Nucleotide-binding Domain And Leucine-rich Repeat Protein 3 Inflammasome Activation Research Articles

The VDAC1 oligomerization regulated by ATP5B leads to the NLRP3 inflammasome activation in the liver cells under PFOS exposure

As a persistent organic pollutant, perfluorooctane sulfonate (PFOS) has a serious detrimental impact on human health. It has been suggested that PFOS is associated with liver inflammation. However, the underlying mechanisms are still unclear. Here, PFOS was found to elevate the oligomerization tendency of voltage-dependent anion channel 1 (VDAC1) in the mice liver and human normal liver cells L-02. Inhibition of VDAC1 oligomerization alleviated PFOS-induced nucleotide-binding domain and leucine-rich repeat protein-3 (NLRP3) inflammasome activation. Cytoplasmic membrane VDAC1 translocated to mitochondria was also observed in response to PFOS. Therefore, the oligomerization of VDAC1 occurred mainly in the mitochondria. VDAC1 was found to interact with the ATP synthase beta subunit (ATP5B) under PFOS treatment. Knockdown of ATP5B or immobilization of ATP5B to the cytoplasmic membrane alleviated the increased VDAC1 oligomerization and NLRP3 inflammasome activation. Therefore, our results suggested that PFOS induced NLRP3 inflammasome activation through VDAC1 oligomerization, a process dependent on ATP5B to transfer VDAC1 from the plasma membrane to the mitochondria. The findings offer novel perspectives on the activation of the NLRP3 inflammasome, the regulatory mode on VDAC1 oligomerization, and the mechanism of PFOS toxicity.

Protective effect of scallop-derived plasmalogen against vascular dysfunction, via the pSTAT3/PIM1/NFATc1 axis, in a novel mouse model of Alzheimer’s disease with cerebral hypoperfusion

A strong relationship between Alzheimer’s disease (AD) and vascular dysfunction has been the focus of increasing attention in aging societies. In the present study, we examined the long-term effect of scallop-derived plasmalogen (sPlas) on vascular remodeling-related proteins in the brain of an AD with cerebral hypoperfusion (HP) mouse model. We demonstrated, for the first time, that cerebral HP activated the axis of the receptor for advanced glycation endproducts (RAGE)/phosphorylated signal transducer and activator of transcription 3 (pSTAT3)/provirus integration site for Moloney murine leukemia virus 1 (PIM1)/nuclear factor of activated T cells 1 (NFATc1), accounting for such cerebral vascular remodeling. Moreover, we also found that cerebral HP accelerated pSTAT3-mediated astrogliosis and activation of the nucleotide-binding domain and leucine-rich repeat protein 3 (NLRP3) inflammasome, probably leading to cognitive decline. On the other hand, sPlas treatment attenuated the activation of the pSTAT3/PIM1/NFATc1 axis independent of RAGE and significantly suppressed NLRP3 inflammasome activation, demonstrating the beneficial effect on AD.

Biotransformation of soluble-insoluble lanthanum species and its induced NLRP3 inflammasome activation and chronic fibrosis

Soluble lanthanum (La)(Ⅲ) species that have been extensively used as fertilizers in agriculture can potentially get into the human body through foods and environment. Most soluble La(Ⅲ) species can rapidly transform into insoluble La(Ⅲ) species under physiological conditions, however, their potential biological behavior and chronic toxicity are rarely investigated. In the present study, insoluble La(Ⅲ) species formed under physiological condition were identified as nanoscale or microscale particles, and their major components were found to experience biotransformation process upon contact with cells. Insoluble La(Ⅲ) species could adhere to extracellular membrane or be internalized into cells, capable of activating a nucleotide-binding domain and leucine-rich repeat protein 3 (NLRP3) inflammasome. The underlying mechanism could be ascribed to K+ efflux and lysosomal rupture because these insoluble La(Ⅲ) species locating at extracellular membrane could reduce the unsaturated fatty acids of cell membrane, leading to potassium (K+) efflux, and those internalized into cells could consume the phospholipids of lysosomal membrane, leading to lysosomal rupture. Mice daily drinking soluble La(Ⅲ) species to mimic drinking tea process for 90 days were found to present NLRP3 inflammasome activation in liver and kidney, as a result of chronic fibrosis, which is potentially correlated to insoluble La(Ⅲ) species formation.

NLRP3 Inflammasome Inhibition Prevents α-Synuclein Pathology by Relieving Autophagy Dysfunction in Chronic MPTP-Treated NLRP3 Knockout Mice.

Recent researches showed that nucleotide-binding domain and leucine-rich repeat protein 3 (NLRP3) inflammasome inhibition exerted dopaminergic neuroprotection in cellular or animal models of Parkinson's disease (PD). NLRP3 inflammasome has been proposed as a drug target for treatment of PD. However, the interplay between chronic NLRP3 inflammasome and progressive α-synuclein pathology keeps poorly understood. Moreover, the potential mechanism keeps unknown. In the present study, we investigate whether NLRP3 inflammasome inhibition prevents α-synuclein pathology by relieving autophagy dysfunction in the chronic 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP) mouse model of PD. NLRP3 knockout mice and their wild-type counterparts were treated with continuous MPTP administration via osmotic mini-pumps. Dopaminergic neuronal degeneration was assessed by western blotting and immunohistochemistry (IHC). The levels of dopamine and its metabolites were determined using high-performance liquid chromatography. NLRP3 inflammasome activation and autophagy biomarkers were assessed by western blot. The expressions of pro-inflammatory cytokines were measured by ELISA. The glial reaction and α-synuclein pathology were assessed by IHC and immunofluorescence. Our results show that NLRP3 inflammasome inhibition via NLRP3 knockout not only protects against nigral dopaminergic degeneration and striatal dopamine deletion but also prevents nigral pathological α-synuclein formation in PD mice. Furthermore, it significantly suppresses MPTP-induced glial reaction accompanied by the secretion of pro-inflammatory cytokines in the midbrain of mice. Most importantly, it relieves autophagy dysfunction in the midbrain of PD mice. Collectively, we demonstrate for the first time that improving autophagy function is involved in the preventive effect of NLRP3 inflammasome inhibition on α-synuclein pathology in PD.

Nrf2/HO-1 partially regulates cytoprotective effects of carbon monoxide against urban particulate matter-induced inflammatory responses in oral keratinocytes

IntroductionExposure to airborne particulate matter (PM) increases the proportion of oral inflammatory diseases. During the formation of inflammatory conditions, the nucleotide-binding domain and leucine-rich repeat protein 3 (NLRP3) inflammasome activation plays an important regulator. Carbon monoxide (CO) arising from heme degradation, catalyzed particularly by heme oxygenase-1 (HO-1), has been shown to own cytoprotective effects including anti-inflammation and antioxidant. Here, we determined the novel mechanisms of carbon monoxide releasing molecule-2 (CORM-2) on PM-induced inflammatory responses in human oral keratinocytes (HOKs). MethodsThe effects of CORM-2 on the expression of various inflammatory proteins induced by PM were determined by Western blot, real-time PCR, promoter assay, and ELISA. The involvement of signaling molecules in these responses was studied by using the selective pharmacological inhibitors and siRNAs. ResultsWe proved that PM enhanced C-reactive protein (CRP) levels, NLRP3 inflammasome and caspase-1 activation, and IL-1β release, which were reduced by preincubation with CORM-2. Transfection with PKCα siRNA and preincubation with the ROS scavenger (N-acetyl-cysteine, NAC), an inhibitor of NADPH oxidase (diphenyleneiodonium, DPI), or the mitochondria-specific superoxide scavenger (MitoTEMPO) inhibited PM-mediated inflammatory responses. In addition, PM-regulated PKCα and NADPH oxidase activation as well as NADPH oxidase- and mitochondria-derived ROS generation were inhibited by CORM-2, but not inactivate CORM-2 (iCORM-2) pretreatment. At the end, we confirmed that CORM-2 improved PM-induced inflammatory responses via the induction of Nrf2 activation and HO-1 expression. ConclusionWe suggest that CORM-2 inhibits PM-induced inflammatory responses in HOKs via the inhibition of PKCα/ROS/NLRP3 inflammasome activation combined with the induction of Nrf2/HO-1 expression.

Atractylenolide III reduces NLRP3 inflammasome activation and Th1/Th2 imbalances in both in vitro and in vivo models of asthma.

Paediatric asthma is a common inflammatory disease in children. Atractylenolide III is an active component of the Atractylodes rhizome, an herbal medicine that has been used as an asthma treatment. This study aimed to explore the effects and underlying mechanisms of atractylenolide III in IL-4-induced 16HBE cells and ovalbumin-induced asthmatic mice. The results showed that IL-4 stimulation significantly decreased, and atractylenolide III treatment increased, growth and apoptosis of 16HBE cells. In 16HBE cells, administration of atractylenolide III also significantly suppressed the IL-4-induced increases in the expression of cleaved caspase-1; apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC); and nucleotide-binding domain and leucine-rich repeat protein 3 (NLRP3). Moreover, the numbers of total leukocytes, neutrophils, eosinophils, and macrophages significantly increased in ovalbumin-induced mice, and then decreased after atractylenolide III treatment. In ovalbumin-induced asthmatic mice, atractylenolide III treatment also significantly inhibited NLRP3 inflammasome activation and restored the Th1/Th2 balance. These results indicate that atractylenolide III reduced NLRP3 inflammasome activation and regulated the Th1/Th2 balance in IL-4 induced 16HBE cells and ovalbumin-induced asthmatic mice, suggesting it has a protective effect that may be useful in the treatment of paediatric asthma.

Resveratrol alleviates chronic “real-world” ambient particulate matter-induced lung inflammation and fibrosis by inhibiting NLRP3 inflammasome activation in mice

BackgroundInhalation of fine particulate matter (PM2.5) induces the occurrence of lung inflammation and fibrosis, but its molecular mechanism remains unclear. Resveratrol (RES) is known to have anti-inflammatory properties in many pulmonary diseases. Here, we aimed to investigate the effect of long-term “real-world” ambient PM exposure on lung inflammation and fibrosis and further explore the protective effect and mechanism of RES. Methods and resultsRES (50 and 100 mg/kg.bw) was administered to C57BL/6J mice that were exposed to ambient PM for 5 months. The control group breathed filtered air without RES, and the PM group was exposed to PM without RES. The inflammatory cytokine levels in bronchoalveolar lavage fluid (BALF) and lung fibrosis were evaluated by enzyme-linked immune sorbent assay (ELISA) kits and Masson's trichrome staining. The real-time PCR and Western blot analysis were used to determine the signal pathway. In vivo, PM exposure markedly elevated the levels of inflammatory cytokines and TGF-β1 in BALF, induced lung fibrosis. Meanwhile, PM exposure triggered autophagy process and activated the nucleotide-binding domain and leucine-rich repeat protein 3 (NLRP3) inflammasome in lung. Also, RES treatment abolished PM-induced lung inflammation and fibrosis, and inhibited autophagic process and NLRP3 inflammasome activation. In vitro, PM2.5-induced cytotoxicity in BEAS-2B cells dose-dependently. Besides, RES alleviated PM2.5-induced cytotoxicity, inhibited autophagic process and NLRP3 inflammasome activity and decreased IL-1β production in BEAS-2B cells. ConclusionLong-term PM exposure induced lung inflammation and fibrosis, and RES intervention alleviated these adverse effects via inhibiting autophagy-related NLRP3 inflammasome activation.

Carbon monoxide releasing molecule-2 protects against particulate matter-induced lung inflammation by inhibiting TLR2 and 4/ROS/NLRP3 inflammasome activation

Exposure to airborne particulate matter (PM) not only causes lung inflammation and chronic respiratory diseases, but also increases the incidence and mortality of cardiopulmonary diseases. The nucleotide-binding domain and leucine-rich repeat protein 3 (NLRP3) inflammasome activation has been shown to play a critical role in the formation of many chronic disorders. On the other hand, carbon monoxide (CO) has been shown to possess anti-inflammatory and antioxidant effects in many tissues and organs. Here, we investigated the effects and mechanisms of carbon monoxide releasing molecule-2 (CORM-2) on PM-induced inflammatory responses in human pulmonary alveolar epithelial cells (HPAEpiCs). We found that PM induced C-reactive protein (CRP) expression, NLRP3 inflammasome activation, IL-1β secretion, and caspase-1 activation, which were inhibited by pretreatment with CORM-2. In addition, transfection with siRNA of Toll-like receptor 2 (TLR2) or TLR4 and pretreatment with an antioxidant (N-acetyl-cysteine, NAC), the inhibitor of NADPH oxidase (diphenyleneiodonium, DPI), or a mitochondria-specific superoxide scavenger (MitoTEMPO) reduced PM-induced inflammatory responses. CORM-2 also inhibited PM-induced NADPH oxidase activity and NADPH oxidase- and mitochondria-derived ROS generation. However, pretreatment with inactivate CORM-2 (iCORM-2) had no effects on PM-induced inflammatory responses. Finally, we showed that CORM-2 inhibited PM-induced CRP, NLRP3 inflammasome, and ASC protein expression in the lung tissues of mice and IL-1β levels in the serum of mice. PM-enhanced leukocyte count in bronchoalveolar lavage fluid in mice was reduced by CORM-2. The results of this study suggested a protective role of CORM-2 in PM-induced lung inflammation by inhibiting the TLR2 and TLR4/ROS-NLRP3 inflammasome-CRP axial.

PPARß/δ agonist alleviates NLRP3 inflammasome-mediated neuroinflammation in the MPTP mouse model of Parkinson’s disease

Recent studies have indicated that peroxisome proliferator-activated receptor β/δ (PPARß/δ) agonists exert neuroprotective effects in the model of Parkinson’s disease (PD). Furthermore, PPARß/δ agonists have been shown to have potential anti-inflammatory activity, but the underlying mechanisms remain obscure. Emerging evidence indicates that the nucleotide-binding domain and leucine-rich-repeat-protein 3 (NLRP3) inflammasome-mediated neuroinflammation plays a crucial role in the pathogenesis of PD. In the present study we investigate whether PPARß/δ agonists alleviate NLRP3-mediated neuroinflammation in the 1- methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP) mouse model of PD. We administered GW501516, a selective and high-affinity PPARß/δ agonist, via intracerebroventricular infusion. Locomotor activities were tested by open field tests and the pole test. The levels of dopamine and its metabolites were determined using highperformance liquid chromatography.Dopaminergic neurodegeneration was assessed via Western blot analysis. The levels of oxidative stress were detected via spectrophotometric assays. The expressions of pro-inflammatory cytokines were measured by performing quantitative real-time RT-PCR and ELISA. Western blot analysis was used to assess NLRP3 inflammasome activation. Our results show that GW501516 reduced movement impairment in PD mice; furthermore, it attenuated dopaminergic neurodegeneration in the midbrain and the depletion of dopamine in the striatum and it inhibited inflammatory reactions and NLRP3 inflammasome activation in the midbrain of PD mice. More importantly, it attenuated astrocytic reaction but had no significant effect on microglial reaction in the midbrain of PD mice. Collectively, our findings demonstrate for the first time that the specific PPARß/δ agonist GW501516 alleviates NLRP3 inflammasome-mediated neuroinflammation in astrocytes in the MPTP mouse model of PD.

Effects on IL-1β signaling activation induced by water and organic extracts of fine particulate matter (PM2.5) in vitro

Fine particulate matter (PM2.5) air pollution poses a major risk to human health worldwide, and absorbed chemicals play a key role in determining the toxicity of PM2.5. After inhalation and entry into the lungs, PM2.5 components induce pro-inflammatory cytokines (e.g., interleukin (IL)-1β) in pulmonary cells. To test whether PM2.5 components induce IL-1β through signing pathways that include the toll-like receptor 4 (TLR4)/nuclear factor-κ-gene binding (NF-κB), nucleotide-binding domain and leucine-rich repeat protein 3 (NLRP3), we exposed the mouse macrophage cell-line RAW264.7 to both water and organic extracts of PM2.5 sampled over a 1-year period in Beijing, China. Varying degrees of oxidative stress and inflammatory responses were induced following exposure, while organic extracts of PM2.5 collected during the heating season induced more significant responses. This response is attributed to high concentrations of polycyclic aromatic hydrocarbons (PAHs) originating from coal combustion and biomass burning for domestic heating. The inhibition of signaling molecules suggested that increased IL-1β was associated with the TLR4/NF-κB pathway and NLRP3 inflammasome activation, with a slightly difference between water and organic extracts exposure groups, which was likely the result of different chemical components. Our study elucidated a potentially important mechanism by which PM2.5 components could trigger pulmonary inflammation, thus improving our understanding of the deleterious effects of this important and prevalent form of air pollution.

The NLRP3 inflammasome modulates sleep and NREM sleep delta power induced by spontaneous wakefulness, sleep deprivation and lipopolysaccharide

Both sleep loss and pathogens can enhance brain inflammation, sleep, and sleep intensity as indicated by electroencephalogram delta (δ) power. The pro-inflammatory cytokine interleukin-1 beta (IL-1β) is increased in the cortex after sleep deprivation (SD) and in response to the Gram-negative bacterial cell-wall component lipopolysaccharide (LPS), although the exact mechanisms governing these effects are unknown. The nucleotide-binding domain and leucine-rich repeat protein-3 (NLRP3) inflammasome protein complex forms in response to changes in the local environment and, in turn, activates caspase-1 to convert IL-1β into its active form. SD enhances the cortical expression of the somnogenic cytokine IL-1β, although the underlying mechanism is, as yet, unidentified. Using NLRP3-gene knockout (KO) mice, we provide evidence that NLRP3 inflammasome activation is a crucial mechanism for the downstream pathway leading to increased IL-1β-enhanced sleep. NLRP3 KO mice exhibited reduced non-rapid eye movement (NREM) sleep during the light period. We also found that sleep amount and intensity (δ activity) were drastically attenuated in NLRP3 KO mice following SD (homeostatic sleep response), as well as after LPS administration, although they were enhanced by central administration of IL-1β. NLRP3, ASC, and IL1β mRNA, IL-1β protein, and caspase-1 activity were greater in the somatosensory cortex at the end of the wake-active period when sleep propensity was high and after SD in wild-type but not NLRP3 KO mice. Thus, our novel and converging findings suggest that the activation of the NLRP3 inflammasome can modulate sleep induced by both increased wakefulness and a bacterial component in the brain.

Activation of Cannabinoid Receptor 2 Ameliorates DSS-Induced Colitis through Inhibiting NLRP3 Inflammasome in Macrophages.

Activation of cannabinoid receptor 2 (CB2R) ameliorates inflammation, but the underlying mechanism remains unclear. In the present study, we examined whether activation of CB2R could suppress the nucleotide-binding domain and leucine-rich repeat protein 3 (NLRP3) inflammasome. In peritoneal macrophages isolated from C57BL/6 mice, LPS/DSS challenge for 24 h increased the expression of the components of NLRP3 inflammasome NLRP3, Casp-1 p20/Casp-1 p45 ratio, proIL-1β and IL-1β and also enhanced autophagy (LC3-II/LC3-I ratio, Beclin-1 and SQSTM1). Pretreatment of peritoneal macrophages with HU 308, a selective CB2R agonist, attenuated LPS/DSS-induced NLRP3 inflammasome activation, but further enhanced autophagy. In comparison with wild-type (WT) control, peritoneal macrophages from CB2R knockout (KO) mice had more robust NLRP3 inflammasome activation and attenuated autophagy upon LPS/DSS challenge. Knockdown autophagy-related gene 5 (Atg5) with a siRNA in peritoneal macrophages attenuated the inhibitory effects of HU 308 on LPS/DSS-induced NLRP3 inflammasome activation in vitro. In vivo, HU308 treatment attenuated DSS-induced colitis mice associated with reduced colon inflammation and inhibited NLRP3 inflammasome activation in wild-type mice. In CB2R KO mice, DSS-induced inflammation and NLRP3 inflammasome activation were more pronounced than those in WT control. Finally, we demonstrated that AMPK-mTOR-P70S6K signaling pathway was involved in this CB2R-mediated process. We conclude that activation of CB2R ameliorates DSS-induced colitis through enhancing autophagy that may inhibit NLRP3 inflammasome activation in macrophages.

The NLRP3 Inflammasome as a Novel Player of the Intercellular Crosstalk in Metabolic Disorders

The combination of obesity and type 2 diabetes is a serious health problem, which is projected to afflict 300 million people worldwide by 2020. Both clinical and translational laboratory studies have demonstrated that chronic inflammation is associated with obesity and obesity-related conditions such as insulin resistance. However, the precise etiopathogenetic mechanisms linking obesity to diabetes remain to be elucidated, and the pathways that mediate this phenomenon are not fully characterized. One of the most recently identified signaling pathways, whose activation seems to affect many metabolic disorders, is the “inflammasome,” a multiprotein complex composed of NLRP3 (nucleotide-binding domain and leucine-rich repeat protein 3), ASC (apoptosis-associated speck-like protein containing a CARD), and procaspase-1. NLRP3 inflammasome activation leads to the processing and secretion of the proinflammatory cytokines interleukin- (IL-) 1β and IL-18. The goal of this paper is to review new insights on the effects of the NLRP3 inflammasome activation in the complex mechanisms of crosstalk between different organs, for a better understanding of the role of chronic inflammation in metabolic disease pathogenesis. We will provide here a perspective on the current research on NLRP3 inflammasome, which may represent an innovative therapeutic target to reverse the detrimental metabolic consequences of the metabolic inflammation.

Reversal of the deleterious effects of chronic dietary HFCS-55 intake by PPAR-δ agonism correlates with impaired NLRP3 inflammasome activation

Although high-fructose corn syrup (HFCS-55) is the major sweetener in foods and soft-drinks, its potential role in the pathophysiology of diabetes and obesity (“diabesity”) remains unclear. Peroxisome-proliferator activated receptor (PPAR)-δ agonists have never been tested in models of sugar-induced metabolic abnormalities. This study was designed to evaluate (i) the metabolic and renal consequences of HFCS-55 administration (15%wt/vol in drinking water) for 30 weeks on male C57Bl6/J mice and (ii) the effects of the selective PPAR-δ agonist GW0742 (1mg/kg/day for 16 weeks) in this condition. HFCS-55 caused (i) hyperlipidemia, (ii) insulin resistance, and (iii) renal injury/inflammation. In the liver, HFCS-55 enhanced the expression of fructokinase resulting in hyperuricemia and caused abnormalities in known insulin-driven signaling events. In the kidney, HFCS-55 enhanced the expression of the NLRP3 (nucleotide-binding domain and leucine-rich-repeat-protein 3) inflammasome complex, resulting in caspase-1 activation and interleukin-1β production. All of the above effects of HFCS-55 were attenuated by the specific PPAR-δ agonist GW0742. Thus, we demonstrate for the first time that the specific PPAR-δ agonist GW0742 attenuates the metabolic abnormalities and the renal dysfunction/inflammation caused by chronic HFCS-55 exposure by preventing upregulation of fructokinase (liver) and activation of the NLRP3 inflammasome (kidney).