IL-18 is a proinflammatory cytokine belonging to the “IL-1 family” that has been shown to play a prominent role in the induction of type 1 immune responses. Here, we show that M-CSFinduces the expression of a membrane-bound form of IL-18 (mIL-18) in a subset of human blood monocytes differentiating toward macrophages. While monocytes, DC, and GM-CSF-treated monocytes did not express mIL-18, its expression was detected in approximately 30–40% of M-CSF-primed macrophages differentiating from both CD16− and CD16+ monocytes. Treatment with the caspase-1 inhibitor significantly reduced mIL-18 expression suggesting the requirement of an assembled inflammasome for IL-18 surface expression. Polarization toward M2 did not modify mIL-18 expression. On the contrary, LPS stimulation of both M0 and M2 (mIL-18+) macrophages induced shedding of mIL-18, which was likely mediated by the activation of cellular protease(s). Importantly, the soluble form IL-18 (sIL-18) induced in autologous resting NK cells both the expression of CCR7 and the production of high amounts of IFN-γ, which was virtually abrogated by Ab-mediated neutralization of sIL-18. Overall our data shed new light on the cells and mechanisms leading to the release of sIL-18, the major IFN-γ-inducing factor in both physiological and pathological immune responses.