The role of NOX4 and its interaction with mitochondria and Nrf2 in hyperexcitability-induced neurodegeneration

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Hyperexcitability is a common mechanism leading to neurodegeneration in a plethora of central nervous system (CNS) diseases. It occurs amongst others in epilepsy and multiple sclerosis (MS) where reactive oxygen species (ROS) have been shown to play a role. Despite overwhelming evidence of their critical role in neurodegeneration, targeting ROS has been challenging, and clinical trials aimed at scavenging ROS, once they have been produced, have largely failed. One explanation for this is that scavenging of ROS with direct antioxidants is futile since large doses and a sustained supply of antioxidants are needed, which by themselves might have other adverse effects. Thus, targeting sources of ROS, rather than ROS once they have already been produced, represents an appealing strategy. This in turn requires a careful characterization of the sources of ROS production, as some may be beneficial to the cell.We have previously shown that nicotinamide adenine dinucleotide phosphate-oxidase (NADPH) oxidases, which are enzymes highly specialized in ROS production, play a pivotal role in hyperexcitability-induced neurodegeneration. Seven different isoforms of NADPH oxidases exist (NOX1-5, and DUOX1 and 2). In preliminary experiments, we recently found high expression of NOX4 in brain resident cells, and that NOX4 interacts both with mitochondria and nuclear factor erythroid 2–related factor 2 (Nrf2). On the one hand, these interactions are important since energy depletion and thus mitochondria, as the “powerhouses of the cell”, play a lead role in hyperexcitability-induced neurodegeneration. On the other hand, the transcription factor Nrf2, which upon activation drives transcription of antioxidant proteins, has also been shown to contribute to neurodegeneration during hyperexcitability. This implies that targeting NOX4 is likely a promising strategy to combat hyperexcitability-induced neurodegeneration.We here propose to first study the impact of NOX4 on hyperexcitability-induced neurodegeneration and the interaction between NOX4 and mitochondria and NOX4 and Nrf2 during hyperexcitability. This will be done with live cell imaging techniques in different brain resident cells (astrocytes, neurons, endothelial cells and microglia) in addition to molecular biology analyses in NOX4-deficient mice. Next, we will use a mouse model of hyperexcitability-induced neurodegeneration to study the role of NOX4, taking advantage of global and tissue-specific NOX4-deficient animals, i.e. mice with a NOX4 deficiency in endothelial or neuronal tissue (NOX4endo -/- and NOX4neuro -/-) and live-cell imaging analyses in brain slices in addition to electrophysiology and immunohistochemistry. Finally, we will corroborate data obtained from animal experiments by analysing post mortem human brain tissues of patients suffering from MS, a disease characterized by hyperexcitability-induced neurodegeneration, using immunohistochemistry.