Identification of proteases involved in the proteolysis of vascular endothelium cadherin during neutrophil transmigration.
Transmigration of neutrophils across the endothelium occurs at the cell-cell junctions where the vascular endothelium cadherin (VE cadherin) is expressed. This adhesive receptor was previously demonstrated to be involved in the maintenance of endothelium integrity. We propose that neutrophil transmigration across the vascular endothelium goes in parallel with cleavage of VE cadherin by elastase and cathepsin G present on the surface of neutrophils. This hypothesis is supported by the following lines of evidence. 1) Proteolytic fragments of VE cadherin are released into the culture medium upon adhesion of neutrophils to endothelial cell monolayers; 2) conditioned culture medium, obtained after neutrophil adhesion to endothelial monolayers, cleaves the recombinantly expressed VE cadherin extracellular domain; 3) these cleavages are inhibited by inhibitors of elastase; 4) VE cadherin fragments produced by conditioned culture medium or by exogenously added elastase are identical as shown by N-terminal sequencing and mass spectrometry analysis; 5) both elastase- and cathepsin G-specific VE cadherin cleavage patterns are produced upon incubation with tumor necrosis factor alpha-stimulated and fixed neutrophils; 6) transendothelial permeability increases in vitro upon addition of either elastase or cathepsin G; and 7) neutrophil transmigration is reduced in vitro in the presence of elastase and cathepsin G inhibitors. Our results suggest that cleavage of VE cadherin by neutrophil surface-bound proteases induces formation of gaps through which neutrophils transmigrate.
Hermant
Bastien
B
Laboratoire d'Ingénierie des Macromolécules, Institut de Biologie Structurale Jean-Pierre Ebel, Commissariat à l'Energie Atomique/CNRS, Université Joseph Fourier, 41 rue Jules Horowitz, 38027 Grenoble Cedex, France.
Bibert
Stephanie
S
Concord
Evelyne
E
Dublet
Bernard
B
Weidenhaupt
Marianne
M
Vernet
Thierry
T
Gulino-Debrac
Danielle
D
D000818:Animals; D015703:Antigens, CD; D015153:Blotting, Western; D016466:CHO Cells; D015820:Cadherins / Q000737:chemistry* / Q000378:metabolism* / Q000502:physiology; D056649:Cathepsin G; D002403:Cathepsins / Q000378:metabolism; D002448:Cell Adhesion; D002465:Cell Movement; D002478:Cells, Cultured; D006224:Cricetinae; D003470:Culture Media / Q000494:pharmacology; D017077:Culture Media, Conditioned / Q000494:pharmacology; D004727:Endothelium / Q000378:metabolism; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism*; D006801:Humans; D007962:Leukocytes / Q000378:metabolism; D013058:Mass Spectrometry; D008856:Microscopy, Fluorescence; D009504:Neutrophils / Q000201:enzymology / Q000378:metabolism*; D010196:Pancreatic Elastase / Q000378:metabolism; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D012697:Serine Endopeptidases; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013997:Time Factors; D014471:Umbilical Veins / Q000166:cytology; D000099152:Cadherin 5
D015703:Antigens, CD; D015820:Cadherins; D056649:Cathepsin G; D002403:Cathepsins; D003470:Culture Media; D017077:Culture Media, Conditioned; D010196:Pancreatic Elastase; D011506:Proteins; D011994:Recombinant Proteins; D012697:Serine Endopeptidases; D000099152:Cadherin 5; C534094:CTSG protein, human