Neutrophil-mediated increased permeability of microcarrier-cultured endothelial monolayers: a model for the in vitro study of neutrophil-dependent mediators of vasopermeability.
Changes in the permeability of human endothelial monolayers in response to activated human neutrophils were examined in a novel, in vitro model of vasopermeability changes. Microcarrier-cultured human umbilical vein endothelial monolayers were used in a system that responds to histamine. Human neutrophils did not increase Evans Blue staining of the endothelium-covered microcarriers if added alone or if added with the neutrophil-dependent mediator of vasopermeability, formyl-methionyl-leucyl-phenylalanine (FMLP, 0.1 microM). In contrast, neutrophils, added to the endothelial cells in a ratio as low as 2.5:1 caused time-dependent increases in microcarrier staining if pretreated with cytochalasin B (5 micrograms/mL) before addition with FMLP. Neutrophil cell-free releasate and purified human sputum elastase also caused concentration-related increases in Evans Blue staining of the endothelial-covered microcarriers and these effects were inhibited by the elastase inhibitor methoxysuccinyl-alanyl-alanyl-prolyl-valyl chloromethyl ketone. This compound also inhibited neutrophil-mediated endothelial permeability increases. The microcarrier-cultured human endothelial monolayer system rapidly detects permeability alterations of endothelial monolayers in response to activated human neutrophils. This model is a potentially useful screening assay for the development of therapeutic agents, directed at neutrophil degranulation or degranulation products, for the control of inflammatory vasopermeability abnormalities.
Killackey
J J
JJ
Syntex Research, Mississauga, Ont., Canada.
Killackey
B A
BA
D002199:Capillary Permeability / Q000502:physiology*; D056649:Cathepsin G; D002403:Cathepsins / Q000378:metabolism; D002478:Cells, Cultured; D003571:Cytochalasin B / Q000494:pharmacology; D004730:Endothelium, Vascular / Q000378:metabolism* / Q000502:physiology; D005609:Free Radicals; D006801:Humans; D066298:In Vitro Techniques; D008954:Models, Biological; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D009504:Neutrophils / Q000502:physiology*; D012697:Serine Endopeptidases; D012835:Silver Nitrate; D013183:Sputum / Q000201:enzymology; D013194:Staining and Labeling; D013481:Superoxides / Q000378:metabolism
D005609:Free Radicals; D013481:Superoxides; D003571:Cytochalasin B; D009240:N-Formylmethionine Leucyl-Phenylalanine; D012835:Silver Nitrate; D002403:Cathepsins; D012697:Serine Endopeptidases; C534094:CTSG protein, human; D056649:Cathepsin G