Induced Caspase-1 Activation Research Articles

Abstract T MP71: Estrogen Attenuates NLRP3 Inflammasome Activation Following Global Cerebral Ischemia

NLRP3 inflammasome formation and activation have recently been reported to be involved in several types of ischemic injury, including middle cerebral artery occlusion (MCAO)-induced cerebral ischemia. However, it is unknown whether or not NLRP3 inflammasome is involved in brain injury following global cerebral ischemia (GCI) in ovariectomized rats, and whether its activation can be regulated by 17β-Estradiol (E2) replacement. As part of its activation, NLRP3 can bind the apoptosis speck protein (ASC) to assemble an NLRP3-ASC-Caspase-1 inflammasome and induce caspase-1 activation, followed by subsequent activation of IL-1β and IL-18. IL-1β and IL-18 are implicated in neurodegeneration in diverse forms of experimental brain injury, including ischemic brain injury. In this study, confocal microscopic analysis revealed that NLRP3 expression was initially highly induced in hippocampal CA1 neurons at 1-day reperfusion in the placebo-treated rats following 10-min GCI. Interestingly, at later time-points (3-7 days reperfusion) NLRP3 became highly expressed in GFAP-positive astrocytes in the hippocampal CA1 region. Additionally, ASC was similarly highly induced in astrocytes and co-localized with NLRP3 at the late reperfusion stage (3-7 day). E2 replacement initiated at the time of ovariectomy (7 days before GCI) blocked the induction of NLRP3 and ASC in astrocytes. In addition, caspase-1 activity was found to be highly induced at 1-day reperfusion, and significantly attenuated following E2 replacement. Cleaved-IL-1β expression was also enhanced in neurons at 1-day reperfusion and in astrocytes at 3-7 day reperfusion - effects that were blocked by E2 replacement. Taken as a whole, the results suggest that NLRP3 inflammasome formation and activation is markedly enhanced in the hippocampal CA1 region following GCI, which may contribute to neuronal injury. Furthermore, E2 replacement markedly attenuated NLRP3 inflammation activation and cytokine induction after GCI, which may contribute to E2 neuroprotective effects following GCI.

Role of NLRP3 Inflammasomes in Hepatic Ischemia-reperfusion Injury

Increasing evidence indicates that inflammation plays an important role in the pathogenesis of hepatic ischemia-reperfusion (I/R) injury. However, it is still unclear how I/R stimuli induce inflammatory responses in the liver. NLRP3 is an intracellular pattern recognition receptor and a component of NLRP3 inflammasomes that can induce caspase-1 activation, and regulate the processing of a potent inflammatory cytokine, interleukin-1β (IL-1β). Several investigations have recently suggested that inflammatory responses are mediated through NLRP3 inflammasomes in hepatic I/R injury. On the other hand, we recently found that NLRP3 regulates neutrophil functions and contributes to hepatic I/R injury independently of inflammasomes. This review summarizes the basic information on NLRP3 inflammasomes and discusses inflammasome-dependent/independent roles of NLRP3 in the pathophysiology of hepatic I/R injury. Rec.12/21/2014, Acc.1/22/2015, pp61-68

Independent roles of the priming and the triggering of the NLRP3 inflammasome in the heart.

The NLRP3 inflammasome is activated in the ischaemic heart promoting caspase-1 activation, inflammation, and cell death. Ischaemic injury establishes both a priming signal (transcription of inflammasome components) and a trigger (NLRP3 activation). Whether NLRP3 activation, without priming, induces cardiac dysfunction and/or failure is unknown. The aim of this study was to assess the independent and complementary roles of the priming and the triggering signals in the heart, in the absence of ischaemia or myocardial injury. We used mice with mutant NLRP3 (constitutively active), NLRP3-A350V, under the control of tamoxifen-driven expression of the Cre recombinase (Nlrp3-A350V/CreT mice). The mice were treated for 10 days with tamoxifen before measuring the activity of caspase-1, the effector enzyme in the inflammasome. Tamoxifen treatment induced the inflammasome in the spleen but not in the heart, despite expression of the mutant NLRP3-A350V. The components of the inflammasome were significantly less expressed in the heart compared with the spleen. Subclinical low-dose lipopolysaccharide (LPS; 2 mg/kg) in Nlrp3-A350V/CreT mice induced the expression of the components of the inflammasome (priming), measured using real-time PCR and western blot, leading to the formation of an active inflammasome (caspase-1 activation) in the heart and LV systolic dysfunction while low-dose LPS was insufficient to induce LV systolic dysfunction in wild-type mice (all P < 0.01 for mutant vs. wild-type mice). The signalling pathway governing the inflammasome formation in the heart requires a priming signal in order for an active NLRP3 to induce caspase-1 activation and LV dysfunction.

Fc gamma receptor-TLR cross-talk elicits pro-inflammatory cytokine production by human M2 macrophages.

M2 macrophages suppress inflammation in numerous disorders, including tumour formation, infection and obesity. However, the exact role of M2 macrophages in the context of several other diseases is still largely undefined. We here show that human M2 macrophages promote inflammation instead of suppressing inflammation on simultaneous exposure to complexed IgG (c-IgG) and TLR ligands, as occurs in the context of diseases such as rheumatoid arthritis (RA). c-IgG-TLR ligand co-stimulation of M2 macrophages selectively amplifies production of pro-inflammatory cytokines TNF-α, IL-1β and IL-6 and promotes Th17 responses, which all play a critical role in RA pathology. Induction of pro-inflammatory cytokines on c-IgG co-stimulation mainly depends on Fc gamma receptor IIa (FcγRIIa), which selectively amplifies cytokine gene transcription and induces caspase-1 activation. These data indicate that FcγR-TLR cross-talk may be targeted for treatment to attenuate inflammation in RA, by restoring the anti-inflammatory function of M2 macrophages.

Cytosolic double-stranded RNA activates the NLRP3 inflammasome via MAVS-induced membrane permeabilization and K+ efflux.

The nucleotide-binding oligomerization domain-like receptor pyrin domain-containing 3 (Nlrp3) inflammasome plays an important role in inflammation by controlling the maturation and secretion of the cytokines IL-1β and IL-18 in response to multiple stimuli including pore-forming toxins, particulate matter, and ATP. Although the pathways activated by the latter stimuli lead to a decrease in intracellular K(+) concentration, which is required for inflammasome activation, the mechanism by which microbial RNA activates Nlrp3, remains poorly understood. In this study, we found that cytosolic poly(I:C), but not total RNA from healthy macrophages, macrophages undergoing pyroptosis, or mitochondrial RNA, induces caspase-1 activation and IL-1β release through the Nlrp3 inflammasome. Experiments with macrophages deficient in Tlr3, Myd88, or Trif, indicate that poly(I:C) induces Nlrp3 activation independently of TLR signaling. Further analyses revealed that the cytosolic sensors Rig-I and melanoma differentiation-associated gene 5 act redundantly via the common adaptor mitochondrial antiviral signaling (Mavs) to induce Nlrp3 activation in response to poly(I:C), but not ATP or nigericin. Mechanistically, Mavs triggered membrane permeabilization and K(+) efflux independently of the inflammasome which were required for poly(I:C)-induced Nlrp3 activation. We conclude that poly (I:C) activates the inflammasome through an Mavs-dependent surveillance pathway that converges into a common K(+) lowering step in the cytosol that is essential for the induction of Nlrp3 activation.

Enterococcus faecalis Activates Caspase-1 Leading to Increased Interleukin-1 Beta Secretion in Macrophages

IntroductionRecent studies of inflammasome activation have focused on the pathogenesis of diverse inflammatory and autoimmune diseases. Inflammasome activation results in caspase-1 activation, which is required for processing of prointerleukin (IL)-1 beta to its secreted form as well as a proinflammatory cell death (ie, pyroptosis). The purpose of this study was to analyze whether Enterococcus faecalis associated with endodontic infection induces inflammasome activation. MethodsTHP-1 macrophages were treated with E. faecalis in the presence or absence of caspase-1 inhibitors. Caspase-1 activation, pro–IL-1 beta expression, and IL-1 beta secretion were detected by immunoblotting, real-time reverse-transcription polymerase chain reaction, and enzyme-linked immunosorbent assay, respectively. Cell death was measured by lactate dehydrogenase release and propidium iodide staining. Adenosine triphosphate (ATP) release was measured by an ATP bioluminescence assay kit. ResultsE. faecalis induced caspase-1 activation and pro–IL-1 beta expression, which resulted in IL-1 beta secretion in macrophages. E. faecalis significantly induced ATP release, which is a mechanism of Nod-like receptor family protein 3 (NLRP3) inflammasome activation, whereas oxATP treatment inhibited E. faecalis–induced caspase-1 activation. E. faecalis significantly increased lactate dehydrogenase release and propidium iodide uptake, which are characteristics of pyroptosis. ConclusionsOur results show that E. faecalis may contribute to the progression of pulpal inflammation by stimulating excessive secretion of IL-1 beta and cell death.

The dual role of nod-like receptors in mucosal innate immunity and chronic intestinal inflammation.

Nucleotide-binding and oligomerization domain NOD-like receptors (NLRs) are highly conserved cytosolic pattern recognition receptors that play, in combination with toll-like receptors, a critical role in innate immunity and inflammation. These proteins are characterized by a central oligomerization domain termed nucleotide-binding domain, and a protein interaction domain containing leucine-rich repeats. Some NLRs, including NOD1 and NOD2, sense the cytosolic presence of conserved bacterial molecular signatures and drive the activation of mitogen-activated protein kinase and the transcription factor NF-κB. A different set of NLRs induces caspase-1 activation through the assembly of large protein complexes known as inflammasomes. Activation of NLR proteins results in secretion of pro-inflammatory cytokines and subsequent inflammatory responses. The critical role of NLRs in innate immunity is underscored by the fact that polymorphisms within their genes are implicated in the development of several immune-mediated diseases, including inflammatory bowel disease. Over the past few years, the role of NLRs in intestinal homeostasis has been highlighted, however the mechanism by which dysfunction in these proteins leads to aberrant inflammation is still the focus of much investigation. The purpose of this review is to systematically evaluate the function of NLRs in mucosal innate immunity and understand how genetic or functional alterations in these components can lead to the disruption of intestinal homeostasis, and the subsequent development of chronic inflammation.

Acute lymphoblastic leukemia induces caspase-1 activation in human bone marrow monocyte/macrophages and is associated with impaired CD8 function. (HUM7P.315)

Abstract B-acute lymphoblastic leukemia (ALL) is a malignancy of bone marrow-derived precursor-B cells and represents the leading cause of pediatric cancer death. Suppressive signals induced in the marrow of ALL patients may explain both the failure of host CD8 surveillance and inconsistent responses to adoptive CD8 therapy. We hypothesize that ALL induces a caspase-1 driven inflammasome in bone marrow monocyte/macrophages (BM-MM) that modulates CD8 function and contributes to relapse. To determine whether BM-MM could produce soluble factors that suppress CD8 function in the setting of ALL, we treated CFSE-labeled CD8 from healthy donor PBMC with supernatants from cocultures of primary human ALL and either CD14+ BM-MM or stromal fibroblast controls. CD8 exposed to supernatants from ALL cultured alone or with stromal fibroblasts maintained 100% proliferation, while those exposed to BM-MM cocultures failed to proliferate, regardless of whether ALL was cultured directly or in transwell with BM-MM. Cleaved caspase-1 was detected, both by qPCR (2.2-fold expression over endogenous control) and Western blot from BM-MM cultured with ALL, but not stromal fibroblasts Similar levels of soluble IL1B were detected from stromal fibroblast and BM-MM cocultures. Current studies are determining whether the reduced proliferative capacity observed in CD8 corresponds to impaired antileukemia killing and identifying specific signals downstream of caspase-1 that may be targeted to reverse this phenomenon.

Effect of Silica Particle Size on Macrophage Inflammatory Responses

Amorphous silica particles, such as nanoparticles (<100 nm diameter particles), are used in a wide variety of products, including pharmaceuticals, paints, cosmetics, and food. Nevertheless, the immunotoxicity of these particles and the relationship between silica particle size and pro-inflammatory activity are not fully understood. In this study, we addressed the relationship between the size of amorphous silica (particle dose, diameter, number, and surface area) and the inflammatory activity (macrophage phagocytosis, inflammasome activation, IL-1β secretion, cell death and lung inflammation). Irrespective of diameter size, silica particles were efficiently internalized by mouse bone marrow-derived macrophages via an actin cytoskeleton-dependent pathway, and induced caspase-1, but not caspase-11, activation. Of note, 30 nm-1000 nm diameter silica particles induced lysosomal destabilization, cell death, and IL-1β secretion at markedly higher levels than did 3000 nm-10000 nm silica particles. Consistent with in vitro results, intra-tracheal administration of 30 nm silica particles into mice caused more severe lung inflammation than that of 3000 nm silica particles, as assessed by measurement of pro-inflammatory cytokines and neutrophil infiltration in bronchoalveolar lavage fluid of mice, and by the micro-computed tomography analysis. Taken together, these results suggest that silica particle size impacts immune responses, with submicron amorphous silica particles inducing higher inflammatory responses than silica particles over 1000 nm in size, which is ascribed not only to their ability to induce caspase-1 activation but also to their cytotoxicity.

Role of the inflammasome in inflammatory responses and subsequent injury after hepatic ischemia-reperfusion injury

We read with great interest the article by Kamo et al.1 in which the authors showed that after hepatic ischemia-reperfusion (I/R) injury, inflammasome activation mediated by apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC) leads to interleukin-1β (IL-1β) production and subsequently promotes high mobility group box 1(HMGB1) induction, which triggers a Toll-like receptor 4 (TLR4)-driven inflammatory response. Consistent with their findings, we recently showed that ASC-mediated inflammasome activation plays an essential role in the initial inflammatory response after myocardial I/R injury.2 Recently, it has been shown that inflammasome components such as ASC and NLR family pyrin domain containing 3 (NLRP3) can function independently of inflammasomes. Shigeoka et al.3 showed that mice deficient in NLRP3 but not ASC or caspase-1 had reduced renal I/R. Because the inflammasome is defined as a molecular platform that induces caspase-1 activation, they concluded that an NLRP3-dependent and inflammasome-independent pathway contributed to the development of I/R injury in the kidney. Similar to their findings, we observed that hepatic I/R injury was significantly ameliorated in mice deficient for NLRP3 but not ASC. This was inconsistent with the finding of Kamo et al.1 Although the reason for this discrepancy is unclear, the differences between our study and Kamo et al.'s study are the hepatic I/R protocol used and the extent of injury. We subjected C57BL/6 wild-type mice to hepatic I/R with 60 minutes ischemia of the left lateral and median lobes (i.e., interruption of the hepatic artery and portal vein) followed by a 6-hour reperfusion, whereas Kamo et al.1 subjected mice to I/R with 90 minutes ischemia followed by a 6-hour reperfusion. Serum alanine transaminase (ALT) levels increased to ∼5,000 IU/mL in our model and 33,000 IU/mL in the model of Kamo et al. Moreover, regarding the role of IL-1β, Kato et al.4 reported that there was no difference in serum ALT levels between wild-type and IL-1 receptor-deficient mice after hepatic I/R injury and suggested a limited role of IL-1β in causing hepatic I/R injury. We agree with the authors' conclusion that the inflammasome plays a substantial role in hepatic I/R injury; however, the contribution of the inflammasome depends on the extent of injury and status of inflammatory responses; therefore, researchers should be aware of experimental disease conditions to discern the precise role of the inflammasome. Yoshiyuki Inoue, M.D.1,2 Yoshikazu Yasuda, M.D., Ph.D.2 Masafumi Takahashi, M.D., Ph.D.1 1Division of Inflammation Research, Center for Molecular Medicine, Tochigi, Japan 2Department of Surgery, Jichi Medical University, Tochigi, Japan

Rotenone Directly Induces BV2 Cell Activation via the p38 MAPK Pathway

Parkinson’s disease (PD) is the second most common neurodegenerative disease. Although its pathogenesis is still unclear, increasing evidence suggests that mitochondrial dysfunction induced by environmental toxins, such as mitochondrial complex I inhibitors, plays a significant role in the disease process. The microglia in PD brains are highly activated, and inflammation is also an essential element in PD pathogenesis. However, the means by which these toxins activate microglia is still unclear. In the present study, we found that rotenone, a mitochondrial complex I inhibitor, could directly activate microglia via the nuclear factor kappa B (NF-κB) signaling pathway, thereby inducing significantly increased expression of inflammatory cytokines. We further observed that rotenone induced caspase-1 activation and mature IL-1β release, both of which are strictly dependent on p38 mitogen-activated protein kinase (MAPK). The activation of p38 is associated with the presence of reactive oxygen species (ROS) produced by rotenone. Removal of these ROS abrogated the activation of the microglia. Therefore, our data suggest that the environmental toxin rotenone can directly activate microglia through the p38 MAPK pathway.

P2X7R/cryopyrin inflammasome axis inhibition reduces neuroinflammation after SAH

Neuroinflammation contributes to the pathogenesis of early brain injury (EBI) after subarachnoid hemorrhage (SAH). Cytotoxic events following SAH, such as extracellular accumulation of adenosine triphosphate (ATP), may activate the P2X purinoceptor 7 (P2X7R)/cryopyrin inflammasome axis, thus inducing the proinflammatory cytokine IL-1β/IL-18 secretion. We therefore hypothesized that inhibition of P2X7R/cryopyrin inflammasome axis would ameliorate neuroinflammation after SAH. In the present study, SAH was induced by the endovascular perforation in rats. Small interfering RNAs (siRNAs) of P2X7R or cryopyrin were administered intracerebroventricularly 24h before SAH. Brilliant blue G (BBG), a non-competitive antagonist of P2X7R, was administered intraperitoneally 30min following SAH. Post-assessments including SAH severity score, neurobehavioral test, brain water content, Western blot and immunofluorescence, were performed. Administration of P2X7R and cryopyrin siRNA as well as pharmacologic blockade of P2X7R by BBG ameliorated neurological deficits and brain edema at 24h following SAH. Inhibition of P2X7R/cryopyrin inflammasome axis suppressed caspase-1 activation, which subsequently decreased maturation of IL-1β/IL-18. To investigate the link between P2X7R and cryopyrin inflammasome in vivo, Benzoylbenzoyl-ATP (BzATP), a P2X7R agonist, was given to lipopolysaccharide (LPS) primed naive rats with scramble or cryopyrin siRNAs. In LPS-primed naive rats, BzATP induced caspase-1 activation and mature IL-1β release were neutralized by cryopyrin siRNA. Thus, the P2X7R/cryopyrin inflammasome axis may contribute to neuroinflammation via activation of caspase-1 and thereafter mature IL-1β/IL-18 production following SAH. Therapeutic interventions targeting P2X7R/cryopyrin pathway may be a novel approach to ameliorate EBI following SAH.

NLRP3 in myocardial ischaemia-reperfusion injury: inflammasome-dependent or -independent role in different cell types

This editorial refers to ‘The NLRP3 inflammasome is up-regulated in cardiac fibroblasts and mediates myocardial ischaemia–reperfusion injury’ by O. Sandanger et al. , pp. 164–174, this issue. An accumulating body of evidence has indicated that inflammation in the absence of pathogens, which is referred to as sterile inflammation, is mediated through an inflammasome, a large multiple protein complex in the cytosol that induces caspase-1 activation. Till date, several types of the inflammasomes have been documented. The best-characterized inflammasome is the nucleotide-binding oligomerization domain-like receptor (NLR) family pyrin domain containing 3 (NLRP3; also known as NALP3 and cryopyrin) inflammasome, which is known to participate in the pathophysiology of sterile inflammation.1 The NLRP3 inflammasome contains NLRP3 that interact with the adaptor molecule an apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC; also known as Pycard), which recruits and activates caspase-1. Because caspase-1 is an interleukin (IL)-1β-converting enzyme (ICE), it induces the processing of pro-IL-1β into mature IL-1β and the release of IL-1β, thereby causing inflammation and tissue damage. However, the induction of IL-1β release requires another signal provided by the transcriptional induction of pro-IL-1β. Hence, a system comprising pro-IL-1β induction and inflammasome-mediated IL-1β processing is believed to be necessary for the tight regulation of this potent inflammatory …

Oxidized low density lipoprotein induced caspase-1 mediated pyroptotic cell death in macrophages: implication in lesion instability?

BackgroundMacrophage death in advanced lesion has been confirmed to play an important role in plaque instability. However, the mechanism underlying lesion macrophage death still remains largely unknown.Methods and ResultsImmunohistochemistry showed that caspase-1 activated in advanced lesion and co-located with macrophages and TUNEL positive reaction. In in-vitro experiments showed that ox-LDL induced caspase-1 activation and this activation was required for ox-LDL induced macrophages lysis, IL-1β and IL-18 production as well as DNA fragmentation. Mechanism experiments showed that CD36 and NLRP3/caspase-1/pathway involved in ox-LDL induced macrophage pyroptosis.ConclusionOur study here identified a novel cell death, pyroptosis in ox-LDL induced human macrophage, which may be implicated in lesion macrophages death and play an important role in lesion instability.

Critical Role of ASC Inflammasomes and Bacterial Type IV Secretion System in Caspase-1 Activation and Host Innate Resistance to Brucella abortus Infection

Pathogens are detected by innate immune receptors that, upon activation, orchestrate an appropriate immune response. Recent studies revealed the intracellular signaling cascades involved in the TLR-initiated immune response to Brucella abortus infection. However, no report has elucidated the role of inflammasome receptors in Brucella recognition. Therefore, we decided to investigate the function of NLRC4, NLRP3, and AIM2 in sensing Brucella. In this study, we showed that NLRC4 is not required to induce caspase-1 activation and further secretion of IL-1β by B. abortus in macrophages. In contrast, we determined that AIM2, which senses Brucella DNA, and NLRP3 are partially required for caspase-1 activation and IL-1β secretion. Additionally, mitochondrial reactive oxygen species induced by Brucella were implicated in IL-1β production. Furthermore, AIM2, NLRP3, ASC, and caspase-1 knockout mice were more susceptible to B. abortus infection than were wild-type animals, suggesting that multiple ASC-dependent inflammasomes contribute to host protection against infection. This protective effect is due to the inflammatory response caused by IL-1β and IL-18 rather than pyroptosis, because we observed augmented bacterial burden in IL-1R and IL-18 knockout mice. Finally, we determined that bacterial type IV secretion system VirB and live, but not heat-killed, Brucella are required for full inflammasome activation in macrophages during infection. Taken together, our results indicate that Brucella is sensed by ASC inflammasomes that collectively orchestrate a robust caspase-1 activation and proinflammatory response.

Critical role of caspase-1 in vascular inflammation and development of atherosclerosis in Western diet-fed apolipoprotein E-deficient mice

ObjectiveRecent investigations have suggested that the inflammasome plays a role in the development of vascular inflammation and atherosclerosis; however, its precise role remains controversial. We produced double-deficient mice for apolipoprotien E (Apoe) and caspase-1 (Casp1), a key component molecule of the inflammasome, and investigated the effect of caspase-1 deficiency on vascular inflammation and atherosclerosis. Methods and resultsAtherosclerotic plaque areas in whole aortas and aortic root of Western diet (WD)-fed Apoe−/−Casp1−/− mice were significantly reduced compared to those in Apoe−/− mice. The amount of macrophages and vascular smooth muscle cells in the plaques was also reduced in Apoe−/−Casp1−/− mice. No significant differences in plasma lipid profiles and body weight change were observed between these mice. Expression of interleukin (IL)-1β in the plaques as well as plasma levels of IL-1β, IL-1α, IL-6, CCL2, and TNF-α, in Apoe−/−Casp1−/− mice were lower than those in Apoe−/− mice. In vitro experiments showed that calcium phosphate crystals induced caspase-1 activation and secretion of IL-1β and IL-1α in macrophages. ConclusionOur findings suggest that caspase-1 plays a critical role in vascular inflammation and atherosclerosis, and that modulation of caspase-1 could be a potential target for prevention and treatment of atherosclerosis.

Critical role of AIM2 in Mycobacterium tuberculosis infection

Absent in melanoma 2 (AIM2) is a sensor of cytosolic DNA that is responsible for activation of the inflammasome and host immune responses to DNA viruses and intracellular bacteria. However, the role of AIM2 in host defenses against Mycobacterium tuberculosis is unknown. Here, we show that AIM2-deficient mice were highly susceptible to intratracheal infection with M. tuberculosis and that this was associated with defective IL-1β and IL-18 production together with impaired T (h) 1 responses. Macrophages from AIM2-deficient mice infected with M. tuberculosis showed severely impaired secretion of IL-1β and IL-18 as well as activation of the inflammasome, determined by caspase-1 cleavage. Genomic DNA extracted from M. tuberculosis (Mtb DNA) induced caspase-1 activation and IL-1β/IL-18 secretion in an AIM2-dependent manner. Mtb DNA, which was present in the cytosol, co-localized with AIM2. Taken together, these findings demonstrate that AIM2 plays an important role in M. tuberculosis infection through the recognition of Mtb DNA.

High-mobility group box 1 activates caspase-1 and promotes hepatocellular carcinoma invasiveness and metastases

Hypoxia is often found in solid tumors and is associated with tumor progression and poor clinical outcomes. The exact mechanisms related to hypoxia-induced invasion and metastasis remain unclear. We elucidated the mechanism by which the nuclear-damage-associated molecular pattern molecule, high-mobility group box 1 (HMGB1), released under hypoxic stress, can induce an inflammatory response to promote invasion and metastasis in hepatocellular carcinoma (HCC) cells. Caspase-1 activation was found to occur in hypoxic HCC cells in a process that was dependent on the extracellular release of HMGB1 and subsequent activation of both Toll-like receptor 4 (TLR4)- and receptor for advanced glycation endproducts (RAGE)-signaling pathways. Downstream from hypoxia-induced caspase-1 activation, cleavage and release of proinflammatory cytokines interleukin (IL)-1β and -18 occurred. We further demonstrate that overexpression of HMGB1 or treatment with recombinant HMGB1 enhanced the invasiveness of HCC cells, whereas stable knockdown of HMGB1 remarkably reduced HCC invasion. Moreover, in a murine model of HCC pulmonary metastasis, stable knockdown of HMGB1 suppressed HCC invasion and metastasis. These results suggest that in hypoxic HCC cells, HMGB1 activates TLR4- and RAGE-signaling pathways to induce caspase-1 activation with the subsequent production of multiple inflammatory mediators, which, in turn, promote cancer invasion and metastasis.

Abstract 484: Acidic pH Triggers Secretion of Proinflammatory Cytokines in Human Macrophages

Extracellular acidification can occur in atherosclerotic lesions, specifically in areas with high numbers of metabolically active macrophages secreting lactate and hydrogen ions. There is a growing body of evidence showing that acidification of the extracellular environment causes fundamental changes in the functions of immune cells. Here we studied the effect of low pH on the secretion of proinflammatory cytokines by cultured human macrophages. Using custom manufactured cell culture media with neutral and acidic pH (pH 7.5-6.0) we discovered that acidic pH triggers secretion of interleukin(IL)-1beta and IL-18, but not of IL-1alpha. IL-1beta and IL-18 are both produced as proforms that are processed into the biologically active secreted form by the caspase-1 protease. The IL-1beta secretion induced by acidic pH was paralleled by caspase-1 activation, and inhibition of caspase-1 completely abolished IL-1beta secretion. Acidic pH-induced IL-1beta secretion was also abolished at high extracellular potassium concentrations, which implies that potassium efflux from cells is a crucial step upstream of caspase-1 activation. We are currently exploring the involvement of inflammasomes in the acidic pH -induced caspase-1 activation. In conclusion, acidic microenvironments of atherosclerotic lesions may induce proinflammatory cytokine secretion in macrophages, so amplifying the progression of atherogenesis.

Attacking tumor cells with a dual ligand for innate immune receptors

Attacking tumor cells with a dual ligand for innate immune receptors