Why does the hemolytic activity of silica predict its pro-inflammatory activity?

The hemolytic activity of inhaled particles such as silica has been widely investigated in the past and represents a usual toxicological endpoint to characterize particle reactivity despite the fact that red blood cells (RBCs) are not involved in the pathogenesis of pulmonary inflammation or fibrosis caused by some inhaled particles. The inflammatory process induced by silica starts with the activation of the inflammasome, which leads to the release of mature IL-1β. One of the upstream mechanisms causing activation of the inflammasome is the labilization of the phagolysosomal membrane after particle phagocytosis. Considering RBC lysis as a model of membrane damage, we evaluated the relationship between hemolytic activity and inflammasome-dependent release of IL-1β for a panel of selected silica particles, in search of the toxicological significance of the hemolytic activity of an inhaled particle. Well-characterized silica particles, including four quartz samples and a vitreous silica, with different surface properties and hemolytic potential were tested for their capacity to induce inflammasome-dependent release of IL-1β in LPS-primed primary murine peritoneal macrophages by ELISA and Western blot analysis. The mechanisms of IL-1β maturation and release were clarified by using ASC-deficient cells and inhibitors of phagocytosis and cathepsin B. The silica samples induced dose-dependent hemolysis and IL-1β release of different amplitudes. A significant correlation between IL-1β release and hemolytic activity was evidenced (r = 0.827) by linear regression analysis. IL-1β release was completely abolished in ASC-deficient cells and reduced by inhibitors, confirming the involvement of the inflammasome and the requirement of phagocytosis and cathepsin B for activation. The same physico-chemical properties of silica particles which are relevant for the lysis of the RBC membrane also appear implicated in the labilization of the phagolysosome, leading to inflammasome activation and release of the pro-inflammatory cytokine IL-1β. These findings strengthen the relevance of the hemolysis assay to predict the pro-inflammatory activity of silica dusts.