title: Reactive oxygen and nitrogen intermediates contribute to Haemophilus somnus lipooligosaccharide-mediated apoptosis of bovine endothelial cells.
abstract: Although Haemophilus somnus causes septicemia and vasculitis in cattle, relatively little is known about how H. somnus affects endothelial cells in vitro. We previously reported that H. somnus lipooligosaccharide (LOS)-induced activation of caspases-3, -8 and -9, and apoptosis of bovine pulmonary artery endothelial cells (BPAEC) in vitro. Previous reports indicate that the generation of reactive oxygen species (ROS) or reactive nitrogen intermediates (RNI) can contribute to the induction of apoptosis. In the present study, we sought to determine whether ROS and RNI are involved in LOS-mediated apoptosis of BPAEC. We found that H. somnus LOS induced the generation of ROS in BPAEC, which was blocked by pretreatment with membrane permeable ROS scavengers, such as dimethylsulfoxide (DMSO) and allopurinol (AP). Addition of DMSO or AP significantly reduced H. somnus LOS-mediated caspase-3 activation. Addition of membrane impermeable ROS scavengers (e.g. catalase and superoxide dismutase), failed to block LOS-mediated caspase-3 activation, suggesting a role for intracellular generation of ROS in LOS-induced apoptosis of BPAEC. Addition of N(G)-nitro-L-arginine methyl ester (L-NAME) or aminoguanidine, which are selective inhibitors of nitric oxide synthase, blocked NO release and significantly reduced caspase-3 activation in LOS treated BPAEC. These data suggest H. somnus LOS triggers endogenous ROS and RNI production by endothelial cells, which contributes to apoptosis.
authors.affiliation: Department of Pathobiological Sciences, School of Veterinary Medicine, 2015 Linden Drive West, Madison, WI 53706, USA.
authors.forename: Matt J
authors.initials: MJ
authors.lastname: Sylte
authors.forename: Thomas J
authors.initials: TJ
authors.lastname: Inzana
authors.forename: Charles J
authors.initials: CJ
authors.lastname: Czuprynski
mesh_terms: D000493:Allopurinol / Q000494:pharmacology; D000818:Animals; D017209:Apoptosis / Q000276:immunology*; D053148:Caspase 3; D020169:Caspases / Q000276:immunology; D002417:Cattle; D002418:Cattle Diseases / Q000276:immunology / Q000382:microbiology*; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D042783:Endothelial Cells / Q000166:cytology / Q000276:immunology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D016166:Free Radical Scavengers / Q000494:pharmacology; D006146:Guanidines / Q000494:pharmacology; D006192:Haemophilus Infections / Q000276:immunology / Q000382:microbiology / Q000662:veterinary*; D044138:Haemophilus somnus / Q000276:immunology*; D008070:Lipopolysaccharides / Q000276:immunology; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D009569:Nitric Oxide / Q000037:antagonists & inhibitors / Q000276:immunology; D026361:Reactive Nitrogen Species / Q000037:antagonists & inhibitors / Q000276:immunology*; D017382:Reactive Oxygen Species / Q000037:antagonists & inhibitors / Q000276:immunology*
chemical_list: D004791:Enzyme Inhibitors; D016166:Free Radical Scavengers; D006146:Guanidines; D008070:Lipopolysaccharides; D026361:Reactive Nitrogen Species; D017382:Reactive Oxygen Species; C023023:lipid-linked oligosaccharides; D009569:Nitric Oxide; D000493:Allopurinol; D053148:Caspase 3; D020169:Caspases; C004479:pimagedine; D019331:NG-Nitroarginine Methyl Ester; D004121:Dimethyl Sulfoxide