragment of cNOS (11). For iNOS, the primer of the sense 5'-GCCTCGCTCTGGAAAGA-3' corresponding to bases 1425–1441 and of the antisense 5'-TCCATGCAGACAACCTT-3' corresponding to bases 1908–1924 were used to amplify a 500-base pair fragment of iNOS (12).

An equal amount of total RNA was used for RT-PCR from control and experimental groups. For quantification, glyceraldehyde-3-phosphate dehydrogenase was amplified from the same amount of RNA to correct for variation of different samples. The PCR products were loaded in 1.5% agarose gel, then scanned and measured on Image-Quant software (Molecular Dynamics, Sunnyvale, CA).

Total protein from lung tissues was obtained by homogenizing in a lysis buffer containing 1% NP-40, 0.5% sodium deoxycholate, and 0.1% sodium dodecyl sulfate (SDS), then centrifuging at 12,000 g for 10 min at 4°C. Protein concentration of the supernatant was measured by spectrophotometry at 595 nm of an aliquot developed for 10 min in Protein Assay Dye Reagent (Bio-Rad, Hercules, CA). Total protein (50 µg/lane) was fractionated on 10% SDS-polyacrilamide gel transferred to a polyvinylidene difluoride membrane (Immobilon-P; Millipore, Bedford, MA). The membrane was incubated with 5% nonfat dry milk powder and 0.05% Tween20 in phosphate-buffered saline (PBS) for 12 h at 4°C to block nonspecific absorption and was then immunoblotted with the monoclonal mouse anti-endothelial cNOS or anti-iNOS antibody (Transduction Laboratories, Lexington, KY) 1:2000 (vol/vol) dilution for 2 h. After washing with PBS, the membrane was incubated for 1 h in 5% milk powder PBS containing 1:3000 diluted goat anti-mouse immunoglobin G conjugated to horseradish peroxidase (Vector Laboratories, Burlingame, CA). Peroxidase activity was visualized using an enhanced chemiluminescence substrate system (Amersham, Arlington Heights, IL). Densitometry of digitized images of immunoprobed membranes was performed using ImageQuant software.

Pulmonary resistance arteries measuring approximately 100 µm in internal diameter were dissected free of the surrounding tissues. Each vessel was placed in a vessel chamber, cannulated with dual micropipettes (50–75 µm in diameter), and secured with 10-0 sutures. The vessel was continuously bathed with modified Krebs buffer, gassed with a 95% O2/5% CO2 mixture, maintained at 37°C and pH of 7.4, and studied in a no-flow state. The pressure in the micropipettes was maintained at 20 mm Hg to provide vessel distention. The vessel was visualized with an inverted phase-contrast microscope (IMT-2; Olympus, Tokyo, Japan) connected to a video camera. The vessel image was projected onto a television screen (Panasonic, Osaka, Japan). Changes in the vessel internal lumen diameter were monitored with an optical density video detection system (Living Systems Instrumentation, Burlington, VT), as previously described (13).

After equilibration of each vessel for at least 30 min in the vessel chamber, a baseline internal diameter was measured (Dbaseline). Each vessel was then preconstricted with the thromboxane analog U46619 1 µM, and the constricted diameter was measured (Dconst). The vessel was then exposed to increasing concentrations of the endothelium-dependent dilator adenosine diphosphate (ADP) 10-9–10-4 M, another endothelium-dependent dilator substance P (SP) 10-12–10-6 M, or the endothelium-independent dilator sodium nitroprusside (NP) 10-9–10-4 M.

In a preliminary study, the fact that ADP and SP were indeed endothelium-dependent dilators and NP an endothelium-independent dilator in pig pulmonary vessels was verified using 18 vessels from control animals. ADP 10-4 M produced 59% ± 8% relaxation of U46619-preconstricted vessels, but produced only 17% ± 4% relaxation (P < 0.001) in the presence of the NOS inhibitor NG-nitro-L-arginine (L-NNA) 10-5 M. Similarly, SP 10-6 M produced 61% ± 9% relaxation in the absence of L-NNA but 24% ± 6% relaxation in the presence of L-NNA (P < 0.001). However, NP 10-4 M produced 82% ± 5% relaxation without L-NNA and 81% ± 5% relaxation with L-NNA (P = not significant).

At each concentration of the dilator, the diameter was measured (Drelax), and percent relaxation from U46619-preconstricted state was calculated as follows: Go


At the end of each experiment, the vessel chamber was flushed with fresh Krebs buffer, and the vessel was reequilibrated at 37°C. KCl was then added to achieve a concentration of 100 mM in the buffer and vessel chamber. ADP was then added to achieve a concentration of 10-5 M to test for the integrity of endothelium. Only those vessels that constricted by at least 15% to KCl and dilated significantly to ADP were considered still viable and included for data analysis. This excluded any vessel that constricted to KCl less than the average by more than approximately 1 SD.

No animal contributed more than one vessel to any one experimental group. Therefore, n for each experimental group is equal to the number of animals, as well as the number of vessels. Vasomotor responses to ADP, SP, or NP among the different experimental groups were compared by using two-way analysis of variance (ANOVA) with a repeated-measures factor, with post hoc Neumann-Keuls test for between-group comparisons. When the initial ANOVA yielded a significant P value, a stratified z test was performed to identify the concentrations of the vasomotor drugs that resulted in different responses. MPO activity and NOS expression among the different groups were compared by using two-tailed t-tests. All statistics were calculated using True Epistat software (Epistat Services, Richardson, TX). All data are presented as mean ± SD.


   Results
Top
 Abstract
 Introduction
 Methods
 Results
Discussion
 References
 
Vasodilation induced by the endothelium-dependent dilator ADP was attenuated after CPB (P < 0.03), but this effect was prevented by the administration of anti-C5a MAb before CPB (P < 0.03) (Fig. 1a), (control: n = 7, vessel size 102 ± 15 µm; VC group: n = 6, vessel size 109 ± 12 µm; MAb group: n = 7, size 104 ± 6 µm). Similarly, dilation to the endothelium-dependent dilator SP was attenuated after CPB (P < 0.001), but this effect was prevented by anti-C5a MAb (P < 0.001) (Fig. 1b) (control: n = 7, size 101 ± 10 µm; VC group: n = 7, size 107 ± 12 µm; MAb group: n = 7, size 110 ± 9 µm).



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  		Figure 1. a, Percent relaxation of U46619-preconstricted pig lung microvessels versus log concentration of the endothelium-dependent dilator adenosine diphosphate (ADP). Vasodilation to ADP was attenuated after cardiopulmonary bypass (CPB) (P < 0.03), and this effect was prevented by the previous administration of anti-C5a monoclonal antibody (MAb). b, Percent relaxation of U46619-preconstricted pig lung vessels versus log concentration of another endothelium-dependent dilator, substance P. Vasodilation to substance P was attenuated after CPB (P < 0.001), and this effect was prevented by the previous administration of MAb. *P < 0.05 versus response of the control vessels.

 
However, vasodilation to the endothelium-independent dilator NP was not affected by CPB or the anti-C5a MAb (P = 0.21) (Fig. 2) (control: n = 7, size 100 ± 17 µm; VC group: n = 6, size 111 ± 11 µm; MAb group: n = 7, size 102 ± 6 µm).



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  		Figure 2. Percent relaxation of U46619-preconstricted pig lung microvessels versus log concentration of the endothelium-independent dilator sodium nitroprusside. Vasodilation to sodium nitroprusside was not affected by cardiopulmonary bypass (CPB) or the previous administration of anti-C5a monoclonal antibody (MAb) (P = 0.21).

 
Mean pulmonary artery pressures tended to be increased and total pulmonary flow was decreased after CPB, without any significant changes in left atrial pressures (Table 1), which indicates that pulmonary vascular resistance increased significantly after CPB. Despite the beneficial effect of the MAb on responses to endothelium-dependent dilators, it failed to attenuate the increase in pulmonary vascular resistance.


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  		Table 1. Pulmonary Hemodynamics
 
MPO activity was increased on CPB (27 ± 16 U/g of tissue), compared with control (4 ± 3 U/g of tissue) (P < 0.01). After CPB with MAb, the increase in MPO activity tended to be less (16 ± 12 U/g of tissue) than that without MAb, but the difference did not reach statistical significance (P = 0.18).

Transcription of neither cNOS nor iNOS in the lung tissues was changed on CPB with or without MAb (Fig. 3, Table 2). However, translation of cNOS was decreased on CPB (control 100% versus CPB 61% ± 24%, n = 3; P < 0.05), and this decrease was prevented by previous administration of MAb (MAb 125% ± 84%, n = 3; control versus MAb P = 0.63) (Fig. 4a). Translation of iNOS was not affected by CPB (control 100% versus CPB 100% ± 1% versus MAb 95% ± 1%, n = 3 each) (Fig. 4b).



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  		Figure 3. Representative reverse transcription polymerase chain reaction products of constitutive nitric oxide synthase (cNOS) and inducible nitric oxide synthase (iNOS) in porcine lung before (control) and after cardiopulmonary bypass in saline and C5a monoclonal antibody (MAb) treatment groups. Expression of neither cNOS nor iNOS was changed on cardiopulmonary bypass with or without MAb. bp = base pair, GAPDH = glyceraldehyde-3-phosphate dehydrogenase.

 

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  		Table 2. Reverse Transcription Polymerase Chain Reaction Products
 


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  		Figure 4. Representative Western blotting products of (a) constitutive nitric oxide synthase (cNOS) and (b) inducible nitric oxide synthase (iNOS) in porcine lung before (control) and after cardiopulmonary bypass (CPB) in saline and C5a monoclonal antibody (MAb) treatment groups. Translation of cNOS was decreased on CPB (P < 0.05), and this decrease was prevented by the previous administration of MAb. Translation of iNOS was unchanged by CPB.

 

   Discussion
Top
 Abstract
 Introduction
 Methods
 Results
 Discussion
References
 
The most important findings of this study are that (a) there is selective attenuation of endothelium-dependent dilation of pig pulmonary microvessels after CPB; (b) this attenuation is prevented by the administration of anti-C5a MAb before institution of CPB; and (c) the beneficial effect of anti-C5a MAb may be related to translation of cNOS.

Endothelial dysfunction has been noted after many types of acute or chronic insults to the endothelial cells, including CPB (1,2), complement activation (7,8), hypercholesterolemia (14), coronary artery disease (15), and mechanical injury, such as from surgical manipulation (16). Even after CPB, endothelial dysfunction has been noted in pulmonary (1), cerebral (2), and coronary (17) arteries. Our demonstration of endothelial dysfunction in swine pulmonary arteries after CPB corroborates a similar finding in ovine pulmonary vessels in an earlier study (1). As in previous studies (1,2,17), relaxation to the endothelium-independent dilator NP was not altered by CPB in this study, which demonstrates that vascular smooth muscle functions downstream of the guanylate cyclase remain intact after CPB. In addition, transcriptional expression of NOS does not seem to be changed by CPB, as demonstrated by our RT-PCR studies. However, our Western blotting studies show that translation of cNOS proteins may be decreased on CPB, and this is prevented by the previous administration of MAb. Whereas certain inflammatory states, such as sepsis, are associated with increased expression of iNOS (18), this was not observed in association with CPB. Although the benefit conferred by MAb may include a mechanism involving translation of cNOS, the current study does not exclude the possibility that the activity of NOS may also have been altered by CPB. Also not excluded is the possibility that post-CPB endothelial dysfunction may also be at a level upstream of NOS and related to the endothelial receptor, the associated G-protein, and/or endothelial Ca2+ entry.

Several factors may contribute to pulmonary endothelial dysfunction after CPB. First, on CPB, there is nearly complete diversion of blood flow to the lungs, except for bronchial circulation. There may be a state of reduced pulmonary perfusion. Ischemia and reperfusion may then lead to endothelial dysfunction, as has been demonstrated with coronary arteries (19). Furthermore, pulmonary endothelial dysfunction is more severe after total CPB than after partial CPB (20), which supports the notion that diversion of blood flow may be a contributory factor. However, microvessels from systemic vascular beds, such as cerebral arterioles, which are not bypassed on CPB, also display endothelial dysfunction (2). Diversion of flow cannot be the sole mechanism of endothelial dysfunction in CPB.

Second, complement activation occurs in CPB both via the alternate pathway due to contact with the foreign surface (21) and via the classical pathway due to variable degrees of ischemia and endotoxemia (22), although activation is attenuated by hypothermia, heparinization, and hemodilution (23). Protamine-heparin complex after separation from CPB can additionally activate the complement system (21). Complement activation results in the generation of two potential offenders of endothelial function—the anaphylatoxins C5a and C3a and the C5b-9 MAC. Whereas MAC can directly attack the endothelial cell membrane, C5a and, to a lesser extent, C3a work indirectly by neutrophil chemotaxis, aggregation, and release of lysosomal enzymes and cytokines (10,21). In our preparation, we did not specifically examine the role of MAC, but we demonstrated that attenuation of endothelium-dependent dilation after CPB was prevented by the previous administration of anti-C5a MAb, which points to a significant role of C5a in the development of endothelial dysfunction in CPB. Prior studies with MAC suggest that it may also have a role (8,24).

MPO is a specific marker of neutrophils, and prior studies have demonstrated concordance between tissue MPO activity and histological determination of neutrophil quantity (10,25). Our assays of pulmonary MPO activity demonstrated that anti-C5a MAb did not significantly alter neutrophil infiltration into the lung tissues during CPB. This finding does not exclude a beneficial effect of anti-C5a MAb related to neutrophil functions. First, despite attenuation of C5a-induced chemotaxis by the MAb, there may have been other neutrophil chemotactic factors, such as oxidized plasma lipids and platelet-derived factors (10), that attracted neutrophils to the lungs. Neutrophil accumulation by itself, without coincident C5a, may not cause tissue injury (26). Second, lower concentrations of C5a are needed to induce neutrophil chemotaxis than to induce neutrophil release of cytotoxic mediators, such as lysosomal enzymes and oxygen-derived free radicals (10). Thus, anti-C5a MAb could have prevented release of cytotoxic mediators, and thereby endothelial dysfunction, without completely suppressing leukosequestration.

Despite prevention of pulmonary endothelial dysfunction on CPB by C5a MAb, the antibody failed to attenuate the increase in pulmonary vascular resistance after CPB. Although endothelial dysfunction may be contributory to post-CPB pulmonary hypertension, there seem to be other factors that were not affected by the antibody. To the extent that endothelial dysfunction may be a significant factor in the development of post-CPB pulmonary hypertension, drugs that reduce complement activation and neutrophil activation may have a therapeutic implication in the prevention and/or treatment of post-CPB pulmonary hypertension. Potential therapeutic drugs would include measures to attenuate complement activation (possibly new synthetic material for bypass circuit least likely to activate complement); drugs to neutralize the products of complement activation, such as anti-C5a MAb; drugs to limit neutrophil production of cytotoxic mediators, such as the dual lipoxygenase-cyclooxygenase inhibitor BW755C (27); and neutralizers of cytotoxic mediators, such as scavengers of oxygen-derived free radicals. Further studies are needed to identify whether any of these may be of benefit.

In summary, we demonstrated that, in pig pulmonary microvessels, there is selective attenuation of endothelium-dependent dilation after CPB. This attenuation may be prevented by the previous administration of anti-C5a MAb.


   Acknowledgments
 
Supported in part by United States Public Health Service Grants HL46716, HL53793, HL52886, and HL56086; and by a grant from the Beth Israel Anesthesia Foundation.


   Footnotes
 
Presented in part at the 1998 annual meeting of Society of Cardiovascular Anesthesiologists, Seattle, WA.


   References
Top
 Abstract
 Introduction
 Methods
 Results
 Discussion
 References
 

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Accepted for publication March 9, 1999.




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Lippincott, Williams & Wilkins Anesthesia & Analgesia® is published for the International Anesthesia Research Society® by Lippincott Williams & Wilkins and Stanford University Libraries' HighWire Press®. Copyright 1999 by the International Anesthesia Research Society. Online ISSN: 1526-7598   Print ISSN: 0003-2999 HighWire Press