Cellular Models and Assays to Study NLRP3 Inflammasome Biology

Abstract

The NLRP3 inflammasome is a multi-protein complex that initiates innate immunity responses when exposed to a wide range of stimuli, including pathogen-associated molecular patterns (PAMPs) and danger-associated molecular patterns (DAMPs). Inflammasome activation leads to the release of the pro-inflammatory cytokines interleukin (IL)-1β and IL-18 and to pyroptotic cell death. Over-activation of NLRP3 inflammasome has been associated with several chronic inflammatory diseases. A deep knowledge of NLRP3 inflammasome biology is required to better exploit its potential as therapeutic target and for the development of new selective drugs. To this purpose, in the past few years, several tools have been developed for the biological characterization of the multimeric inflammasome complex, the identification of the upstream signaling cascade leading to inflammasome activation, and the downstream effects triggered by NLRP3 activation. In this review, we will report cellular models and cellular, biochemical, and biophysical assays that are currently available for studying inflammasome biology. A special focus will be on those models/assays that have been used to identify NLRP3 inhibitors and their mechanism of action.

Keywords: NLRP3; NLRP3 inhibitors; biochemical assays; biophysical assays; cell models; inflammasome; read-outs.

Figure 1

Schematic representation of the mechanisms regulating inflammasome activation in canonical, non-canonical and alternative pathway. Abbreviations: PAMP, pathogen-associate molecular pattern; TLRs, toll-like receptors; NF-kB, nuclear factor kappa-light-chain-enhancer of activated B cells; PTM, post-translational modifications; eATP, extracellular adenosine triphosphate; mROS, mitochondrial reactive oxygen species; ASC, apoptosis-associated speck-like protein containing a CARD; IL, interleukin; GSDMD, gasdermin D; iLPS, intracellular LPS; TRIF, TIR-domain-containing adapter-inducing interferon-β; RIPK, receptor-interacting serine/threonine-protein kinase 1; FADD, Fas-associated protein with death domain; NLRP3, NLR family pyrin domain containing 3.