Nox4 is a novel inducible source of reactive oxygen species in monocytes and macrophages and mediates oxidized low density lipoprotein-induced macrophage death

RATIONALE: The enhanced formation of intracellular reactive oxygen species (ROS) induced by oxidized low-density lipoprotein (OxLDL) promotes macrophage death, a process likely to contribute to the formation of necrotic cores and the progression of atherosclerotic lesions. Yet macrophage deficiency of phagocytic NADPH oxidase (Nox2), the primary source of ROS in macrophages, does not reduce atherosclerotic lesion development in mice. This suggests an as yet unidentified NADPH oxidase may be present in macrophages and responsible for the intracellular ROS formation induced by OxLDL. OBJECTIVE: The aim of this study was to identify the source of intracellular ROS involved in macrophage death. METHODS AND RESULTS: Nox4 was expressed in human monocytes and mature macrophages, and was localized to the endoplasmic reticulum and to defined foci within the nucleus. Nox4 colocalized with p22^phox, and both proteins were upregulated in response to OxLDL stimulation, whereas Nox2/gp91 ^phox levels remained unchanged. Induction of Nox4 expression, intracellular ROS formation and macrophage cytotoxicity induced by OxLDL were blocked by MEK1/2 inhibition, but not by inhibitors of p38-MAPK (mitogen-activated protein kinase), JNK (Jun N-terminal kinase), or JAK2 (Janus kinase 2). Small interfering RNA knockdown of Nox4 inhibited both intracellular ROS production and macrophage cytotoxicity induced by OxLDL, whereas Nox4 overexpression enhanced both OxLDL-stimulated ROS formation and macrophage death. CONCLUSIONS: Nox4 is a novel source of intracellular ROS in human monocytes and macrophages. Induction of Nox4 by OxLDL is mediated by the MEK1/ERK pathway and required for OxLDL cytotoxicity in human macrophages, implicating monocytic Nox4 in atherogenesis.