Crystal-mediated inflammation in alveolar macrophages of a mouse model of lung fibrosis

Background: Accumulation of crystals of unknown nature was previously observed in alveolar macrophages (AM) in an idiopathic pulmonary fibrosis patient as well as in in Surfactant Protein C deficient mice (SP-C KO), a model of lung fibrosis, where their appearance in AM precedes fibrotic injury. Aim: To explore the nature of crystals in AM and whether they lead to NLRP3 inflammasome activation in SP-C KO mice. Methods: The presence of crystals in AMs was assessed by electron microscopy in aging (10, 30, 60weeks) SP-C KO mice. Bronchoalveolar lavage fluid (BALF)-cell transcriptomic and lipidomic analyses were performed in SP-C KO versus age-matched wild type mice. NLRP3 inflammasome activation was evaluated by western blot of inflammasome proteins and immunofluorescence detection of the inflammasome complex in AM, and by enzymatic immunoassay for the release of inflammatory cytokines in BALF. Results: Needle-shaped crystals were detected in AM in SP-C KO mice from 30weeks of age. At the same age, lipidomics showed higher levels (p<0.05, after FDR correction) of different species of cardiolipins, triglycerides, and cholesterol esters in SP-C KO compared to age-matched wild type mice. SP-C KO mice showed up-regulation of genes coding for steroid catabolism, cytokines and interleukins. Increased levels of NLRP3 related proteins, detection of the complex and release of interleukin-18 in BALF in older SP-C KO mice confirmed activation of inflammasome. Conclusions: These data highlight a link between altered cholesterol metabolism, crystal nucleation and inflammatory response in the context of lung fibrosis, suggesting a mechanism similar to that reported for atherosclerosis.