The Nod-like receptor protein 3 (NLRP3) inflammasome is considered to be a pivotal host platform responsible for sensing of exogenous and endogenous danger signals, including those generated as a result of metabolic dysregulation, and for the subsequent, IL-1β-mediated orchestration of inflammatory and innate immunity responses. In this way, although the molecular link between diet-induced obesity and inflammasome activation is still unclear, free fatty acids (FFA) have been proposed as a triggering event. We report that dietary fatty acid (FA) composition is sensed by the NLRP3 inflammasome in human macrophages. For this purpose, we have analysed three roles of FA supplementation: as a priming signal for ATP-activated macrophages, in determining where the administration of dietary FAs interferes with LPS-mediated inflammasome activation and by inducing inflammasome activation per se. In this study, we confirm that saturated (SFAs) activated the NLRP3 inflammasome and stimulated the secretion of the IL-1β cytokine, while PUFAs were mainly inhibitors. Moreover, in general, DHA (n-3 PUFA) was more effective in preventing inflammasome activation than arachidonic acid (n-6 PUFA).