Macrophages play essential roles in a wide variety of physiological and pathological processes. One of the unique features of these phagocytic leukocytes is their ability to fuse, forming multinucleated giant cells. Multinucleated giant cells are important mediators of tissue remodeling and repair and are also responsible for removal or sequestration of foreign material, intracellular bacteria and non-phagocytosable pathogens, such as parasites and fungi. Depending on the tissue where fusion occurs and the inflammatory insult, multinucleated giant cells assume distinctly different phenotypes. Nevertheless, the ultimate outcome is the formation of large cells that can resorb bone tissue (osteoclasts) or foreign material and pathogens (giant cells) extracellularly. While progress has been made in recent years, the mechanisms and factors involved in macrophage fusion are still not fully understood. In addition to cytokines and a number of adhesion proteins and receptors, it is becoming increasingly clear that NADPH oxidase-generated reactive oxygen species (ROS) also play an important role in macrophage fusion. In this review, we provide an overview of macrophage multinucleation, with a specific focus on the role of NADPH oxidases and ROS in macrophage fusion and in the function of multinucleated giant cells. In addition, we provide an updated overview of the role of these cells in inflammation and various autoimmune diseases.