Recent insights into the molecular mechanisms of the NLRP3 inflammasome activation

Inflammasomes are high-molecular-weight protein complexes that are formed in the cytosolic compartment in response to danger- or pathogen-associated molecular patterns. These complexes enable activation of an inflammatory protease caspase-1, leading to a cell death process called pyroptosis and to proteolytic cleavage and release of pro-inflammatory cytokines interleukin (IL)-1beta and IL-18. Along with caspase-1, inflammasome components include an adaptor protein, ASC, and a sensor protein, which triggers the inflammasome assembly in response to a danger signal. The inflammasome sensor proteins are pattern recognition receptors belonging either to the NOD-like receptor (NLR) or to the AIM2-like receptor family. While the molecular agonists that induce inflammasome formation by AIM2 and by several other NLRs have been identified, it is not well understood how the NLR family member NLRP3 is activated. Given that NLRP3 activation is relevant to a range of human pathological conditions, significant attempts are being made to elucidate the molecular mechanism of this process. In this review, we summarize the current knowledge on the molecular events that lead to activation of the NLRP3 inflammasome in response to a range of K (+) efflux-inducing danger signals. We also comment on the reported involvement of cytosolic Ca (2+) fluxes on NLRP3 activation. We outline the recent advances in research on the physiological and pharmacological mechanisms of regulation of NLRP3 responses, and we point to several open questions regarding the current model of NLRP3 activation

Is facet joint distraction a cause of postoperative axial neck pain after ACDF surgery?

Introduction: Intervertebral distraction in anterior cervical discectomy and fusion (ACDF) has been postulated to injure the degenerative facet joints posteriorly and increase postoperative pain and disability. This study aims to determine if there is a correlation between the amount of facet distraction and postoperative patient reported outcomes. Methods: A retrospective cohort analysis of patients undergoing ACDF for degenerative pathologies was performed. Each patient received lateral cervical spine x-rays at the immediate postoperative time point and were split into groups based on the amount of facet distraction measured on these films: Group A: \u3c 1.5 mm; Group B: 1.5-2.0 mm; and Group C: \u3e 2.0 mm. Patients reported outcome measures were obtained preoperatively and at 1-year postoperatively. Univariate and multivariate analyses were performed to compare outcomes between groups. Results: A total of 229 patients were included with an average follow-up of 19.8 [19.0, 20.7] months with a mean facet joint distraction of 1.7mm. There were 87 patients in Group A, 76 patients in Group B, and 66 patients in Group C. Patients significantly improved across all outcome measures from baseline to postoperatively (p \u3c 0.05). There was no difference between groups at any time point with respect to outcome scores (p \u3e 0.05). Multiple regression analysis did not identify increasing distraction as a predictor of patient outcomes. Conclusions: There were no significant differences between patient outcomes and the amount of facet distraction after ACDF surgery. Multivariate analysis did not find a correlation between facet distraction and overall HRQOL outcome

Still no convincing evidence for cognitive map use by honeybees

Cheeseman et al. (1) claim that an ability of honey bees to travel home through a landscape with conflicting information from a celestial compass proves the bees' use of a cognitive map. Their claim involves a curious assumption about the visual information that can be extracted from the terrain: that there is sufficient information for a bee to identify where it is, but insufficient to guide its path without resorting to a cognitive map. We contend that the authors’ claims are unfounded

Stochastic Delay Accelerates Signaling in Gene Networks

The creation of protein from DNA is a dynamic process consisting of numerous reactions, such as transcription, translation and protein folding. Each of these reactions is further comprised of hundreds or thousands of sub-steps that must be completed before a protein is fully mature. Consequently, the time it takes to create a single protein depends on the number of steps in the reaction chain and the nature of each step. One way to account for these reactions in models of gene regulatory networks is to incorporate dynamical delay. However, the stochastic nature of the reactions necessary to produce protein leads to a waiting time that is randomly distributed. Here, we use queueing theory to examine the effects of such distributed delay on the propagation of information through transcriptionally regulated genetic networks. In an analytically tractable model we find that increasing the randomness in protein production delay can increase signaling speed in transcriptional networks. The effect is confirmed in stochastic simulations, and we demonstrate its impact in several common transcriptional motifs. In particular, we show that in feedforward loops signaling time and magnitude are significantly affected by distributed delay. In addition, delay has previously been shown to cause stable oscillations in circuits with negative feedback. We show that the period and the amplitude of the oscillations monotonically decrease as the variability of the delay time increases

Proton Pump Inhibitor Use Affects Pseudarthrosis Rates and Influences Patient-Reported Outcomes.

Study Design: Retrospective cohort review. Objectives: Cervical pseudarthrosis is a frequent cause of need for revision anterior cervical discectomy and fusion (ACDF) and may lead to worse patient-reported outcomes. The effect of proton pump inhibitors on cervical fusion rates are unknown. The purpose of this study was to determine if patients taking PPIs have higher rates of nonunion after ACDF. Methods: A retrospective cohort review was performed to compare patients who were taking PPIs preoperatively with those not taking PPIs prior to ACDF. Patients younger than 18 years of age, those with less than 1-year follow-up, and those undergoing surgery for trauma, tumor, infection, or revision were excluded. The rates of clinically diagnosed pseudarthrosis and radiographic pseudarthrosis were compared between PPI groups. Patient outcomes, pseudarthrosis rates, and revision rates were compared between PPI groups using either multiple linear or logistic regression analysis, controlling for demographic and operative variables. Results: Out of 264 patients, 58 patients were in the PPI group and 206 were in the non-PPI group. A total of 23 (8.71%) patients were clinically diagnosed with pseudarthrosis with a significant difference between PPI and non-PPI groups (P = .009). Using multiple linear regression, PPI use was not found to significantly affect any patient-reported outcome measure. However, based on logistic regression, PPI use was found to increase the odds of clinically diagnosed pseudarthrosis (odds ratio 3.552, P = .014). Additionally, clinically diagnosed pseudarthrosis negatively influenced improvement in PCS-12 scores (P = .022). Conclusions: PPI use was found to be a significant predictor of clinically diagnosed pseudarthrosis following ACDF surgery. Furthermore, clinically diagnosed pseudarthrosis negatively influenced improvement in PCS-12 scores

Does Smoking Status Influence Health-Related Quality of Life Outcome Measures in Patients Undergoing ACDF?

STUDY DESIGN: Retrospective comparative study. OBJECTIVE: Whereas smoking has been shown to affect the fusion rates for patients undergoing an anterior cervical discectomy and fusion (ACDF), the relationship between smoking and health-related quality of life outcome measurements after an ACDF is less clear. The purpose of this study was to evaluate whether smoking negatively affects patient outcomes after an ACDF for cervical degenerative pathology. METHODS: Patients with tumor, trauma, infection, and previous cervical spine surgery and those with less than a year of follow-up were excluded. Smoking status was assessed by self-reported smoking history. Patient outcomes, including Neck Disability Index, Short Form 12 Mental Component Score, Short Form 12 Physical Component Score (PCS-12), Visual Analogue Scale (VAS) arm pain, VAS neck pain, and pseudarthrosis rates were evaluated. Outcomes were compared between smoking groups using multiple linear and logistic regression, controlling for age, sex, and body mass index (BMI), among other factors. A P value \u3c.05 was considered significant. RESULTS: A total of 264 patients were included, with a mean follow-up of 19.8 months, age of 53.1 years, and BMI of 29.6 kg/m2. There were 43 current, 69 former, and 152 nonsmokers in the cohort. At baseline, nonsmokers had higher PCS-12 scores than current smokers (P = .010), lower VAS neck pain than current (P = .035) and former (P = .014) smokers, as well as lower VAS arm pain than former smokers (P = .006). Postoperatively, nonsmokers had higher PCS-12 scores than both current (P = .030) and former smokers (P = .035). Smoking status was not a significant predictor of change in patient outcome in multivariate analysis. CONCLUSIONS: Whereas nonsmokers had higher function and lower pain than former or current smokers preoperatively, smoking status overall was not found to be an independent predictor of outcome scores after ACDF. This supports the notion that smoking status alone should not deter patients from undergoing ACDF for cervical degenerative pathology

Dynamic Assignment and Maintenance of Positional Identity in the Ventral Neural Tube by the Morphogen Sonic Hedgehog

During development of the vertebrate neural tube, cells acquire their positional identity from not only the spatial level of the Sonic Hedgehog signaling gradient, but also the temporal duration

Recent insights into the molecular mechanisms of the NLRP3 inflammasome activation [version 1; referees: 2 approved]

Inflammasomes are high-molecular-weight protein complexes that are formed in the cytosolic compartment in response to danger- or pathogen-associated molecular patterns. These complexes enable activation of an inflammatory protease caspase-1, leading to a cell death process called pyroptosis and to proteolytic cleavage and release of pro-inflammatory cytokines interleukin (IL)-1β and IL-18. Along with caspase-1, inflammasome components include an adaptor protein, ASC, and a sensor protein, which triggers the inflammasome assembly in response to a danger signal. The inflammasome sensor proteins are pattern recognition receptors belonging either to the NOD-like receptor (NLR) or to the AIM2-like receptor family. While the molecular agonists that induce inflammasome formation by AIM2 and by several other NLRs have been identified, it is not well understood how the NLR family member NLRP3 is activated. Given that NLRP3 activation is relevant to a range of human pathological conditions, significant attempts are being made to elucidate the molecular mechanism of this process. In this review, we summarize the current knowledge on the molecular events that lead to activation of the NLRP3 inflammasome in response to a range of K+ efflux-inducing danger signals. We also comment on the reported involvement of cytosolic Ca2+ fluxes on NLRP3 activation. We outline the recent advances in research on the physiological and pharmacological mechanisms of regulation of NLRP3 responses, and we point to several open questions regarding the current model of NLRP3 activation

Transcriptional licensing is required for Pyrin inflammasome activation in human macrophages and bypassed by mutations causing familial Mediterranean fever.

Pyrin is a cytosolic immune sensor that nucleates an inflammasome in response to inhibition of RhoA by bacterial virulence factors, triggering the release of inflammatory cytokines, including IL-1β. Gain-of-function mutations in the MEFV gene encoding Pyrin cause autoinflammatory disorders, such as familial Mediterranean fever (FMF) and Pyrin-associated autoinflammation with neutrophilic dermatosis (PAAND). To precisely define the role of Pyrin in pathogen detection in human immune cells, we compared initiation and regulation of the Pyrin inflammasome response in monocyte-derived macrophages (hMDM). Unlike human monocytes and murine macrophages, we determined that hMDM failed to activate Pyrin in response to known Pyrin activators Clostridioides difficile (C. difficile) toxins A or B (TcdA or TcdB), as well as the bile acid analogue BAA-473. The Pyrin inflammasome response was enabled in hMDM by prolonged priming with either LPS or type I or II interferons and required an increase in Pyrin expression. Notably, FMF mutations lifted the requirement for prolonged priming for Pyrin activation in hMDM, enabling Pyrin activation in the absence of additional inflammatory signals. Unexpectedly, in the absence of a Pyrin response, we found that TcdB activated the NLRP3 inflammasome in hMDM. These data demonstrate that regulation of Pyrin activation in hMDM diverges from monocytes and highlights its dysregulation in FMF

1-Deoxysphingolipids cause autophagosome and lysosome accumulation and trigger NLRP3 inflammasome activation

1-Deoxysphingolipids (deoxySLs) are atypical sphingolipids of clinical relevance as they are elevated in plasma of patients suffering from hereditary sensory and autonomic neuropathy (HSAN1) or type 2 diabetes. Their neurotoxicity is described best but they inflict damage to various cell types by an uncertain pathomechanism. Using mouse embryonic fibroblasts and an alkyne analog of 1-deoxysphinganine (doxSA), the metabolic precursor of all deoxySLs, we here study the impact of deoxySLs on macroautophagy/autophagy, the regulated degradation of dysfunctional or expendable cellular components. We find that deoxySLs induce autophagosome and lysosome accumulation indicative of an increase in autophagic flux. The autophagosomal machinery targets damaged mitochondria that have accumulated N-acylated doxSA metabolites, presumably deoxyceramide and deoxydihydroceramide, and show aberrant swelling and tubule formation. Autophagosomes and lysosomes also interact with cellular lipid aggregates and crystals that occur upon cellular uptake and N-acylation of monomeric doxSA. As crystals entering the lysophagosomal apparatus in phagocytes are known to trigger the NLRP3 inflammasome, we also treated macrophages with doxSA. We demonstrate the activation of the NLRP3 inflammasome by doxSLs, prompting the release of IL1B from primary macrophages. Taken together, our data establish an impact of doxSLs on autophagy and link doxSL pathophysiology to inflammation and the innate immune system.Abbreviations: alkyne-doxSA: (2S,3R)-2-aminooctadec-17yn-3-ol; alkyne-SA: (2S,3R)-2- aminooctadec-17yn-1,3-diol; aSA: alkyne-sphinganine; ASTM-BODIPY: azido-sulfo-tetramethyl-BODIPY; CerS: ceramide synthase; CMR: clonal macrophage reporter; deoxySLs: 1-deoxysphingolipids; dox(DH)Cer: 1-deoxydihydroceramide; doxCer: 1-deoxyceramide; doxSA: 1-deoxysphinganine; FB1: fumonisin B1; HSAN1: hereditary sensory and autonomic neuropathy type 1; LC3: MAP1LC3A and MAP1LC3B; LPS: lipopolysaccharide; MEF: mouse embryonal fibroblasts; MS: mass spectrometry; N3635P: azido-STAR635P; N3Cy3: azido-cyanine 3; N3picCy3: azido-picolylcyanine 3; NLRP3: NOD-like receptor pyrin domain containing protein 3; P4HB: prolyl 4-hydroxylase subunit beta; PINK1: PTEN induced putative kinase 1; PYCARD/ASC: PYD and CARD domain containing; SPTLC1: serine palmitoyltransferase long chain base subunit 1; SQSTM1: sequestosome 1; TLC: thin layer chromatography