The adoption of Warburg metabolism is critical for the activation of macrophages in response to lipopolysaccharide. Macrophages stimulated with lipopolysaccharide increase their expression of nicotinamide phosphoribosyltransferase (NAMPT), a key enzyme in NAD⁺ salvage, and loss of NAMPT activity alters their inflammatory potential. However, the events that lead to the cells' becoming dependent on NAD⁺ salvage remain poorly defined. We found that depletion of NAD⁺ and increased expression of NAMPT occurred rapidly after inflammatory activation and coincided with DNA damage caused by reactive oxygen species (ROS). ROS produced by complex III of the mitochondrial electron-transport chain were required for macrophage activation. DNA damage was associated with activation of poly(ADP-ribose) polymerase, which led to consumption of NAD⁺. In this setting, increased NAMPT expression allowed the maintenance of NAD⁺ pools sufficient for glyceraldehyde-3-phosphate dehydrogenase activity and Warburg metabolism. Our findings provide an integrated explanation for the dependence of inflammatory macrophages on the NAD⁺ salvage pathway.