It is widely believed that DC, but not macrophages, prime na??ve T cells in vivo. Here, we investigated the ability of CD68-expressing cells (commonly defined as macrophages) in priming autoreactive T cells and initiating collagen-induced arthritis (CIA) in the mouse. For this purpose, a transgenic mouse was developed (MBQ mouse) where macrophages exclusively expressed the MHC class II H2-A q (A q) on an H2-A p (A p) background. A q, but not A p expression mediates susceptibility to CIA through presentation of type II collagen (CII) to T cells. CIA severity is enhanced by a mutation in the Ncf1 gene, impairing reactive oxygen species (ROS) production by the phagocyte NADPH oxidase (NOX2) complex. Expression of functional Ncf1 on macrophages was previously shown to protect from severe CIA. To study the effect of ROS on macrophage-mediated priming of T cells, the Ncf1 mutation was introduced in the MBQ mouse. Upon CII immunization, Ncf1-mutated MBQ mice, but not Ncf1 wild-type MBQ mice nor Ncf1-mutated A p mice, activated autoreactive T cells and developed CIA. These findings demonstrate for the first time that macrophages can initiate arthritis and that the process is negatively regulated by ROS produced via the NOX2 complex. Copyright A? 2011 WILEY-VCH Verlag GmbH Co. KGaA, Weinheim.