Abstract Glycolysis is critical for NLRP3 inflammasome activation, yet the link between lactic acid metabolism and inflammasome signaling remains unclear. Here, we show that stimulation of macrophages with the NLRP3 activators nigericin or ATP induces lactic acid production and efflux via a lactate dehydrogenase–dependent pathway. Accumulation of intracellular lactic acid leads to cytoplasmic acidification, which promotes NLRP3 inflammasome activation. Concurrently, elevated extracellular lactic acid impairs lactate efflux, exacerbating intracellular acidification and amplifying ASC speck formation, caspase-1 activation, and IL-1β secretion. Alkalinization of the extracellular milieu prevents intracellular acidification and abolishes inflammasome activation. Mechanistically, intracellular lactic acidification promoted mitochondrial dysfunction and reactive oxygen species production, and concurrently induced phosphorylation of the stress kinase PKR, which facilitated PKR–NLRP3 interaction and inflammasome assembly through parallel pathways. Independently of inflammasome signaling, lactic acid also directly cleaves pro-IL-1β and pro-IL-18 into mature forms through a mechanism requiring its carboxyl group and mimicking caspase-1 substrate specificity. Mass spectrometry analysis revealed lactic acid–mediated cleavage of pro-IL-1β at Asp116, the canonical caspase-1 site. In a murine model of polymicrobial sepsis induced by cecal ligation and puncture, systemic lactate administration exacerbated inflammation, increased IL-1β levels and neutrophil infiltration, induced hypothermia, and worsened survival. Together, these findings identify intracellular lactic acidification as a metabolic signal that promotes inflammation predominantly through NLRP3 inflammasome activation, while also revealing a potential inflammasome-independent cytokine processing mechanism under conditions of severe metabolic stress.
Lin et al. (2026) studied this question.