Inactivation of endothelin by polymorphonuclear leukocyte-derived lytic enzymes.
Cultured bovine aortic endothelial cells (BAEC) released endothelin-1 (ET-1) in the culture medium in a time-dependent fashion. Coincubation of fMLP-activated human polymorphonuclear leukocytes (PMN) with BAEC caused a fast (maximal activity was reached within 15 minutes) and cell number-dependent disappearance of ET-1 from the medium. This effect was direct to ET-1, because it was also present when PMN were incubated with the synthetic peptide in the absence of BAEC. PMN-dependent disappearance of ET-1 was associated with loss of constrictor activity on isolated rabbit aorta. PMN-released products were responsible for ET-1 degrading activity, because supernatants of activated PMN were equally effective as the intact cells. Resting PMN, in the same time frame, were uneffective. Eglin C, a potent blocker of PMN-derived elastase and cathepsin G, reversed the ET-1 inhibitory activity of fMLP-stimulated PMN and of their supernatant. Direct addition of elastase and cathepsin G to synthetic ET-1 destroyed its immunoreactivity and this effect was blocked by eglin C. High-performance liquid chromatography (HPLC) analysis supported the hypothesis that ET-1 degradation by PMN was due to enzymatic proteolysis. These data provide evidence that activated PMN are able to degrade ET-1 through the release of proteases. Because physiologic concentrations of PMN can destroy high amounts (up to 100 nmol/L) of ET-1 within a few minutes, we propose that this mechanism of ET-1 inactivation has biologic relevance.
Patrignani
P
P
University of Chieti G. D'Annunzio, School of Medicine, Italy.
Del Maschio
A
A
Bazzoni
G
G
Daffonchio
L
L
Hernandez
A
A
Modica
R
R
Montesanti
L
L
Volpi
D
D
Patrono
C
C
Dejana
E
E
D000818:Animals; D001011:Aorta / Q000187:drug effects / Q000502:physiology; D056649:Cathepsin G; D002403:Cathepsins / Q000378:metabolism; D002417:Cattle; D002478:Cells, Cultured; D003470:Culture Media; D010450:Endopeptidases / Q000378:metabolism*; D016232:Endothelins / Q000378:metabolism* / Q000494:pharmacology; D004730:Endothelium, Vascular / Q000502:physiology*; D006801:Humans; D066298:In Vitro Techniques; D007700:Kinetics; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000502:physiology; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D009504:Neutrophils / Q000187:drug effects / Q000201:enzymology*; D010196:Pancreatic Elastase / Q000378:metabolism; D011480:Protease Inhibitors / Q000494:pharmacology; D011817:Rabbits; D012697:Serine Endopeptidases; D014661:Vasoconstriction / Q000187:drug effects
D003470:Culture Media; D016232:Endothelins; D011480:Protease Inhibitors; D009240:N-Formylmethionine Leucyl-Phenylalanine; D002403:Cathepsins; D010450:Endopeptidases; D012697:Serine Endopeptidases; C534094:CTSG protein, human; D056649:Cathepsin G; D010196:Pancreatic Elastase