SUMMARY We report that in the presence of signal 1 (NF-κB), the NLRP3 inflammasome was activated by mitochondrial apoptotic signaling that licensed production of interleukin-1β (IL-1β). NLRP3 secondary signal activators such as ATP induced mitochondrial dysfunction and apoptosis, resulting in release of oxidized mitochondrial DNA (mtDNA) into the cytosol, where it bound to and activated the NLRP3 inflammasome. The anti-apoptotic protein Bcl-2 inversely regulated mitochondrial dysfunction and NLRP3 inflammasome activation. Mitochondrial DNA directly induced NLRP3 inflammasome activation, because macrophages lacking mtDNA had severely attenuated IL-1β production, yet still underwent apoptosis. Both binding of oxidized mtDNA to the NLRP3 inflammasome and IL-1β secretion could be competitively inhibited by the oxidized nucleoside, 8-OH-dG. Thus, our data reveal that oxidized mtDNA released during programmed cell death causes activation of the NLRP3 inflammasome. These results provide a missing link between apoptosis and inflammasome activation, via binding of cytosolic oxidized mtDNA to the NLRP3 inflammasome.

Compared to bacteria, the role of fungi within the intestinal microbiota is poorly understood. In this study we investigated whether the presence of a “healthy” fungal community in the gut is important for modulating immune function. Prolonged oral treatment of mice with antifungal drugs resulted in increased disease severity in acute and chronic models of colitis, and also exacerbated the development of allergic airway disease. Microbiota profiling revealed restructuring of fungal and bacterial communities. Specifically, representation of Candida spp. was reduced, while Aspergillus, Wallemia, and Epicoccum spp. were increased. Oral supplementation with a mixture of three fungi found to expand during antifungal treatment (Aspergillus amstelodami, Epicoccum nigrum, and Wallemia sebi) was sufficient to recapitulate the exacerbating effects of antifungal drugs on allergic airway disease. Taken together these results indicate that disruption of commensal fungal populations can influence local and peripheral immune responses and enhance relevant disease states.

Background Kawasaki disease (KD) is the most common cause of acute vasculitis and acquired cardiac disease in US children. Untreated, children may develop coronary artery aneurysms, myocardial infarction and sudden death as a result of the illness. Up to a third of KD patients fail to respond to intravenous gammaglobulin (IVIG), the standard therapy, and alternative treatments are being investigated. Genetic studies have indicated a possible role for IL-1β in KD. We therefore explored the role of IL-1β in a murine model of KD. Methods and Results Using an established mouse model of KD that involves injection of Lactobacillus casei cell wall extract (LCWE), we investigated the role of IL- 1β and caspase-1 (activated by the inflammasome and required for IL-1β maturation) in coronary arteritis, and evaluated the efficacy of IL-1 receptor antagonist (IL-1Ra) as a potential treatment. LCWE-induced IL-1β maturation and secretion was dependent on the NLRP3 inflammasome in macrophages. Both caspase1-deficient and IL-1R-deficient mice were protected from LCWE-induced coronary lesions. Injection of recombinant IL-1β to caspase-1-deficient mice restored the ability of LCWE to cause coronary lesions in response to LCWE. Furthermore, daily injections of the IL-1Ra prevented LCWE-mediated coronary lesions, up to three days after LCWE injection. Conclusions Our results strongly suggest that caspase-1 and IL-1β play critical roles in the development of coronary lesions in this KD mouse model, blocked by IL-1Ra. Therefore, anti-IL-1β treatment strategies may constitute an effective, more targeted treatment of KD to prevent coronary lesions.