title: Lung endothelial heparan sulfates mediate cationic peptide-induced barrier dysfunction: a new role for the glycocalyx.
abstract: The endothelial glycocalyx is believed to play a major role in microvascular permeability. We tested the hypothesis that specific components of the glycocalyx, via cytoskeletal-mediated signaling, actively participate in barrier regulation. With the use of polymers of arginine and lysine as a model of neutrophil-derived inflammatory cationic proteins, we determined size- and dose-dependent responses of cultured bovine lung microvascular endothelial cell permeability as assessed by transendothelial electrical resistance (TER). Polymers of arginine and lysine >11 kDa produced maximal barrier dysfunction as demonstrated by a 70% decrease in TER. Monomers of l-arginine and l-lysine did not alter barrier function, suggesting a cross-linking requirement of cell surface "receptors". To test the hypothesis that glycosaminoglycans (GAGs) are candidate receptors for this response, we used highly selective enzymes to remove specific GAGs before polyarginine (PA) treatment and examined the effect on TER. Heparinase III attenuated PA-induced barrier dysfunction by 50%, whereas heparinase I had no effect. To link changes in barrier function with structural alterations, we examined actin organization and syndecan localization after PA. PA induced actin stress fiber formation and clustering of syndecan-1 and syndecan-4, which were significantly attenuated by heparinase III. PA-induced cytoskeletal rearrangement and barrier function did not involve myosin light chain kinase (MLCK) or p38 MAPK, as ML-7, a specific MLCK inhibitor, or SB-20358, a p38 MAPK inhibitor, did not alter PA-induced barrier dysfunction. In summary, lung endothelial cell heparan sulfate proteoglycans are key participants in inflammatory cationic peptide-induced signaling that links cytoskeletal reorganization with subsequent barrier dysfunction.
authors.affiliation: Anesthesiology and Critical Care Medicine, Department of Medicine, Johns Hopkins School of Medicine, Baltimore, Maryland 21287, USA. Randal.Dull@hsc.utah.edu
authors.forename: Randal O
authors.initials: RO
authors.lastname: Dull
authors.forename: Ramani
authors.initials: R
authors.lastname: Dinavahi
authors.forename: Lawrence
authors.initials: L
authors.lastname: Schwartz
authors.forename: Donald E
authors.initials: DE
authors.lastname: Humphries
authors.forename: David
authors.initials: D
authors.lastname: Berry
authors.forename: Ram
authors.initials: R
authors.lastname: Sasisekharan
authors.forename: Joe G N
authors.initials: JG
authors.lastname: Garcia
mesh_terms: D000818:Animals; D002412:Cations; D002417:Cattle; D002463:Cell Membrane Permeability / Q000187:drug effects; D002478:Cells, Cultured; D002810:Chondroitinases and Chondroitin Lyases / Q000378:metabolism; D004187:Disaccharides / Q000737:chemistry / Q000494:pharmacology; D004594:Electrophysiology / Q000379:methods; D004730:Endothelium, Vascular / Q000187:drug effects / Q000502:physiology* / Q000503:physiopathology*; D019276:Glycocalyx / Q000187:drug effects / Q000502:physiology*; D006025:Glycosaminoglycans / Q000502:physiology; D019763:Heparin Lyase / Q000378:metabolism; D006497:Heparan Sulfate / Q000502:physiology*; D008562:Membrane Glycoproteins / Q000378:metabolism; D008833:Microcirculation / Q000502:physiology; D010455:Peptides / Q000494:pharmacology*; D011107:Polylysine / Q000494:pharmacology*; D011509:Proteoglycans / Q000378:metabolism; D011652:Pulmonary Circulation / Q000502:physiology*; D053671:Syndecan-4
chemical_list: D002412:Cations; D002810:Chondroitinases and Chondroitin Lyases; D004187:Disaccharides; D006025:Glycosaminoglycans; D019763:Heparin Lyase; D006497:Heparan Sulfate; D008562:Membrane Glycoproteins; D010455:Peptides; D011107:Polylysine; D011509:Proteoglycans; D053671:Syndecan-4; C015462:polyarginine