title: Protein kinase C-α and arginase I mediate pneumolysin-induced pulmonary endothelial hyperpermeability.
abstract: Antibiotics-induced release of the pore-forming virulence factor pneumolysin (PLY) in patients with pneumococcal pneumonia results in its presence days after lungs are sterile and is a major factor responsible for the induction of permeability edema. Here we sought to identify major mechanisms mediating PLY-induced endothelial dysfunction. We evaluated PLY-induced endothelial hyperpermeability in human lung microvascular endothelial cells (HL-MVECs) and human lung pulmonary artery endothelial cells in vitro and in mice instilled intratracheally with PLY. PLY increases permeability in endothelial monolayers by reducing stable and dynamic microtubule content and modulating VE-cadherin expression. These events, dependent upon an increased calcium influx, are preceded by protein kinase C (PKC)-α activation, perturbation of the RhoA/Rac1 balance, and an increase in myosin light chain phosphorylation. At later time points, PLY treatment increases the expression and activity of arginase in HL-MVECs. Arginase inhibition abrogates and suppresses PLY-induced endothelial barrier dysfunction by restoring NO generation. Consequently, a specific PKC-α inhibitor and the TNF-derived tonoplast intrinsic protein peptide, which blunts PLY-induced PKC-α activation, are able to prevent activation of arginase in HL-MVECs and to reduce PLY-induced endothelial hyperpermeability in mice. Arginase I (AI)(+/-)/arginase II (AII)(-/-) C57BL/6 mice, displaying a significantly reduced arginase I expression in the lungs, are significantly less sensitive to PLY-induced capillary leak than their wild-type or AI(+/+)/AII(-/-) counterparts, indicating an important role for arginase I in PLY-induced endothelial hyperpermeability. These results identify PKC-α and arginase I as potential upstream and downstream therapeutic targets in PLY-induced pulmonary endothelial dysfunction.
authors.affiliation: Vascular Biology Center and Dept. of Pharmacology and Toxicology, Georgia Health Sciences University, 1459 Laney-Walker Blvd., Augusta, GA 30912-2500, USA. rlucas@georgiahealth.edu
authors.forename: Rudolf
authors.initials: R
authors.lastname: Lucas
authors.forename: Guang
authors.initials: G
authors.lastname: Yang
authors.forename: Boris A
authors.initials: BA
authors.lastname: Gorshkov
authors.forename: Evgeny A
authors.initials: EA
authors.lastname: Zemskov
authors.forename: Supriya
authors.initials: S
authors.lastname: Sridhar
authors.forename: Nagavedi S
authors.initials: NS
authors.lastname: Umapathy
authors.forename: Agnieszka
authors.initials: A
authors.lastname: Jezierska-Drutel
authors.forename: Irina B
authors.initials: IB
authors.lastname: Alieva
authors.forename: Martin
authors.initials: M
authors.lastname: Leustik
authors.forename: Hamid
authors.initials: H
authors.lastname: Hossain
authors.forename: Bernhard
authors.initials: B
authors.lastname: Fischer
authors.forename: John D
authors.initials: JD
authors.lastname: Catravas
authors.forename: Alexander D
authors.initials: AD
authors.lastname: Verin
authors.forename: Jean-François
authors.initials: JF
authors.lastname: Pittet
authors.forename: Ruth B
authors.initials: RB
authors.lastname: Caldwell
authors.forename: Timothy J
authors.initials: TJ
authors.lastname: Mitchell
authors.forename: Stephen D
authors.initials: SD
authors.lastname: Cederbaum
authors.forename: David J
authors.initials: DJ
authors.lastname: Fulton
authors.forename: Michael A
authors.initials: MA
authors.lastname: Matthay
authors.forename: Robert W
authors.initials: RW
authors.lastname: Caldwell
authors.forename: Maritza J
authors.initials: MJ
authors.lastname: Romero
authors.forename: Trinad
authors.initials: T
authors.lastname: Chakraborty
mesh_terms: D000818:Animals; D015703:Antigens, CD / Q000378:metabolism; D001119:Arginase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D001426:Bacterial Proteins / Q000494:pharmacology; D015820:Cadherins / Q000378:metabolism; D020013:Calcium Signaling; D002199:Capillary Permeability; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000201:enzymology / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D008168:Lung / Q000098:blood supply / Q000276:immunology / Q000473:pathology*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008870:Microtubules / Q000378:metabolism; D055806:Microvessels / Q000473:pathology; D011014:Pneumonia / Q000201:enzymology / Q000276:immunology / Q000473:pathology; D051571:Protein Kinase C-alpha / Q000037:antagonists & inhibitors / Q000378:metabolism*; D013301:Streptolysins / Q000494:pharmacology*; D020742:rhoA GTP-Binding Protein / Q000378:metabolism; D000099152:Cadherin 5
chemical_list: D015703:Antigens, CD; D001119:Arginase; D001426:Bacterial Proteins; D015820:Cadherins; D004791:Enzyme Inhibitors; D051571:Protein Kinase C-alpha; D013301:Streptolysins; D020742:rhoA GTP-Binding Protein; D000099152:Cadherin 5; C032290:plY protein, Streptococcus pneumoniae; C574619:ARG1 protein, human; C477631:ARG2 protein, human