Role of endothelial nitric oxide synthase-derived nitric oxide in activation and dysfunction of cerebrovascular endothelial cells during early onsets of sepsis.
Sepsis-associated encephalopathy is an early manifestation of sepsis, resulting in a diffuse dysfunction of the brain. Recently, nitric oxide (NO) has been proposed to be one of the key molecules involved in the modulation of inflammatory responses in the brain. The aim of this study was to assess the role of NO in cerebrovascular endothelial cell activation/dysfunction during the early onsets of sepsis. To this end, we employed an in vitro model of sepsis in which cultured mouse cerebrovascular endothelial cells (MCVEC) were challenged with blood plasma (20% vol/vol) obtained from sham or septic (feces-induced peritonitis, FIP; 6 h) mice. Exposing MCVEC to FIP plasma for 1 h resulted in increased production of reactive oxygen species and NO as assessed by intracellular oxidation of oxidant-sensitive fluorochrome, dihydrorhodamine 123 (DHR 123), and nitrosation of NO-specific probe, DAF-FM, respectively. The latter events were accompanied by dissociation of tight junction protein, occludin, from MCVEC cytoskeletal framework and a subsequent increase in FITC-dextran (3-kDa mol mass) flux across MCVEC grown on the permeable cell culture supports, whereas Evans blue-BSA (65-kDa mol mass) or FITC-dextran (10-kDa mol mass) flux were not affected. FIP plasma-induced oxidant stress, occludin rearrangement, and MCVEC permeability were effectively attenuated by antioxidant, 1-pyrrolidinecarbodithioic acid (PDTC; 0.5 mM), or interfering with nitric oxide synthase (NOS) activity [0.1 mM nitro-L-arginine methyl ester (L-NAME) or endothelial NOS (eNOS)-deficient MCVEC]. However, treatment of MCVEC with PDTC failed to interfere with NO production, suggesting that septic plasma-induced oxidant stress in MCVEC is primarily a NO-dependent event. Taken together, these data indicate that during early sepsis, eNOS-derived NO exhibits proinflammatory characteristics and contributes to the activation and dysfunction of cerebrovascular endothelial cells.
Handa
Osamu
O
Centre for Critical Illness Research, Lawson Health Research Institute, London, Ontario, Canada.
Stephen
Jancy
J
Cepinskas
Gediminas
G
D000818:Animals; D000975:Antioxidants / Q000494:pharmacology; D001921:Brain / Q000098:blood supply*; D002463:Cell Membrane Permeability / Q000187:drug effects; D002478:Cells, Cultured; D004195:Disease Models, Animal; D004730:Endothelium, Vascular / Q000187:drug effects / Q000201:enzymology* / Q000503:physiopathology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D008565:Membrane Proteins / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D009569:Nitric Oxide / Q000378:metabolism*; D052247:Nitric Oxide Synthase Type II / Q000037:antagonists & inhibitors / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D052250:Nitric Oxide Synthase Type III; D062793:Occludin; D018384:Oxidative Stress; D017382:Reactive Oxygen Species / Q000378:metabolism; D018805:Sepsis / Q000097:blood / Q000201:enzymology* / Q000503:physiopathology; D013997:Time Factors
D000975:Antioxidants; D004791:Enzyme Inhibitors; D008565:Membrane Proteins; D062793:Occludin; C568986:Ocln protein, mouse; D017382:Reactive Oxygen Species; D009569:Nitric Oxide; D052247:Nitric Oxide Synthase Type II; D052250:Nitric Oxide Synthase Type III; C496315:Nos3 protein, mouse