title: Controlled enzymatic production of astrocytic hydrogen peroxide protects neurons from oxidative stress via an Nrf2-independent pathway.
abstract: Neurons rely on their metabolic coupling with astrocytes to combat oxidative stress. The transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2) appears important for astrocyte-dependent neuroprotection from oxidative insults. Indeed, Nrf2 activators are effective in stroke, Parkinson disease, and Huntington disease models. However, key endogenous signals that initiate adaptive neuroprotective cascades in astrocytes, including activation of Nrf2-mediated gene expression, remain unclear. Hydrogen peroxide (H(2)O(2)) plays an important role in cell signaling and is an attractive candidate mediator of adaptive responses in astrocytes. Here we determine (i) the significance of H(2)O(2) in promoting astrocyte-dependent neuroprotection from oxidative stress, and (ii) the relevance of H(2)O(2) in inducing astrocytic Nrf2 activation. To control the duration and level of cytoplasmic H(2)O(2) production in astrocytes cocultured with neurons, we heterologously expressed the H(2)O(2)-producing enzyme Rhodotorula gracilis D-amino acid oxidase (rgDAAO) selectively in astrocytes. Exposure of rgDAAO-astrocytes to D-alanine lead to the concentration-dependent generation of H(2)O(2). Seven hours of low-level H(2)O(2) production (∼3.7 nmol·min·mg protein) in astrocytes protected neurons from oxidative stress, but higher levels (∼130 nmol·min·mg protein) were neurotoxic. Neuroprotection occurred without direct neuronal exposure to astrocyte-derived H(2)O(2), suggesting a mechanism specific to astrocytic intracellular signaling. Nrf2 activation mimicked the effect of astrocytic H(2)O(2) yet H(2)O(2)-induced protection was independent of Nrf2. Astrocytic protein tyrosine phosphatase inhibition also protected neurons from oxidative death, representing a plausible mechanism for H(2)O(2)-induced neuroprotection. These findings demonstrate the utility of rgDAAO for spatially and temporally controlling intracellular H(2)O(2) concentrations to uncover unique astrocyte-dependent neuroprotective mechanisms.
authors.affiliation: Department of Neurology and Neuroscience, Weill Medical College of Cornell University, The Burke Medical Research Institute, White Plains, NY 10605, USA. rrr2001@med.cornell.edu
authors.forename: Renée E
authors.initials: RE
authors.lastname: Haskew-Layton
authors.forename: Jimmy B
authors.initials: JB
authors.lastname: Payappilly
authors.forename: Natalya A
authors.initials: NA
authors.lastname: Smirnova
authors.forename: Thong C
authors.initials: TC
authors.lastname: Ma
authors.forename: Kelvin K
authors.initials: KK
authors.lastname: Chan
authors.forename: Timothy H
authors.initials: TH
authors.lastname: Murphy
authors.forename: Hengchang
authors.initials: H
authors.lastname: Guo
authors.forename: Brett
authors.initials: B
authors.lastname: Langley
authors.forename: Rukhsana
authors.initials: R
authors.lastname: Sultana
authors.forename: D Allan
authors.initials: DA
authors.lastname: Butterfield
authors.forename: Sandro
authors.initials: S
authors.lastname: Santagata
authors.forename: Melissa J
authors.initials: MJ
authors.lastname: Alldred
authors.forename: Irina G
authors.initials: IG
authors.lastname: Gazaryan
authors.forename: George W
authors.initials: GW
authors.lastname: Bell
authors.forename: Stephen D
authors.initials: SD
authors.lastname: Ginsberg
authors.forename: Rajiv R
authors.initials: RR
authors.lastname: Ratan
mesh_terms: D000818:Animals; D001253:Astrocytes / Q000166:cytology / Q000378:metabolism*; D002478:Cells, Cultured; D018920:Coculture Techniques; D003605:D-Amino-Acid Oxidase / Q000378:metabolism; D005978:Glutathione / Q000378:metabolism; D006861:Hydrogen Peroxide / Q000378:metabolism*; D046228:Microarray Analysis; D051267:NF-E2-Related Factor 2 / Q000378:metabolism; D009474:Neurons / Q000166:cytology / Q000378:metabolism*; D018696:Neuroprotective Agents / Q000378:metabolism*; D016877:Oxidants / Q000378:metabolism*; D018384:Oxidative Stress / Q000502:physiology*; D051381:Rats; D012248:Rhodotorula / Q000201:enzymology
chemical_list: D051267:NF-E2-Related Factor 2; D018696:Neuroprotective Agents; D016877:Oxidants; D006861:Hydrogen Peroxide; D003605:D-Amino-Acid Oxidase; D005978:Glutathione