Inflammasomes are cytosolic multiprotein complexes that sense microbial infection and trigger cytokine production and cell death. However, the molecular components of inflammasomes, and what they sense, remain poorly defined. Here we demonstrate that 35 amino acids from the Cterminus of flagellin triggered inflammasome activation in the absence of bacterial contaminants or secretion systems. To further elucidate the host flagellin-sensing pathway, we generated mice deficient in Naip5. Naip5-deficient mice failed to activate the inflammasome in response to the 35 amino acids of flagellin or in response to Legionella pneumophila infection. Taken together, these data clarify the molecular basis for the cytosolic response to flagellin.Inflammasomes are cytosolic multiprotein complexes that are critical regulators of inflammation, and are required for proteolytic activation of the cysteine protease caspase-1 (refs. 1-3). Caspase-1 (A000492; http://www.signaling-gateway.org/molecule/query?afcsid=A000492) is itself required for the proteolytic processing and release of inflammatory cytokines such as interleukin 1β (IL-1β) and IL-18, as well as for induction of a necrotic-like cell death called pyroptosis1-3. The molecular components and structures of inflammasomes remain poorly defined. It is believed that multiple distinct inflammasomes may exist, each containing a key scaffold protein of the NLR (nucleotide-binding domain, leucine-rich repeat) superfamily that confers specificity for particular microbial products. For example, NLR proteins of the NLRP1 family (also called NALP1) appear to activate the inflammasome in response to anthrax lethal toxin4 and bacterial muramyl dipeptide5. In contrast, the NLR protein NLRP3 (also called NALP3 or cryopyrin) has been proposed to sense a wide range of stimuli including bacterial RNA6, viral DNA7, uric acid crystals8, muramyl dipeptide9,10, nigericin11, amyloid-beta12, and other irritants13-16. There is at present no molecular explanation for how a single NLR protein can NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author Manuscript be activated by all these microbial products and the precise molecular nature of what is sensed by any inflammasome remains undefined.The inflammasome containing the NLR protein IPAF (also called NLRC4) is one of the best characterized inflammasomes, and has been proposed by several groups to respond to the presence of flagellin in the cytosol17-19. Flagellin-deficient mutants of Salmonella typhimurium and Legionella pneumophila are defective in IPAF-dependent inflammasome activation, and flagellin, purified from or expressed in bacteria, triggers IPAF-dependent caspase-1 activation when delivered to the cytosol of macrophages by use of a pore-forming toxin (listeriolysin O (LLO)) or transfection reagents17-21. It was proposed that during natural infections, flagellin triggers inflammasome activation upon secretion into the host cytosol via bacterial type III/IV secretion systems17-21. However, doubts have been expressed as to w...

Inflammasomes are cytosolic multiprotein complexes that assemble in response to infectious or noxious stimuli and activate the CASPASE-1 protease. The inflammasome containing the nucleotide binding domainleucine-rich repeat (NBD-LRR) protein NLRC4 (interleukin-converting enzyme protease-activating factor [IPAF]) responds to the cytosolic presence of bacterial proteins such as flagellin or the inner rod component of bacterial type III secretion systems (e.g., Salmonella PrgJ). In some instances, such as infection with Legionella pneumophila, the activation of the NLRC4 inflammasome requires the presence of a second NBD-LRR protein, NAIP5. NAIP5 also is required for NLRC4 activation by the minimal C-terminal flagellin peptide, which is sufficient to activate NLRC4. However, NLRC4 activation is not always dependent upon NAIP5. In this report, we define the molecular requirements for NAIP5 in the activation of the NLRC4 inflammasome. We demonstrate that the N terminus of flagellin can relieve the requirement for NAIP5 during the activation of the NLRC4 inflammasome. We also demonstrate that NLRC4 responds to the Salmonella protein PrgJ independently of NAIP5. Our results indicate that NAIP5 regulates the apparent specificity of the NLRC4 inflammasome for distinct bacterial ligands.

BackgroundImpact of liver disease on development of atrial fibrillation (AF) is unclear. The purpose of the study was to evaluate prevalence of AF in the setting of liver disease and whether increasing severity of liver disease, using Model for End‐Stage Liver Disease (MELD), is independently associated with increased risk of AF.Methods and ResultsRetrospective data analysis of 1727 patients with liver disease evaluated for liver transplantation between 2006 and 2015 was performed, and patient characteristics were analyzed from billing codes and review of medical records. Multivariable time‐dependent Cox proportional hazards model was performed to determine effect of increasing MELD score on risk of developing AF. Prevalence of AF was 11.2%. Incidence of AF at median follow‐up time of 1.04 years was 8.5%. Both prevalence and incidence of AF increased with increasing MELD scores. Prevalence of AF was 3.7%, 6.4%, 16.7%, and 20.2% corresponding with MELD quartiles 1 to 10, 11 to 20, 21 to 30, and >30, respectively. Compared with patients with MELD quartile 1 to 10, patients with MELD quartile of 11 to 20 had hazard ratio of 2.73 (confidence interval, 1.47–5.07), those in the MELD quartile of 21 to 30 had a hazard ratio of 5.17 (confidence interval, 2.65–10.09), and those with MELD values >30 had hazard ratio of 9.33 (confidence interval, 3.93–22.14) for development of new‐onset AF. Other significant variables associated with new‐onset AF were age, sleep apnea, valvular heart disease, hemodynamic instability, and reduced left ventricular ejection fraction <50% (hazard ratio, of 1.06, 2.17, 3.21, 2.00, and 2.44, respectively).ConclusionsPrevalence and incidence of AF in patients with liver disease is high. Severity of liver disease, as measured by MELD, is an important predictor of new‐onset AF. This novel finding suggests an interaction between inflammatory and neurohormonal changes in liver disease and pathogenesis of AF.

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