Single-cell analysis reveals cell death as driver of NLRP3-mediated secretion of IL-1β in human monocytes

Interleukin-1β (IL-1β) is a key proinflammatory cytokine with critical roles in infections and inflammatory diseases, yet the mechanisms regulating its release from human monocytes remain unclear. Here we used a suite of single-cell approaches, including integrated live-cell imaging of secretion and cell fate, flow cytometry and high-content imaging, to investigate IL-1β secretion dynamics in lipopolysaccharide-stimulated primary human peripheral blood CD14⁺ monocytes. We found marked heterogeneity: a large fraction of cells remained viable and contributed negligibly to IL-1β secretion, challenging established models. Instead, a small subset (5-10%) undergoing canonical NLRP3 inflammasome activation and GSDMD-dependent pyroptosis produced the majority of secreted IL-1β, with a smaller contribution from apoptotic cells transitioning to secondary necrosis. Single-cell profiling of CD14⁺ monocytes from patients with cryopyrin-associated periodic syndrome confirmed lytic cell death as the driver of pathological IL-1β release. These findings redefine IL-1β as a damage-associated molecular pattern, secreted predominantly by dying monocytes.