ATP‐dependent activation of an inflammasome in primary gingival epithelial cells infected by Porphyromonas gingivalis

Production of IL‐1β typically requires two‐separate signals. The first signal, from a pathogen‐associated molecular pattern, promotes intracellular production of immature cytokine. The second signal, derived from a danger signal such as extracellular ATP, results in assembly of an inflammasome, activation of caspase‐1 and secretion of mature cytokine. The inflammasome component, Nalp3, plays a non‐redundant role in caspase‐1 activation in response to ATP binding to P2X₇ in macrophages. Gingival epithelial cells (GECs) are an important component of the innate‐immune response to periodontal bacteria. We had shown that GECs express a functional P2X₇ receptor, but the ability of GECs to secrete IL‐1β during infection remained unknown. We find that GECs express a functional Nalp3 inflammasome. Treatment of GECs with LPS or infection with the periodontal pathogen, Porphyromonas gingivalis, induced expression of the il‐1β gene and intracellular accumulation of IL‐1β protein. However, IL‐1β was not secreted unless LPS‐treated or infected cells were subsequently stimulated with ATP. Conversely, caspase‐1 is activated in GECs following ATP treatment but not P. gingivalis infection. Furthermore, depletion of Nalp3 by siRNA abrogated the ability of ATP to induce IL‐1β secretion in infected cells. The Nalp3 inflammasome is therefore likely to be an important mediator of the inflammatory response in gingival epithelium.     

Caspase‐1 inflammasome activity in patients with Staphylococcus aureus bacteremia

The inflammasome is a multiprotein complex that mediates caspase‐1 activation with subsequent maturation of the proinflammatory cytokines IL‐1β and IL‐18. The NLRP3 inflammasome is known to be activated by Staphylococcus aureus, one of the leading causes of bacteremia worldwide. Inflammasome activation and regulation in response to bacterial infection have been found to be of importance for a balanced host immune response. However, inflammasome signaling in vivo in humans initiated by S. aureus is currently sparsely studied. This study therefore aimed to investigate NLRP3 inflammasome activity in 20 patients with S. aureus bacteremia (SAB), by repeated measurement during the first week of bacteremia, compared with controls. Caspase‐1 activity was measured in monocytes and neutrophils by flow cytometry detecting FLICA (fluorescent‐labeled inhibitor of caspase‐1), while IL‐1β and IL‐18 was measured by Luminex and ELISA, respectively. As a measure of inflammasome priming, messenger RNA (mRNA) expression of NLRP3, CASP1 (procaspase‐1), and IL1B (pro‐IL‐1β) was analyzed by quantitative PCR. We found induced caspase‐1 activity in innate immune cells with subsequent release of IL‐18 in patients during the acute phase of bacteremia, indicating activation of the inflammasome. There was substantial interindividual variation in caspase‐1 activity between patients with SAB. We also found an altered inflammasome priming with low mRNA levels of NLRP3 accompanied by elevated mRNA levels of IL1B. This increased knowledge of the individual host immune response in SAB could provide support in the effort to optimize management and treatment of each individual patient.     

Activation of the NLRP3 inflammasome in microglia: the role of ceramide

Inflammation within the CNS is a major component of many neurodegenerative diseases. A characteristic feature is the generation of microglia‐derived factors that play an essential role in the immune response. IL‐1β is a pro‐inflammatory cytokine released by activated microglia, able to exacerbate injury at elevated levels. In the presence of caspase‐1, pro‐IL‐1β is cleaved to the mature cytokine following NOD‐like receptor pyrin domain containing 3 (NLRP3) inflammasome activation. Growing evidence suggests that ceramide plays a critical role in NLRP3 inflammasome assembly, however, the relationship between ceramide and inflammasome activation in microglia remains unknown. Here, we investigated potential mechanistic links between ceramide as a modulator of NLRP3 inflammasome assembly and the resulting secretion of IL‐1β using small bioactive enzyme stimulators and inhibitors of ceramide signaling in wild‐type and apoptosis‐associated speck‐like protein containing a CARD knockout (ASC^?/?) primary microglia. To induce the expression of inflammasome components, microglia were primed prior to experiments. Treatment with sodium palmitate (PA) induced de novo ceramide synthesis via modulation of its synthesizing protein serine palmitoyl transferase resulting in increased IL‐1β secretion in microglia. Exposure of microglia to the serine palmitoyl transferase‐inhibitor l ‐cycloserine significantly prevented PA‐induced IL‐1β secretion. Application of the ceramide analogue C2 and the sphingosine‐1‐phosphate‐receptor agonist Fingolimod (FTY720) up‐regulated levels of IL‐1β and cleaved caspase‐1 in wild‐type microglia, whereas ASC^?/? microglia were unaffected. HPA‐12 inhibition of ceramide transport did not affect inflammasome activation. Taken together, our findings reveal a critical role for ceramide as a positive modulator of NLRP3 inflammasome assembly and the resulting release of IL‐1β.

     

Critical role for the AIM2 inflammasome during acute CNS bacterial infection

Interleukin‐1β (IL‐1β) is essential for eliciting protective immunity during the acute phase of Staphylococcus aureus (S. aureus) infection in the central nervous system (CNS). We previously demonstrated that microglial IL‐1β production in response to live S. aureus is mediated through the Nod‐like receptor protein 3 (NLRP3) inflammasome, including the adapter protein ASC (apoptosis‐associated speck‐like protein containing a caspase‐1 recruitment domain), and pro‐caspase 1. Here, we utilized NLRP3, ASC, and caspase 1/11 knockout (KO) mice to demonstrate the functional significance of inflammasome activity during CNS S. aureus infection. ASC and caspase 1/11 KO animals were exquisitely sensitive, with approximately 50% of mice succumbing to infection within 24 h. Unexpectedly, the survival of NLRP3 KO mice was similar to wild‐type animals, suggesting the involvement of an alternative upstream sensor, which was later identified as absent in melanoma 2 (AIM2) based on the similar disease patterns between AIM2 and ASC KO mice. Besides IL‐1β, other key inflammatory mediators, including IL‐6, CXCL1, CXCL10, and CCL2 were significantly reduced in the CNS of AIM2 and ASC KO mice, implicating autocrine/paracrine actions of IL‐1β, as these mediators do not require inflammasome processing for secretion. These studies demonstrate a novel role for the AIM2 inflammasome as a critical molecular platform for regulating IL‐1β release and survival during acute CNS S. aureus infection.

     

Fusobacterium nucleatum infection of gingival epithelial cells leads to NLRP3 inflammasome‐dependent secretion of IL‐1β and the danger signals ASC and HMGB1

Fusobacterium nucleatum is an invasive anaerobic bacterium that is associated with periodontal disease. Previous studies have focused on virulence factors produced by F. nucleatum, but early recognition of the pathogen by the immune system remains poorly understood. Although an inflammasome in gingival epithelial cells (GECs) can be stimulated by danger‐associated molecular patterns (DAMPs) (also known as danger signals) such as ATP, inflammasome activation by this periodontal pathogen has yet to be described in these cells. This study therefore examines the effects of F. nucleatum infection on pro‐inflammatory cytokine expression and inflammasome activation in GECs. Our results indicate that infection induces translocation of NF‐κB into the nucleus, resulting in cytokine gene expression. In addition, infection activates the NLRP3 inflammasome, which in turn activates caspase‐1 and stimulates secretion of mature IL‐1β. Unlike other pathogens studied until now, F. nucleatum activates the inflammasome in GECs in the absence of exogenous DAMPs such as ATP. Finally, infection promotes release of other DAMPs that mediate inflammation, such as high‐mobility group box 1 protein and apoptosis‐associated speck‐like protein, with a similar time‐course as caspase‐1 activation. Thus, F. nucleatum expresses the pathogen‐associated molecular patterns necessary to activate NF‐κB and also provides an endogenous DAMP to stimulate the inflammasome and further amplify inflammation through secretion of secondary DAMPs.     

Activation of nucleotide‐binding domain‐like receptor containing protein 3 inflammasome in dendritic cells and macrophages by Streptococcus sanguinis

Streptococcus sanguinis is frequently isolated from the blood of patients with infective endocarditis and contributes to the pathology of this disease through induction of interleukin (IL)‐1β responsible for the development of the disease. However, the mechanism of IL‐1β induction remains unknown. In this study, S. sanguinis activated a murine dendritic cell (DC) to induce IL‐1β and this activity was attenuated by silencing the mRNAs of nucleotide‐binding domain‐like receptor containing protein 3 (NLRP3) and caspase‐1. S. sanguinis induced IL‐1β production in murine bone marrow‐derived macrophage, but this activity was significantly reduced in bone marrow‐derived macrophages from NLRP3‐, apoptosis‐associated speck‐like protein containing a caspase‐recruitment domain‐, and caspase‐1‐deficient mice. DC phagocytosed S. sanguinis cells, followed by the release of adenosine triphosphate (ATP). The ATP‐degradating enzyme attenuated the release of ATP and IL‐1β. The inhibitors for ATP receptor reduced IL‐1β release in DC. These results strongly suggest that S. sanguinis has the activity to induce IL‐1β through the NLRP3 inflammasome in macrophage and DC and interaction of purinergic receptors with ATP released is involved in expression of the activity.     

Soluble uric acid induces myocardial damage through activating the NLRP3 inflammasome

Uric acid crystal is known to activate the NLRP3 inflammasome and to cause tissue damages, which can result in many diseases, such as gout, chronic renal injury and myocardial damage. Meanwhile, soluble uric acid (sUA), before forming crystals, is also related to these diseases. This study was carried out to investigate whether sUA could also activate NLRP3 inflammasome in cardiomyocytes and to analyse the mechanisms. The cardiomyocyte activity was monitored, along with the levels of mature IL‐1β and caspase‐1 from H9c2 cells following sUA stimulus. We found that sUA was able to activate NLRP3 inflammasome, which was responsible for H9c2 cell apoptosis induced by sUA. By elevating TLR6 levels and then activating NF‐κB/p65 signal pathway, sUA promoted NLRP3, pro‐caspase 1 and pro‐IL‐1β production and provided the first signal of NLRP3 inflammasome activation. Meanwhile, ROS production regulated by UCP2 levels also contributed to NLRP3 inflammasome assembly and subsequent caspase 1 activation and mature IL‐1β secretion. In addition, the tlr6 knockdown rats suffering from hyperuricemia showed the lower level of IL‐1β and an ameliorative cardiac function. These findings suggest that sUA activates NLRP3 inflammasome in cardiomyocytes and they may provide one therapeutic strategy for myocardial damage induced by sUA.     

Involvement of caspase‐1 in inflammasomes activation and bacterial clearance in S. aureus‐infected osteoblast‐like MG‐63 cells

Staphylococcus aureus, a versatile Gram‐positive bacterium, is the main cause of bone and joint infections (BJI), which are prone to recurrence. The inflammasome is an immune signaling platform that assembles after pathogen recognition. It activates proteases, most notably caspase‐1 that proteolytically matures and promotes the secretion of mature IL‐1β and IL‐18. The role of inflammasomes and caspase‐1 in the secretion of mature IL‐1β and in the defence of S. aureus‐infected osteoblasts has not yet been fully investigated. We show here that S. aureus‐infected osteoblast‐like MG‐63 but not caspase‐1 knock‐out CASP1 ^?/?MG‐63 cells, which were generated using CRISPR‐Cas9 technology, activate the inflammasome as monitored by the release of mature IL‐1β. The effect was strain‐dependent. The use of S. aureus deletion and complemented phenole soluble modulins (PSMs) mutants demonstrated a key role of PSMs in inflammasomes‐related IL‐1β production. Furthermore, we found that the lack of caspase‐1 in CASP1 ^?/?MG‐63 cells impairs their defense functions, as bacterial clearance was drastically decreased in CASP1 ^?/? MG‐63 compared to wild‐type cells. Our results demonstrate that osteoblast‐like MG‐63 cells play an important role in the immune response against S. aureus infection through inflammasomes activation and establish a crucial role of caspase‐1 in bacterial clearance.     

Inflammasome activation by Pseudomonas aeruginosa's ExlA pore‐forming toxin is detrimental for the host

During acute Pseudomonas aeruginosa infection, the inflammatory response is essential for bacterial clearance. Neutrophil recruitment can be initiated following the assembly of an inflammasome within sentinel macrophages, leading to activation of caspase‐1, which in turn triggers macrophage pyroptosis and IL‐1β/IL‐18 maturation. Inflammasome formation can be induced by a number of bacterial determinants, including Type III secretion systems (T3SSs) or pore‐forming toxins, or, alternatively, by lipopolysaccharide (LPS) via caspase‐11 activation. Surprisingly, previous studies indicated that a T3SS‐induced inflammasome increased pathogenicity in mouse models of P. aeruginosa infection. Here, we investigated the immune reaction of mice infected with a T3SS‐negative P. aeruginosa strain (IHMA879472). Virulence of this strain relies on ExlA, a secreted pore‐forming toxin. IHMA879472 promoted massive neutrophil infiltration in infected lungs, owing to efficient priming of toll‐like receptors, and thus enhanced the expression of inflammatory proteins including pro‐IL‐1β and TNF‐α. However, mature‐IL‐1β and IL‐18 were undetectable in wild‐type mice, suggesting that ExlA failed to effectively activate caspase‐1. Nevertheless, caspase‐1/11 deficiency improved survival following infection with IHMA879472, as previously described for T3SS+ bacteria. We conclude that the detrimental effect associated with the ExlA‐induced inflammasome is probably not due to hyperinflammation, rather it stems from another inflammasome‐dependent process.     

P2X7 receptor inhibition attenuated sympathetic nerve sprouting after myocardial infarction via the NLRP3/IL‐1β pathway

Mounting evidence supports the hypothesis that inflammation modulates sympathetic sprouting after myocardial infarction (MI). The myeloid P2X₇ signal has been shown to activate the nucleotide‐binding and oligomerization domain‐like receptor family pyrin domain‐containing 3 (NLRP3) inflammasome, a master regulator of inflammation. We investigated whether P2X₇ signal participated in the pathogenesis of sympathetic reinnervation after MI, and whether NLRP3/interleukin‐1β (IL‐1β) axis is involved in the process. We explored the relationship between P2X₇ receptor (P2X₇R) and IL‐1β in the heart tissue of lipopolysaccharide (LPS)‐primed naive rats. 3′‐O‐(4‐benzoyl) benzoyl adenosine 5′‐triphosphate (BzATP), a P2X₇R agonist, induced caspase‐1 activation and mature IL‐1β release, which was further neutralized by a NLRP3 inhibitor (16673‐34‐0). MI was induced by coronary artery ligation. Following infarction, a marked increase in P2X₇R was localized within infiltrated macrophages and observed in parallel with an up‐regulation of NLRP3 inflammasome levels and the release of IL‐1β in the left ventricle. The administration of A‐740003 (a P2X₇R antagonist) significantly prevented the NLRP3/IL‐1β increase. A‐740003 and/or Anakinra (an IL‐1 receptor antagonist) significantly reduced macrophage infiltration as well as macrophage‐based IL‐1β and NGF (nerve growth factor) production and eventually blunted sympathetic hyperinnervation, as assessed by the immunofluorescence of tyrosine hydroxylase (TH) and growth‐associated protein 43 (GAP 43). Moreover, the use of Anakinra partly attenuated sympathetic sprouting. This indicated that the effect of P2X₇ on neural remodelling was mediated at least partially by IL‐1β. The arrhythmia score of programmed electric stimulation was in accordance with the degree of sympathetic hyperinnervation. In vitro studies showed that BzATP up‐regulated secretion of nerve growth factor (NGF) in M1 macrophages via IL‐1β. Together, these data indicate that P2X₇R contributes to neural and cardiac remodelling, at least partly mediated by NLRP3/IL‐1β axis. Therapeutic interventions targeting P2X₇ signal may be a novel approach to ameliorate arrhythmia following MI.     

H3 relaxin inhibits the collagen synthesis via ROS‐ and P2X7R‐mediated NLRP3 inflammasome activation in cardiac fibroblasts under high glucose

Excessive production of reactive oxygen species (ROS) and P2X7R activation induced by high glucose increases NLRP3 inflammasome activation, which contributes to the pathogenesis of diabetic cardiomyopathy. Although H3 relaxin has been shown to inhibit cardiac fibrosis induced by isoproterenol, the mechanism has not been well studied. Here, we demonstrated that high glucose (HG) induced the collagen synthesis by activation of the NLRP3 inflammasome, leading to caspase‐1 activation, interleukin‐1β (IL‐1β) and IL‐18 secretion in neonatal rat cardiac fibroblasts. Moreover, we used a high‐glucose model with neonatal rat cardiac fibroblasts and showed that the activation of ROS and P2X7R was augmented and that ROS‐ and P2X7R‐mediated NLRP3 inflammasome activation was critical for the collagen synthesis. Inhibition of ROS and P2X7R decreased NLRP3 inflammasome‐mediated collagen synthesis, similar to the effects of H3 relaxin. Furthermore, H3 relaxin reduced the collagen synthesis via ROS‐ and P2X7R‐mediated NLRP3 inflammasome activation in response to HG. These results provide a mechanism by which H3 relaxin alleviates NLRP3 inflammasome‐mediated collagen synthesis through the inhibition of ROS and P2X7R under HG conditions and suggest that H3 relaxin represents a potential drug for alleviating cardiac fibrosis in diabetic cardiomyopathy.     

Structure of the NLRP1 caspase recruitment domain suggests potential mechanisms for its association with procaspase‐1

The NLRP1 inflammasome responds to microbial challenges such as Bacillus anthracis infection and is implicated in autoimmune disease such as vitiligo. Human NLRP1 contains both an N‐terminal pyrin domain (PYD) and a C‐terminal caspase recruitment domain (CARD), with the latter being essential for its association with the downstream effector procaspase‐1. Here we report a 2.0 Å crystal structure of the human NLRP1 CARD as a fusion with the maltose‐binding protein. The structure reveals the six‐helix bundle fold of the NLRP1 CARD, typical of the death domain superfamily. The charge surface of the NLRP1 CARD structure and a procaspase‐1 CARD model suggests potential mechanisms for their association through electrostatic attraction. Proteins 2013; 81:1266–1270. © 2013 Wiley Periodicals, Inc.     

Gypenosides improve diabetic cardiomyopathy by inhibiting ROS‐mediated NLRP3 inflammasome activation

NLRP3 inflammasome activation plays an important role in diabetic cardiomyopathy (DCM), which may relate to excessive production of reactive oxygen species (ROS). Gypenosides (Gps), the major ingredients of Gynostemma pentaphylla (Thunb.) Makino, have exerted the properties of anti‐hyperglycaemia and anti‐inflammation, but whether Gps improve myocardial damage and the mechanism remains unclear. Here, we found that high glucose (HG) induced myocardial damage by activating the NLRP3 inflammasome and then promoting IL‐1β and IL‐18 secretion in H9C2 cells and NRVMs. Meanwhile, HG elevated the production of ROS, which was vital to NLRP3 inflammasome activation. Moreover, the ROS activated the NLRP3 inflammasome mainly by cytochrome c influx into the cytoplasm and binding to NLRP3. Inhibition of ROS and cytochrome c dramatically down‐regulated NLRP3 inflammasome activation and improved the cardiomyocyte damage induced by HG, which was also detected in cells treated by Gps. Furthermore, Gps also reduced the levels of the C‐reactive proteins (CRPs), IL‐1β and IL‐18, inhibited NLRP3 inflammasome activation and consequently improved myocardial damage in vivo. These findings provide a mechanism that ROS induced by HG activates the NLRP3 inflammasome by cytochrome c binding to NLRP3 and that Gps may be potential and effective drugs for DCM via the inhibition of ROS‐mediated NLRP3 inflammasome activation.     

Naoxintong attenuates Ischaemia/reperfusion Injury through inhibiting NLRP3 inflammasome activation

Naoxintong (NXT) is a Chinese Materia Medica standardized product extracted from 16 various kinds of Chinese traditional herbal medicines including Salvia miltiorrhiza, Angelica sinensis, Astragali Radix. Naoxintong is clinically effective in treating ischaemia heart disease. Nucleotide‐binding oligomerization domain‐Like Receptor with a Pyrin domain 3 (NLRP3) inflammasome has been critically involved in myocardial ischaemia/reperfusion (I/R) injury. Here, we have been suggested that NXT might attenuate myocardial I/R injury via suppression of NLRP3 inflammasome activation. Male C57BL6 mice were subjected to myocardial I/R injury via 45 min. coronary ligation and release for the indicated times. Naoxintong (0.7 g/kg/day) and PBS were orally administrated for 2 weeks before surgery. Cardiac function assessed by echocardiography was significantly improved in the NXT group compared to PBS group at day 2 after myocardial I/R. NLRP3 inflammasome activation is crucially involved in the initial inflammatory response after myocardial I/R injury, leading to cleaved caspase‐1, mature interleukin (IL)‐1β production, accompanying by macrophage and neutrophil infiltration. The cardioprotective effect of NXT was associated with a diminished NLRP3 inflammasome activation, decreased pro‐inflammatory macrophage (M1 macrophages) and neutrophil infiltration after myocardial I/R injury. In addition, serum levels of IL‐1β, indicators of NLRP3 inflammasome activation, were also significantly suppressed in the NXT treated group after I/R injury. Naoxintong exerts cardioprotive effects at least partly by suppression of NLRP3 inflammasome activation in this I/R injury model.     

Vascular endothelial cells senescence is associated with NOD-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome activation via reactive oxygen species (ROS)/thioredoxin-interacting protein (TXNIP) pathway

Endothelial dysfunction caused by endothelial cells senescence and chronic inflammation is tightly linked to the development of cardiovascular diseases. NLRP3 (NOD-like receptor family pyrin domain-containing3) inflammasome plays a central role in inflammatory response that is associated with diverse inflammatory diseases. This study explores the effects and possible mechanisms of NLRP3 inflammasome in endothelial cells senescence. Results show an increment of pro-inflammatory cytokine interleukin (IL) −1β secretion and caspase-1 activation during the senescence of endothelial cells induced by bleomycin. Moreover, secreted IL-1β promoted endothelial cells senescence through up-regulation of p53/p21 protein expression. NLRP3 inflammasome was found to mediate IL-1β secretion through the production of ROS (reactive oxygen species) during the senescence of endothelial cells. Furthermore, the association of TXNIP (thioredoxin-interacting protein) with NLRP3 induced by ROS promoted NLRP3 inflammasome activation in senescent endothelial cells. In addition, the expressions of NLRP3 inflammasome related genes, ASC (apoptosis associated speck-like protein containing a CARD), TXNIP, cleaved caspase-1 and IL-1β, were also increased in vitro and in vivo studies. These findings indicate that endothelial senescence could be mediated through ROS and NLRP3 inflammasome signaling pathways, suggesting a potential target for the prevention of endothelial senescence-related cardiovascular diseases.     

Serum amyloid A activates the NLRP3 inflammasome via P2X7 receptor and a cathepsin B-sensitive pathway

Serum amyloid A (SAA) is an acute-phase protein, the serum levels of which can increase up to 1000-fold during inflammation. SAA has a pathogenic role in amyloid A-type amyloidosis, and increased serum levels of SAA correlate with the risk for cardiovascular diseases. IL-1β is a key proinflammatory cytokine, and its secretion is strictly controlled by the inflammasomes. We studied the role of SAA in the regulation of IL-1β production and activation of the inflammasome cascade in human and mouse macrophages, as well as in THP-1 cells. SAA could provide a signal for the induction of pro-IL-1β expression and for inflammasome activation, resulting in secretion of mature IL-1β. Blocking TLR2 and TLR4 attenuated SAA-induced expression of IL1B, whereas inhibition of caspase-1 and the ATP receptor P2X(7) abrogated the release of mature IL-1β. NLRP3 inflammasome consists of the NLRP3 receptor and the adaptor protein apoptosis-associated speck-like protein containing CARD (a caspase-recruitment domain) (ASC). SAA-mediated IL-1β secretion was markedly reduced in ASC(-/-) macrophages, and silencing NLRP3 decreased IL-1β secretion, confirming NLRP3 as the SAA-responsive inflammasome. Inflammasome activation was dependent on cathepsin B activity, but it was not associated with lysosomal destabilization. SAA also induced secretion of cathepsin B and ASC. In conclusion, SAA can induce the expression of pro-IL-1β and activation of the NLRP3 inflammasome via P2X(7) receptor and a cathepsin B-sensitive pathway. Thus, during systemic inflammation, SAA may promote the production of IL-1β in tissues. Furthermore, the SAA-induced secretion of active cathepsin B may lead to extracellular processing of SAA and, thus, potentially to the development of amyloid A amyloidosis.     

Hepatoprotective effect of allicin against acetaminophen‐induced liver injury: Role of inflammasome pathway,apoptosis, and liver regeneration

Acetaminophen (APAP) overdose leads to liver injury. NLRP3 inflammasome is a key player in APAP‐induced inflammation. Also, apoptosis and liver regeneration play an important role in liver injury. Therefore, we assessed allicin's protective effect on APAP‐induced hepatotoxicity and studied its effect on NLRP3 inflammasome and apoptosis. Mice in the APAP group were injected by APAP (250 mg/kg, intraperitoneal). The allicin‐treated group received allicin orally (10 mg/kg/d) during 7 days before APAP injection. Serum and hepatic tissues were separated 24 hours after APAP injection. Serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), albumin, alkaline phosphatase (ALP), and hepatic malondialdehyde (MDA) were assessed using the colorimetric method. Hepatic NLRP3 inflammasome, caspase‐1, and interleukin‐1β (IL‐1β) were estimated using enzyme‐linked immunosorbent assay. Hepatic Bcl‐2 and Ki‐67 were investigated by immunohistochemistry. APAP significantly increased AST, ALT, and ALP, whereas allicin significantly decreased their levels. Also, APAP significantly decreased albumin and allicin significantly improved it. APAP produced changes in liver morphology, including inflammation and massive coagulative necrosis. Allicin protected the liver from APAP‐induced necrosis, apoptosis, and hepatocellular degeneration via increasing Bcl‐2 and Ki‐67 levels. APAP significantly increased the hepatic MDA, whereas allicin significantly prevented this increase. APAP markedly activated the NLRP3 inflammasome pathway and consequently increased the production of caspase‐1 and IL‐1β. Interestingly, we found that allicin significantly inhibited NLRP3 inflammasome activation, which resulted in decreased caspase‐1 and IL‐1β levels. Allicin has a hepatoprotective effect against APAP‐induced liver injury via the decline of oxidative stress and inhibition of the inflammasome pathway and apoptosis. Therefore, allicin might be a novel tool to halt the progression of APAP‐stimulated hepatotoxicity.     

Mechanisms of NLRP3 Inflammasome Activation: Its Role in the Treatment of Alzheimer’s Disease

Alzheimer’s disease (AD) is a common neurodegenerative disease of progressive dementia which is characterized pathologically by extracellular neuritic plaques containing aggregated amyloid beta (Aβ) and intracellular hyperphosphorylated tau protein tangles in cerebrum. It has been confirmed that microglia-specific nucleotide-binding oligomerization domain (NOD)-like receptor protein 3 (NLRP3) inflammasome-mediated chronic neuroinflammation plays a crucial role in the pathogenesis of AD. Stimulated by Aβ deposition, NLRP3 assembles and activates within microglia in the AD brain, leading to caspase-1 activation along with downstream interleukin (IL)-1β secretion, and subsequent inflammatory events. Activation of the NLRP3 inflammasome mediates microglia to exhibit inflammatory M1 phenotype, with high expression of caspase-1 and IL-1β. This leads to Aβ deposition and neuronal loss in the amyloid precursor protein (APP)/human presenilin-1 (PS1) mouse model of AD. However, NLRP3 or caspase-1 deletion in APP/PS1 mice promotes microglia to transform to an anti-inflammatory M2 phenotype, with decreased secretion of caspase-1 and IL-1β. It also results in improved cognition, enhanced Aβ clearance, and a lower cerebral inflammatory response. This result suggests that the NLRP3 inflammasome may be an appropriate target for reducing neuroinflammation and alleviating pathological processes in AD. In the present review, we summarize the generally accepted regulatory mechanisms of NLRP3 inflammasome activation, and explore its role in neuroinflammation. Furthermore, we speculate on the possible roles of microglia-specific NLRP3 activation in AD pathogenesis and consider potential therapeutic interventions targeting the NLRP3 inflammasome in AD.

     

Cytolethal distending toxin‐induced release of interleukin‐1β by human macrophages is dependent upon activation of glycogen synthase kinase 3β, spleen tyrosine kinase (Syk) and the noncanonical inflammasome

Cytolethal distending toxins (Cdt) are a family of toxins produced by several human pathogens which infect mucocutaneous tissue and induce inflammatory disease. We have previously demonstrated that the Aggregatibacter actinomycetemcomitans Cdt induces a pro‐inflammatory response from human macrophages which involves activation of the NLRP3 inflammasome. We now demonstrate that in addition to activating caspase‐1 (canonical inflammasome), Cdt treatment leads to caspase‐4 activation and involvement of the noncanonical inflammasome. Cdt‐treated cells exhibit pyroptosis characterised by cleavage of gasdermin‐D (GSDMD), release of HMGB1 at 24 hr and LDH at 48 hr. Inhibition of either the canonical (caspase‐1) or noncanonical (caspase‐4) inflammasome blocks both Cdt‐induced release of IL‐1β and induction of pyroptosis. Analysis of upstream events indicates that Cdt induces Syk phosphorylation (activation); furthermore, blockade of Syk expression and inhibition of pSyk activity inhibit both Cdt‐induced cytokine release and pyroptosis. Finally, we demonstrate that increases in pSyk are dependent upon Cdt‐induced activation of GSK3β. These studies advance our understanding of Cdt function and provide new insight into the virulence potential of Cdt in mediating the pathogenesis of disease caused by Cdt‐producing organisms such as A. actinomycetemcomitans.