Complement-Mediated Loss of Endothelium-Dependent Relaxation of Porcine Coronary Arteries

Abstract

Reperfusion of the ischemic myocardium results in the loss of endothelium-dependent relaxation. We have shown recently that the alternate complement pathway is activated immediately on reperfusion of the ischemic porcine myocardium. We hypothesized that complement activation directly attenuates endothelium-dependent relaxation of porcine coronary arteries. Bradykinin (BK) or substance P concentration-dependently relaxed precontracted (U46619, 50 nmol/L) left anterior descending coronary artery (LAD) rings in vitro. Addition of zymosan to human (10%) or porcine (10%) serum for 30 minutes significantly (P < 0.05) increased the EC50 of BK-induced LAD relaxation from 4 +/- 1 to 418 +/- 159 nmol/L (n = 8) and from 9 +/- 3 to 281 +/- 132 nmol/L (n = 7), respectively. Similarly, addition of zymosan to 10% human serum (HS) for 30 minutes increased the EC50 of substance P-induced LAD relaxation from 0.4 +/- 0.1 to 30 +/- 14 nmol/L (n = 9, P < .05). Basal release of nitric oxide was reduced significantly in LAD rings exposed to zymosan-activated HS compared with HS alone. Addition of soluble CR1 (sCR1, 10 nmol/L) to zymosan-activated HS preserved BK-induced relaxation (EC50) of the LAD rings (control, 4 +/- 1 nmol/L; sCR1 + zymosan+serum, 2 +/- 1 nmol/L; n = 6). Zymosan-activated C8-depleted HS (10%) did not attenuate the EC50 of BK-induced coronary artery relaxation (3 +/- 1 to 3 +/- 1 nmol/L, n = 7, P = NS). Zymosan-activated C8-depleted HS plus C8 (6 micrograms/mL) increased the EC50 of BK-induced coronary artery relaxation from 4 +/- 1 to 423 +/- 141 nmol/L (n = 12, P < .05). We have further demonstrated that C5b-9 complexes can be found on the luminal surface of LAD endothelial cells after 5 minutes of exposure to zymosan-activated HS by using C5b-9 reactive monoclonal antibody fluorescent immunohistochemistry and confocal microscopy.(ABSTRACT TRUNCATED AT 250 WORDS)

Endothelial Nuclear Factor-κB Translocation and Vascular Cell Adhesion Molecule-1 Induction by Complement

“…[13][14][15] We thus investigated whether C5b-9 attenuates ACh-induced increases in endothelial cGMP. Intracellular concentrations of cGMP were measured by ELISA in normoxic and hypoxic (12 hours) HUVECs reoxygenated (30 minutes) in the presence of buffer or 30% HS treated with 0 or 10 g/mL h5G1.1-scFv ( Figure 3).…”

“…We conclude that (1) C5b-9 deposition, NF-B translocation, and VCAM-1 protein expression are increased in hypoxic HUVECs reoxygenated in HS; (2) reoxygenation of hypoxic HUVECs in HS, but not buffer alone, attenuates ACh-induced cGMP synthesis; and (3) Key Words: adhesion molecules Ⅲ hypoxia Ⅲ inflammation Ⅲ nitric oxide Ⅲ immunotherapy I ncreasing evidence suggests that the terminal complement complex (C5b-9) plays an integral role in the pathogenesis of atherosclerosis 1-5 and vascular injury after ischemiareperfusion, cardiopulmonary bypass, and acute myocardial infarction. 6 -11 In addition to amplifying the local inflammatory response by inducing endothelial interleukin-8 (IL-8) and monocyte chemoattractant protein-1 (MCP-1) secretion, 12 C5b-9 also influences endothelial vascular tone [13][14][15] 16 ). Yet the mechanisms by which C5b-9 influences endothelium-dependent relaxation and leukocyte adhesion molecule expression have not been fully elucidated.…”

“…6 -11 In addition to amplifying the local inflammatory response by inducing endothelial interleukin-8 (IL-8) and monocyte chemoattractant protein-1 (MCP-1) secretion, 12 C5b-9 also influences endothelial vascular tone [13][14][15] and expression of leukocyte adhesion molecules (eg, vascular cell adhesion molecule-1 [VCAM-1], 16 intercellular adhesion molecule-1 [ICAM-1], 17 and endothelial leukocyte adhesion molecule-1 [E-selectin] 16 ). Yet the mechanisms by which C5b-9 influences endothelium-dependent relaxation and leukocyte adhesion molecule expression have not been fully elucidated.…”

Isolation, Characterization, and Cloning of Porcine Complement Component C7

“…Furthermore, C5b-9 and not C5a appears to be the major mediator of renal ischemia/reperfusion injury (16). These are not unexpected observations, considering the multitude of proinflammatory actions induced by the terminal complement components (9,13,14,18,35). Thus, identification and formation of specific inhibitors of C5b-9 may have therapeutic value.…”

“…In addition, C5b-9 induces endothelial expression of IL-8 and P-selectin (8,9), augments TNF-induced ICAM-1 and E-selectin expression (9,10), and directly attenuates endothelium-dependent relaxation of vascular smooth muscle (11)(12)(13). C5b-9 also induces a loss of acetylcholine-induced increases in intracellular cGMP in human endothelial cells, leading to translocation of NF-B and up-regulation of VCAM-1 (14).…”