title: PR3 and elastase alter PAR1 signaling and trigger vWF release via a calcium-independent mechanism from glomerular endothelial cells.
abstract: Neutrophil proteases, proteinase-3 (PR3) and elastase play key roles in glomerular endothelial cell (GEC) injury during glomerulonephritis. Endothelial protease-activated receptors (PARs) are potential serine protease targets in glomerulonephritis. We investigated whether PAR1/2 are required for alterations in GEC phenotype that are mediated by PR3 or elastase during active glomerulonephritis. Endothelial PARs were assessed by flow cytometry. Thrombin, trypsin and agonist peptides for PAR1 and PAR2, TFLLR-NH(2) and SLIGKV-NH(2,) respectively, were used to assess alterations in PAR activation induced by PR3 or elastase. Endothelial von Willebrand Factor (vWF)release and calcium signaling were used as PAR activation markers. Both PR3 and elastase induced endothelial vWF release, with elastase inducing the highest response. PAR1 peptide induced GEC vWF release to the same extent as PR3. However, knockdown of PARs by small interfering RNA showed that neither PAR1 nor PAR2 activation caused PR3 or elastase-mediated vWF release. Both proteases interacted with and disarmed surface GEC PAR1, but there was no detectable interaction with cellular PAR2. Neither protease induced a calcium response in GEC. Therefore, PAR signaling and serine protease-induced alterations in endothelial function modulate glomerular inflammation via parallel but independent pathways.
authors.affiliation: Schools of Immunity and Infection, College of Medicine and Dentistry, University of Birmingham, Birmingham, United Kingdom. s.p.tull@bham.ac.uk
authors.forename: Samantha P
authors.initials: SP
authors.lastname: Tull
authors.forename: Anne
authors.initials: A
authors.lastname: Bevins
authors.forename: Sahithi Jyothsna
authors.initials: SJ
authors.lastname: Kuravi
authors.forename: Simon C
authors.initials: SC
authors.lastname: Satchell
authors.forename: Bahjat
authors.initials: B
authors.lastname: Al-Ani
authors.forename: Stephen P
authors.initials: SP
authors.lastname: Young
authors.forename: Lorraine
authors.initials: L
authors.lastname: Harper
authors.forename: Julie M
authors.initials: JM
authors.lastname: Williams
authors.forename: George Ed
authors.initials: GE
authors.lastname: Rainger
authors.forename: Caroline O S
authors.initials: CO
authors.lastname: Savage
mesh_terms: D002118:Calcium / Q000378:metabolism; D042783:Endothelial Cells / Q000166:cytology* / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D007678:Kidney Glomerulus / Q000166:cytology*; D053146:Myeloblastin / Q000378:metabolism*; D010196:Pancreatic Elastase / Q000378:metabolism*; D059748:Proteolysis; D044463:Receptor, PAR-1 / Q000378:metabolism*; D015398:Signal Transduction; D014841:von Willebrand Factor / Q000378:metabolism*
chemical_list: D044463:Receptor, PAR-1; D014841:von Willebrand Factor; D010196:Pancreatic Elastase; D053146:Myeloblastin; D002118:Calcium