Macrophages activated by the gram negative bacterial product lipopolysaccharide (LPS) switch their core metabolism from oxidative phosphorylation to glycolysis1. Inhibition of glycolysis with 2-deoxyglucose (2DG) suppressed LPS-induced Interleukin-1 beta (IL-1β) but not Tumour necrosis factor alpha (TNFα) in macrophages. A comprehensive metabolic map of LPS-activated macrophages revealed up-regulation of glycolytic and down-regulation of mitochondrial genes, which correlated directly with the expression profiles of altered metabolites. LPS strongly increased the TCA cycle intermediate succinate. Glutamine-dependent anerplerosis was the major source of succinate with the ‘Gamma-Aminobutyric Acid (GABA)-shunt’ pathway also playing a role. LPS-induced succinate stabilized Hypoxia-inducible factor 1α (HIF-1α), an effect inhibited by 2DG, with IL-1β as an important target. LPS also increases succinylation of several proteins. Succinate is therefore identified as a metabolite in innate immune signalling which leads to enhanced IL-1β production during inflammation.

The NLRP3 inflammasome is a component of the inflammatory process and its aberrant activation is pathogenic in inherited disorders such as the cryopyrin associated periodic syndromes (CAPS) and complex diseases such as multiple sclerosis, type 2 diabetes and atherosclerosis. We describe the development of MCC950, a potent, selective, small molecule inhibitor of NLRP3. MCC950 blocks canonical and non-canonical NLRP3 activation at nanomolar concentrations. MCC950 specifically inhibits NLRP3 but not AIM2, NLRC4 or NLRP1 activation. MCC950 reduces Interleukin-1p (IL-1β) production in vivo and attenuates the severity of experimental autoimmune encephalomyelitis (EAE), a disease model of multiple sclerosis. Furthermore, MCC950 treatment rescues neonatal lethality in a mouse model of CAPS and is active in ex vivo samples from individuals with Muckle-Wells syndrome. MCC950 is thus a potential therapeutic for NLRP3-associated syndromes, including autoinflammatory and autoimmune diseases, and a tool for the further study of the NLRP3 inflammasome in human health and disease.

Although microRNA (miRNA) regulation of TLR signaling is well established, this has not yet been observed for NLR proteins or the inflammasomes they form. We have now validated a highly conserved miR-223 target site in the NLRP3 3′-untranslated region. miR-223 expression decreases as monocytes differentiate into macrophages, whereas NLRP3 protein increases during this time. However, overexpression of miR-223 prevents accumulation of NLRP3 protein and inhibits IL-1β production from the inflammasome. Virus inhibition of the inflammasome is an emerging theme, and we have also identified an EBV miRNA that can target the miR-223 binding site in the NLRP3 3′-untranslated region. Furthermore, this virus miRNA can be secreted from infected B cells via exosomes to inhibit the NLRP3 inflammasome in noninfected cells. Therefore, we have identified both the first endogenous miRNA that limits NLRP3 inflammatory capacity during myeloid cell development and also a viral miRNA that takes advantage of this, limiting inflammation for its own purposes.

Abstract. Haneklaus M,

Neudecker et al. define a role for a microRNA, miR-223, in regulating the inflammatory tone of the intestine by constraining nlrp3 inflammasome activation in CCR2+ monocytes and attenuating excessive IL-1β–driven inflammation. Therapeutic nanoparticle delivery of miR-223 mimetics limits experimental colitis.