title: Neuronal NAD(P)H oxidases contribute to ROS production and mediate RGC death after ischemia.
abstract: OBJECTIVE
  To study the role of neuronal nicotinamide adenine dinucleotide phosphate [NAD(P)H] oxidase-dependent reactive oxygen species (ROS) production in retinal ganglion cell (RGC) death after ischemia.
  
  
  METHODS
  Ischemic injury was induced by unilateral elevation of intraocular pressure via direct corneal cannulation. For in vitro experiments, RGCs isolated by immunopanning from retinas were exposed to oxygen and glucose deprivation (OGD). The expression levels of NAD(P)H oxidase subunits were evaluated by quantitative PCR, immunocytochemistry, and immunohistochemistry. The level of ROS generated was assayed by dihydroethidium. The NAD(P)H oxidase inhibitors were then tested to determine if inhibition of NAD(P)H oxidase altered the production of ROS within the RGCs and promoted cell survival.
  
  
  RESULTS
  It was reported that RGCs express catalytic Nox1, Nox2, Nox4, Duox1, as well as regulatory Ncf1/p47phox, Ncf2/p67phox, Cyba/p22phox, Noxo1, and Noxa1 subunits of NAD(P)H oxidases under normal conditions and after ischemia. However, whereas RGCs express only low levels of catalytic Nox2, Nox4, and Duox1, and regulatory Ncf1/p47, Ncf2/p67 subunits, they exhibit significantly higher levels of catalytic subunit Nox1 and the subunits required for optimal activity of Nox1. It was observed that the nonselective NAD(P)H oxidase inhibitors VAS-2870, AEBSF, and the Nox1 NAD(P)H oxidase-specific inhibitor ML-090 decreased the ROS burst stimulated by OGD, which was associated with a decreased level of RGC death.
  
  
  CONCLUSIONS
  The findings suggest that NAD(P)H oxidase activity in RGCs renders them vulnerable to ischemic death. Importantly, high levels of Nox1 NAD(P)H oxidase subunits in RGCs suggest that this enzyme could be a major source of ROS in RGCs produced by NAD(P)H oxidases.
authors.affiliation: Bascom Palmer Eye Institute, Department of Ophthalmology, University of Miami Miller School of Medicine, Miami, FL 33136, USA.
authors.forename: Galina
authors.initials: G
authors.lastname: Dvoriantchikova
authors.forename: Jeff
authors.initials: J
authors.lastname: Grant
authors.forename: Andrea Rachelle C
authors.initials: AR
authors.lastname: Santos
authors.forename: Eleut
authors.initials: E
authors.lastname: Hernandez
authors.forename: Dmitry
authors.initials: D
authors.lastname: Ivanov
mesh_terms: D000818:Animals; D016923:Cell Death / Q000502:physiology*; D002478:Cells, Cultured; D004195:Disease Models, Animal; D007150:Immunohistochemistry; D007511:Ischemia / Q000201:enzymology / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D019255:NADPH Oxidases / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism*; D012165:Retinal Ganglion Cells / Q000201:enzymology / Q000378:metabolism*
chemical_list: D012333:RNA, Messenger; D017382:Reactive Oxygen Species; D019255:NADPH Oxidases