Complement C5a (desArg) generation in serum exposed to damaged aortic endothelium.
Complement activation by injured endothelial cells was investigated using ex vivo rabbit thoracic aortas as a source of endothelium and a neutrophil aggregation (NA) bioassay to detect the complement cleavage product C5a (desArg). Endothelium, oxygen-starved by incubating aortas 30 min with Tyrode's buffer, activated complement as demonstrated by a 57% increase in the NA response induced by serum from buffer-treated aortas as compared to serum from untreated control aortas. Incubation of MgEGTA serum in injured aortas resulted in a 27% (P less than 0.025) weaker NA response than normal serum, indicating participation by both classical and alternative pathways of complement activation. Serum from aortas incubated 30 min with 100 micrograms/ml cholestane-3 beta, 5 alpha, 6 beta-triol, a cytotoxic cholesterol oxidation derivative, induced a NA response comparable to that from serum from aortas treated 30 min with Tyrode's buffer. Heat inactivation of serum prior to aortic incubation abolished NA activity and serum incubated in deendothelialized aortas lacked NA activity. Fractionation of serum samples from these experiments on Sephadex G-100 revealed a single peak of NA activity corresponding to the molecular weight of C5a (desArg). Endothelial cell injury was demonstrated by the inability to exclude Trypan blue dye and by scanning electron microscopy. These data demonstrate that damaged arterial endothelium can effectively activate the complement system, resulting in the production of an anaphylatoxic inflammatory mediator.
Seifert
P S
PS
Department of Pathology, Albany Medical College, New York 12201.
Catalfamo
J L
JL
Dodds
W J
WJ
D000818:Animals; D001013:Aorta, Thoracic / Q000503:physiopathology*; D002449:Cell Aggregation; D003167:Complement Activation; D003182:Complement C5 / Q000031:analogs & derivatives* / Q000032:analysis; D015937:Complement C5a, des-Arginine; D004730:Endothelium, Vascular / Q000503:physiopathology* / Q000648:ultrastructure; D007511:Ischemia / Q000097:blood / Q000503:physiopathology*; D008855:Microscopy, Electron, Scanning; D009504:Neutrophils / Q000473:pathology; D011817:Rabbits
D003182:Complement C5; D015937:Complement C5a, des-Arginine