Neurobiology of Aging 27 (2006) 1577–1587 A key role for the microglial NADPH oxidase in APP-dependent killing of neurons Bin Qin, Laetitia Cartier, Michel Dubois-Dauphin, Bin Li, Lena Serrander, Karl-Heinz Krause ∗ Biology of Ageing Laboratory, Department of Geriatrics, Geneva University Hospitals, 2 chemin Petit Bel-Air, 1225 Chˆ ene-Bourg, Geneva, Switzerland Received 15 July 2005; received in revised form 14 September 2005; accepted 20 September 2005 Available online 2 November 2005 Abstract Reactive oxygen species (ROS) and deposition of cleaved products of amyloid precursor protein (APP) are thought to contribute to neuronal loss observed in Alzheimer’s disease (AD). The relationship between these factors was studied in a neuroblastoma and microglia co-culture system. Overexpression of wild-type APP (APP-wt) or APP with three mutations typical of familial AD (APP-3m) in SH-SY5Y neuroblastoma cells did not directly alter their morphology, growth rate, cell cycle or H 2 O 2 sensitivity. In a co-culture of APP-wt neuroblastoma cells with microglia, microglial cells generated ROS and neuronal cells died. The cell death was more pronounced in APP-3m-expressing neurons. Neuroblastoma cell death was attenuated by ROS-scavengers and was dose-dependently inhibited by the NADPH oxidase inhibitor diphenyleneiodonium chloride (DPI). Macrophage cell lines behaved similarly to microglia in the co-culture model. However, a macrophage cell line deficient in the NADPH oxidase subunit, gp91phox, failed to kill neurons. These results suggest that APP-dependent microglia activation and subsequent ROS generation by the phagocyte NADPH oxidase play a crucial role in neuronal killing in a cellular model of AD. © 2005 Elsevier Inc. All rights reserved. Keywords: Alzheimers disease; Amyloid precursor protein; NOX2; NADPH oxidase; Microglia; Neuroblastoma cells 1. Introduction Alzheimer’s disease (AD) is the most common cause of dementia in the Western world and probably affects around one-third of the population over 80 years of age. From a bio- chemical point of view, a key step in the pathogenesis of AD is the neuronal processing of the amyloid precursor pro- tein (APP) yielding small, hydrophobic peptides, in particular A40 and A42, which in turn will aggregate to form amy- loid plaques. The rare familial form of Alzheimer’s disease may be caused by APP mutations, which lead to an enhanced generation of amyloidogenic peptides (for general review see Ref. [21]). The relationship between -amyloid and neuronal death in AD is complex. Exogenous addition or endogenous gen- eration of amyloidogenic peptides may show a direct neu- ∗ Corresponding author. Tel.: +41 22 3055450; fax: +41 22 3055455. E-mail address: karl-heinz.krause@medecine.unige.ch (K.-H. Krause). rotoxicity [15,20,33]. However, under the same conditions, activation of microglia, and subsequent release of a variety of potentially neurotoxic compounds, including reactive oxy- gen species also occurs [14,8,17]. Thus, it appears likely that neuronal death in AD is only in part due to a direct neuro- toxicity of amyloidogenic peptides, and involves in addition neuronal killing through activated microglia. Activated microglia may kill neurons through mechanisms involving cytokine and chemokine release, such as TNF, Fas-ligand and -chemokines [5,28,6]. However, there is also ample evidence for the involvement of reactive oxygen species (ROS). Recently, we have reviewed presently exist- ing, indirect evidence that NOX family NADPH oxidases might be involved in the neuronal killing in the context of AD [34]. In this study, we have investigated killing of neurons in a co-culture model of APP-expressing neurons and microglia with a focus on the role and the source of ROS in this system. 0197-4580/$ – see front matter © 2005 Elsevier Inc. All rights reserved. doi:10.1016/j.neurobiolaging.2005.09.036