Controlled enzymatic production of astrocytic hydrogen peroxide protects neurons from oxidative stress via an Nrf2-independent pathway

摘要

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_2O_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_2O_2 in promoting astrocyte-dependent neuroprotection from oxidative stress, and (ii) the relevance of H_2O_2 in inducing astrocytic Nrf2 activation. To control the duration and level of cyto-plasmic H_2O_2 production in astrocytes cocultured with neurons, we heterologously expressed the H_2O_2-producing enzyme Rho-dotorula gracilis D-amino acid oxidase (rgDAAO) selectively in astrocytes. Exposure of rgDAAO-astrocytes to D-alanine lead to the concentration-dependent generation of H_2O_2. Seven hours of low-level H_2O_2 production (～3.7 nmolminmg 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_2O_2, suggesting a mechanism specific to astrocytic intracellular signaling. Nrf2 activation mimicked the effect of astrocytic H_2O_2 yet H_2O_2-induced protection was independent of Nrf2. Astrocytic protein tyrosine phosphatase inhibition also protected neurons from oxidative death, representing a plausible mechanism for H_2O_2-induced neuroprotection. These findings demonstrate the utility of rgDAAO for spatially and temporally controlling intracellular H_2O_2 concentrations to uncover unique astrocyte-dependent neuroprotective mechanisms.