NOD-like Receptor Family Pyrin Domain Research Articles

Melatonin suppresses TLR4-mediated RSV infection in the central nervous cells by inhibiting NLRP3 inflammasome formation and autophagy.

Respiratory syncytial virus (RSV) infects neuronal cells in the central nervous system (CNS), resulting in neurological symptoms. In the present study, we intended to explore the mechanism of RSV infection-induced neuroinflammatory injury from the perspective of the immune response and sought to identify effective protective measures against the injury. The findings showed that toll-like receptor 4 (TLR4) was activated after RSV infection in human neuronal SY5Y cells. Furthermore, TLR4 activation induced autophagy and apoptosis in neuronal cells, promoted the formation of the NOD-like receptor family pyrin domain containing 3 (NLRP3) inflammasome, and increased the secretion of downstream inflammatory cytokines such as interleukin-1β (IL-1β), interleukin-18 (IL-18) and tumour necrosis factor-α (TNF-α). Interestingly, blockade of TLR4 or treatment with exogenous melatonin significantly suppressed TLR4 activation as well as TLR4-mediated apoptosis, autophagy and immune responses. Therefore, we infer that melatonin may act on the TLR4 to ameliorate RSV-induced neuronal injury, which provides a new therapeutic target for RSV infection.

Immune Regulation Patterns in Response to Environmental Pollutant Chromate Exposure-Related Genetic Damage: A Cross-Sectional Study Applying Machine Learning Methods.

Exposure to hexavalent chromium damages genetic materials like DNA and chromosomes, further elevating cancer risk, yet research rarely focuses on related immunological mechanisms, which play an important role in the occurrence and development of cancer. We investigated the association between blood chromium (Cr) levels and genetic damage biomarkers as well as the immune regulatory mechanism involved, such as costimulatory molecules, in 120 workers exposed to chromates. Higher blood Cr levels were linearly correlated with higher genetic damage, reflected by urinary 8-hydroxy-2'-deoxyguanosine (8-OHdG) and blood micronucleus frequency (MNF). Exploratory factor analysis revealed that both positive and negative immune regulation patterns were positively associated with blood Cr. Specifically, higher levels of programmed cell death protein 1 (PD-1; mediated proportion: 4.12%), programmed cell death ligand 1 (PD-L1; 5.22%), lymphocyte activation gene 3 (LAG-3; 2.11%), and their constitutive positive immune regulation pattern (5.86%) indirectly positively influenced the relationship between blood Cr and urinary 8-OHdG. NOD-like receptor family pyrin domain containing 3 (NLRP3) positively affected the association between blood Cr levels and inflammatory immunity. This study, using machine learning, investigated immune regulation and its potential role in chromate-induced genetic damage, providing insights into complex relationships and emphasizing the need for further research.

Compatibility with Fructus Ligustri Lucidi Effectively Mitigates Idiosyncratic Liver Injury of Epimedii Folium by Modulating NOD-like Receptor Family Pyrin Domain Containing 3 Inflammasome Activation

Abstract Background: Idiosyncratic drug-induced liver injury (IDILI) is a serious side effect of drugs, Epimedii Folium (EF) is unequivocally implicated in idiosyncratic liver injury onset, potentially due to its ability to perturb the NOD-like receptor family pyrin domain containing 3 (NLRP3) inflammasome. Fructus Ligustri Lucidi (FLL), a frequently used medicinal combination with EF, has not yet been investigated for its ability to ameliorate EF-associated hepatotoxicity. Aims and Objectives: Study on the mechanism of compatibility of FLL to alleviate liver injury caused by EF. Materials and Methods: Western blot was used to determine the expression of related proteins, ELISA was used to detect the secretion of related inflammatory factors IL-1β, IL-18, IL-6 and TNF-α, liver injury indexes were detected and liver pathological tissue staining was used to evaluate the liver injury. Results: Our results demonstrated that EF exerted a particular augmenting effect on the stimulation of the NLRP3 inflammasome mediated by nigericin or ATP, whereas FLL suppressed the NLRP3 inflammasome stimulation. Furthermore, an equal EF to FLL ratio significantly reduced the stimulatory effects of EF. Moreover, EF has the potential to induce hepatic injury and augment pro-inflammatory cytokine synthesis in rats subjected to LPS. However, when combined with FLL, the detrimental effects of EF were mitigated. Conclusion: FLL possesses the capacity to attenuate EF-associated hepatotoxicity by suppressing EF-triggered NLRP3 inflammasome activation. Thus, FLL holds promise for improving the clinical safety profile of EF, shedding light on the potential of compatibility and detoxification theories in traditional Chinese medicine.

GNAI3 mediated by Lin28A regulates lipopolysaccharide-induced inflammation and osteogenic differentiation in periodontal stem cells by mediating the NF-κB/NLRP3 inflammasome pathway

ObjectivesThe aim of this study was to investigate the regulatory role of G protein subunit alpha i3 (GNAI3) in periodontitis. DesignFollowing the induction of human periodontal ligament stem cells (hPDLSCs) with lipopolysaccharide (LPS), the mRNA and protein expressions of GNAI3 and Lin28A were detected by real-time quantitative polymerase chain reaction (RT-qPCR) and western blot. The transfection efficiency of Oe-GNAI3 and sh-Lin28A was examined by virtue of RT-qPCR and western blot. With the application of ELISA and flow cytometry, the releases of inflammatory cytokines and cell apoptosis were appraised. Alkaline phosphatase (ALP) staining and alizarin red S (ARS) staining were conducted to evaluate osteogenic differentiation. Next, the binding ability of Lin28A with GNAI3 mRNA was estimated by radioimmunoprecipitation (RIP) assay while the stability of GNAI3 mRNA was assessed utilizing RT-qPCR. Western blot was employed for the measurement of inflammation-, apoptosis- and nuclear factor-kappaB (NF-κB)/NOD-like receptor family pyrin domain containing 3 (NLRP3) inflammasome pathway-related proteins and osteogenic markers. ResultsThe expression of GNAI3 was down-regulated in LPS-induced hPDLSCs. After the transfection with Oe-GNAI3, the inflammation and apoptosis in LPS-induced hPDLSCs were inhibited while osteogenic differentiation was promoted. Moreover, Lin28A could stabilize GNAI3 mRNA and Lin28A knockdown significantly reduced GNAI3 expression. Further experiments verified that the inhibitory effects of GNAI3 overexpression on LPS-induced cellular inflammation and cell apoptosis as well as the promotive effects on osteogenic differentiation in hPDLSCs were all partially counteracted by Lin28A depletion, which may possibly be mediated via the regulation of the NF-κB/NLRP3 inflammasome pathway. ConclusionGNAI3 that mediated by Lin28A regulates the inflammation and osteogenic differentiation in LPS-induced hPDLSCs by mediating the NF-κB/NLRP3 inflammasome pathway.

KLF5-mediated pyroptosis of airway epithelial cells leads to airway inflammation in asthmatic mice through the miR-182–5p/TLR4 axis

Asthma is viewed as an airway disease and an inflammatory condition. This study aims to reveal the role of Kruppel-like factor 5 (KLF5)-mediated pyroptosis of airway epithelial cells in airway inflammation in asthma. The asthmatic mouse model was established. The mice were infected with the lentivirus containing sh-KLF5, antagomiR-182–5p, and pc-Toll-like receptor 4 (TLR4). Airway hyperresponsiveness was measured, and the cells in bronchoalveolar lavage fluid (BALF) were sorted and counted. The expression levels of interleukin (IL)-4/IL-13/IL-6/IL-18/IL-1β/NOD-like receptor family pyrin domain containing 3 (NLRP3)/N-gasdermin D (GSDMD-N)/cleaved caspase-1 were detected. The pathological changes in lung tissue were observed. The enrichment of KLF5 in the miR-182–5p promoter region was measured. The binding relationship among KLF5, miR-182–5p, and TLR4 were analyzed. KLF5 was highly expressed in asthmatic mice. Silencing KLF5 improved airway resistance and lung dynamic compliance, reduced the cells in BALF and the expression of IL-4/IL-13/IL-6/NLRP3/GSDMD-N/cleaved caspase-1/IL-18/IL-1β, and alleviated the pathological changes. Mechanistically, KLF5 bonded to the miR-182–5p promoter to inhibit miR-182–5p expression, and miR-182–5p inhibited TLR4. Silencing miR-182–5p or TLR4 overexpression reversed the improvement of silencing KLF5 on airway inflammation and pyroptosis in asthmatic mice. In conclusion, KLF5 inhibited miR-182–5p to promote TLR4 expression, thus aggravating pyroptosis and airway inflammation in asthmatic mice.

Inhibition of xanthine oxidase alleviated pancreatic necrosis via HIF-1α-regulated LDHA and NLRP3 signaling pathway in acute pancreatitis

Acute pancreatitis (AP) is a potentially fatal condition with no targeted treatment options. Although inhibiting xanthine oxidase (XO) in the treatment of AP has been studied in several experimental models and clinical trials, whether XO is a target of AP and what its the main mechanism of action is remains unclear. Here, we aimed to re-evaluate whether XO is a target aggravating AP other than merely generating reactive oxygen species that trigger AP. We first revealed that XO expression and enzyme activity were significantly elevated in the serum and pancreas of necrotizing AP models. We also found that allopurinol and febuxostat, as purine-like and non-purine XO inhibitors, respectively, exhibited protective effects against pancreatic acinar cell death in vitro and pancreatic damage in vivo at different doses and treatment time points. Moreover, we observed that conditional Xdh overexpression aggravated pancreatic necrosis and severity. Further mechanism analysis showed that XO inhibition restored the hypoxia-inducible factor 1-alpha (HIF-1α)-regulated lactate dehydrogenase A (LDHA) and NOD-like receptor family pyrin domain containing 3 (NLRP3) signaling pathways and reduced the enrichment of 13C6-glucose to 13C3-lactate. Lastly, we observed that clinical circulatory XO activity was significantly elevated in severe cases and correlated with C-reactive protein levels, while pancreatic XO and urate were also increased in severe AP patients. These results together indicated that proper inhibition of XO might be a promising therapeutic strategy for alleviating pancreatic necrosis and preventing progression of severe AP by downregulating HIF-1α-mediated LDHA and NLRP3 signaling pathways.

V-set and immunoglobulin domain containing 4 inhibits oxidative stress, mitochondrial dysfunction, and inflammation to attenuate Parkinson's disease progression by activating the JAK2/STAT3 pathway

ObjectiveV-set and immunoglobulin domain containing 4 (VSIG4) inhibits neurological dysfunction, microglial M1 polarization, and inflammation to participate in the progression of neurological disorders, but evidence regarding Parkinson's disease (PD) is scarce. The present study intended to investigate the engagement of VSIG4 in PD progression, and the potential mechanism. MethodsBV-2 cells were treated with 1-Methyl-4-phenylpyridinium (MPP+) to establish PD model. MPP+ treated BV-2 cells were infected with VSIG4 overexpression adenovirus-associated virus (AAV) (oeVSIG4) and negative control AAV (oeNC), and AZD1480 (JAK2 inhibitor) was added to these cells. ResultsMPP+ reduced VSIG4 mRNA (P < 0.05) and protein (P < 0.05) in BV-2 cells. Interestingly, VSIG4 reduced malondialdehyde (P < 0.01), reactive oxygen species (P < 0.01), NOD-like receptor family pyrin domain containing 3 (P < 0.05), cleaved-caspase1 (P < 0.05), tumor necrosis factor-α (P < 0.05), and interleukin-1β (P < 0.05), but increased glutathione (P < 0.05), mitochondrial membrane potential (P < 0.05), phosphorylation (p)-JAK2 (P < 0.05), and p-STAT3 (P < 0.01) in MPP+ treated BV-2 cells, which indicated that VSIG4 inhibited oxidative stress, mitochondrial dysfunction, and inflammation, as well as activated the JAK2/STAT3 pathway in PD model. Moreover, AZD1480 inhibited the JAK2/STAT3 pathway and aggravated oxidative stress, mitochondrial dysfunction, and inflammation in PD model (all P < 0.05). Importantly, AZD1480 attenuated the influence of VSIG4 on oxidative stress, mitochondrial dysfunction, inflammation, and the JAK2/STAT3 pathway in PD model (all P < 0.05). ConclusionVSIG4 suppresses oxidative stress, mitochondrial dysfunction, and inflammation by activating the JAK2/STAT3 pathway, which may be helpful in attenuating PD progression.

Jianpi Gushen Huayu decoction ameliorated diabetic nephropathy through modulating metabolites in kidney, and inhibiting TLR4/NF-κB/NLRP3 and JNK/P38 pathways.

Jianpi Gushen Huayu Decoction (JPGS) has been used to clinically treat diabetic nephropathy (DN) for many years. However, the protective mechanism of JPGS in treating DN remains unclear. To evaluate the therapeutic effects and the possible mechanism of JPGS on DN. We first evaluated the therapeutic potential of JPGS on a DN mouse model. We then investigated the effect of JPGS on the renal metabolite levels of DN mice using non-targeted metabolomics. Furthermore, we examined the effects of JPGS on c-Jun N-terminal kinase (JNK)/P38-mediated apoptosis and the inflammatory responses mediated by toll-like receptor 4 (TLR4)/nuclear factor-kappa B (NF-κB)/NOD-like receptor family pyrin domain containing 3 (NLRP3). The ameliorative effects of JPGS on DN mice included the alleviation of renal injury and the control of inflammation and oxidative stress. Untargeted metabolomic analysis revealed that JPGS altered the metabolites of the kidneys in DN mice. A total of 51 differential metabolites were screened. Pathway analysis results indicated that nine pathways significantly changed between the control and model groups, while six pathways significantly altered between the model and JPGS groups. Pathways related to cysteine and methionine metabolism; alanine, tryptophan metabolism; aspartate and glutamate metabolism; and riboflavin metabolism were identified as the key pathways through which JPGS affects DN. Further experimental validation showed that JPGS treatment reduced the expression of TLR4/NF-κB/NLRP3 pathways and JNK/P38 pathway-mediated apoptosis related factors. JPGS could markedly treat mice with streptozotocin (STZ)-induced DN, which is possibly related to the regulation of several metabolic pathways found in kidneys. Furthermore, JPGS could improve kidney inflammatory responses and ameliorate kidney injuries in DN mice via the TLR4/NF-κB/NLRP3 pathway and inhibit JNK/P38 pathway-mediated apoptosis in DN mice.

DT-13 attenuates inflammation by inhibiting NLRP3-inflammasome related genes in RAW264.7 macrophages

Plant derived saponins or other glycosides are widely used for their anti-inflammatory, antioxidant, and anti-viral properties in therapeutic medicine. In this study, we focus on understanding the function of the less known steroidal saponin from the roots of Liriope muscari L. H. Bailey – saponin C (also known as DT-13) in lipopolysaccharide (LPS)-stimulated RAW264.7 macrophages in comparison to the well-known saponin ginsenoside Rk1 and anti-inflammatory drug dexamethasone. We proved that DT-13 reduces LPS-induced inflammation by inhibiting nitric oxide (NO) production, interleukin-6 (IL-6) release, cycloxygenase-2 (COX-2), tumour necrosis factor-alpha (TNF-α) gene expression, and nuclear factor kappa-B (NFκB) translocation into the nucleus. It also inhibits the inflammasome component NOD-like receptor family pyrin domain containing protein 3 (NLRP3) regulating the inflammasome activation. This was supported by the significant inhibition of caspase-1 and interleukin-1 beta (IL-1β) expression and release. This study demonstrates the anti-inflammatory effect of saponins on LPS-stimulated macrophages. For the first time, an in vitro study shows the attenuating effect of DT-13 on NLRP3-inflammasome activation. In comparison to the existing anti-inflammatory drug, dexamethasone, and triterpenoid saponin Rk1, DT-13 more efficiently inhibits inflammation in the applied cell culture model. Therefore, DT-13 may serve as a lead compound for the development of new more effective anti-inflammatory drugs with minimised side effects.

FGF21 overexpression alleviates VSMC senescence in diabetic mice by modulating the SYK-NLRP3 inflammasome-PPARγ-catalase pathway.

Diabetes accelerates vascular senescence, which is the basis for atherosclerosis and stiffness. The activation of the NOD-like receptor family pyrin domain containing 3 (NLRP3) inflammasome and oxidative stress are closely associated with progressive senescence in vascular smooth muscle cells (VSMCs). The vascular protective effect of FGF21 has gradually gained increasing attention, but its role in diabetes-induced vascular senescence needs further investigation. In this study, diabetic mice and primary VSMCs are transfected with an FGF21 activation plasmid and treated with a peroxisome proliferator-activated receptor γ (PPARγ) agonist (rosiglitazone), an NLRP3 inhibitor (MCC950), and a spleen tyrosine kinase (SYK)-specific inhibitor, R406, to detect senescence-associated markers. We find that FGF21 overexpression significantly restores the level of catalase (CAT), vascular relaxation, inhibits the intensity of ROSgreen fluorescence and p21 immunofluorescence, and reduces the area of SA-β-gal staining and collagen deposition in the aortas of diabetic mice. FGF21 overexpression restores CAT, inhibits the expression of p21, and limits the area of SA-β-gal staining in VSMCs under high glucose conditions. Mechanistically, FGF21 inhibits SYK phosphorylation, the production of the NLRP3 dimer, the expression of NLRP3, and the colocalization of NLRP3 with PYCARD (ASC), as well as NLRP3 with caspase-1, to reverse the cleavage of PPARγ, preserve CAT levels, suppress ROSgreen density, and reduce the expression of p21 in VSMCs under high glucose conditions. Our results suggest that FGF21 alleviates vascular senescence by regulating the SYK-NLRP3 inflammasome-PPARγ-catalase pathway in diabetic mice.

Gegen Qinlian decoction ameliorates TNBS-induced ulcerative colitis by regulating Th2/Th1 and Tregs/Th17 cells balance, inhibiting NLRP3 inflammasome activation and reshaping gut microbiota

Ethnopharmacological relevanceChinese herbal medicine Gegen Qinlian Decoction (GQD) has been clinically shown to be an effective treatment of ulcerative colitis (UC) in China. However, the underlying mechanism of GQD's anti-ulcerative colitis properties and its effect on gut microbiota still deserve further exploration. Aim of the studyThis study observed the regulatory effects of GQD on Th2/Th1 and Tregs/Th17 cells balance, the NOD-like receptor family pyrin domain containing 3 (NLRP3) infammasome and gut microbiota in TNBS-induced UC in BALB/c mice. Materials and methods61 main chemical compounds in the GQD were determined by UPLC-Q-TOF/MS. The UC BALB/c model was established by intrarectal administration of trinitrobenzene sulfonic acid (TNBS), and GQD was orally administered at low and high dosages of 2.96 and 11.83 g/kg/day, respectively. The anti-inflammatory effects of GQD for ulcerative colitis were evaluated by survival rate, body weight, disease activity index (DAI) score, colonic weight and index, spleen index, hematoxylin-eosin (HE) staining and histopathological scores. Flow cytometry was used to detect the percentage of CD4, Th1, Th2, Th17 and Tregs cells. The levels of Th1-/Th2-/Th17-/Tregs-related inflammatory cytokines and additional proinflammatory cytokines (IL-1β, IL-18) were detected by CBA, ELISA, and RT-PCR. The expressions of GATA3, T-bet, NLRP3, Caspase-1, IL-Iβ, Occludin and Zonula occludens-1 (ZO-1) on colon tissues were detected by Western blot and RT-PCR. Transcriptome sequencing was performed using colon tissue and 16S rRNA gene sequencing was performed on intestinal contents. Fecal microbiota transplantation (FMT) was employed to assess the contribution of intestinal microbiota and its correlation with CD4 T cells and the NLRP3 inflammasome. ResultsGQD increased the survival rate of TNBS-induced UC in BALB/c mice, and significantly improved their body weight, DAI score, colonic weight and index, spleen index, and histological characteristics. The intestinal barrier dysfunction was repaired after GQD administration through promoting the expression of tight junction proteins (Occludin and ZO-1). GQD restored the balance of Th2/Th1 and Tregs/Th17 cells immune response of colitis mice, primarily inhibiting the increase in Th2/Th1 ratio and their transcription factor production (GATA3 and T-bet). Morever, GQD changed the secretion of Th1-/Th2-/Th17-/Tregs-related cytokines (IL-2, IL-12, IL-5, IL-13, IL-6, IL-10, and IL-17A) and reduced the expressions of IL-1β, IL-18. Transcriptome results suggested that GQD could also remodel the immune inflammatory response of colitis by inhibiting NOD-like receptor signaling pathway, and Western blot, immunohistochemistry and RT-PCR further revealed that GQD exerted anti-inflammatory effects by inhibiting the NLRP3 inflammasome, such as down-regulating the expression of NLRP3, Caspase-1 and IL-1β. More interestingly, GQD regulated gut microbiota dysbiosis, suppressed the overgrowth of conditional pathogenic gut bacteria like Helicobacter, Proteobacteria, and Mucispirillum, while the probiotic gut microbiota, such as Lactobacillus, Muribaculaceae, Ruminiclostridium_6, Akkermansia, and Ruminococcaceae_unclassified were increased. We further confirmed that GQD-treated gut microbiota was sufficient to relieve TNBS-induced colitis by FMT, involving the modulation of Th2/Th1 and Tregs/Th17 balance, inhibition of NLRP3 inflammasome activation, and enhancement of colonic barrier function. ConclusionsGQD might alleviate TNBS-induced UC via regulating Th2/Th1 and Tregs/Th17 cells Balance, inhibiting NLRP3 inflammasome and reshaping gut microbiota, which may provide a novel strategy for patients with colitis.

Demotion of canonical/non-canonical inflammasome and pyroptosis alleviates ischemia/reperfusion-induced acute kidney injury: Novel role of the D2/D3 receptor agonist ropinirole

Ropinirole used to treat Parkinson's disease highly targets the dopaminergic receptor D3 over the D2 receptor but although both are expressed in the kidneys the ropinirole potential to treat kidney injury provoked by ischemia/reperfusion (I/R) is undraped. We investigated whether ropinirole can alleviate renal I/R by studying its anti-inflammatory, antioxidant, and anti-pyroptotic effects targeting its aptitude to inhibit the High-mobility group box 1/Toll-like receptor 4/Nuclear factor-kappa B (HMGB1/TLR4/NF-κB) cue and the canonical/non-canonical NOD-like receptor family pyrin domain containing 3 (NLRP3) inflammasome trajectories. Herein, bilateral I/R surgery was induced in animals to be either untreated or treated with ropinirole for three days after the insult. Ropinirole successfully improved the histopathological picture and renal function which was confirmed by reducing cystatin C and the standard parameters creatinine and blood urea nitrogen (BUN). Ropinirole achieved this through its anti-inflammatory capacity mediated by reducing the HMGB1/TLR4 axis and inactivating NF-κB, which are upstream regulators of the NLRP3 pathway. As a result, the injurious inflammasome markers (NLRP3, apoptosis-associated speck-like protein (ASC), active caspase-1) and their target cytokines interleukin-1 beta (IL-1β) and IL-18 were decreased. Ropinirole also reduced the pyroptotic cell death markers caspase-11 and gasdermin-D. Furthermore, ropinirole by replenishing antioxidants and decreasing malondialdehyde helped to reduce oxidative stress in the kidneys. The docking findings confirmed that ropinirole highly binds to the dopaminergic D3 receptor more than to the D2 receptor. In conclusion, ropinirole has the potential to be a reno-therapeutic treatment against I/R insult by abating the inflammatory NLRP3 inflammasome signal, pyroptosis, and oxidative stress.

The Mechanism of Astragaloside IV in NOD-like Receptor Family Pyrin Domain Containing 3 Inflammasome-mediated Pyroptosis after Intracerebral Hemorrhage.

Intracerebral hemorrhage (ICH) is one of the most common subtypes of stroke. This study aimed to investigate the mechanism of Astragaloside IV (AS-IV) on inflammatory injury after ICH. The ICH model was established by the injection of collagenase and treated with ASIV (20 mg/kg or 40 mg/kg). The neurological function, water content of the bilateral cerebral hemisphere and cerebellum, and pathological changes in brain tissue were assessed. The levels of interleukin-1 beta (IL-1β), IL-18, tumor necrosis factor-alpha, interferon-gamma, and IL-10 were detected by enzyme-linked immunosorbent assay. The levels of Kruppel-like factor 2 (KLF2), NOD-like receptor family pyrin domain containing 3 (NLRP3), GSDMD-N, and cleaved-caspase-1 were detected by reverse transcription-quantitative polymerase chain reaction and Western blot assay. The binding relationship between KLF2 and NLRP3 was verified by chromatin-immunoprecipitation and dual-luciferase assays. KLF2 inhibition or NLRP3 overexpression was achieved in mice to observe pathological changes. The decreased neurological function, increased water content, severe pathological damage, and inflammatory response were observed in mice after ICH, with increased levels of NLRP3/GSDMD-N/cleaved-caspase-1/IL-1β/IL-18 and poorly-expressed KLF2 in brain tissue. After AS-IV treatment, the neurological dysfunction, high brain water content, inflammatory response, and pyroptosis were alleviated, while KLF2 expression was increased. KLF2 bonded to the NLRP3 promoter region and inhibited its transcription. Down-regulation of KLF2 or upregulation of NLRP3 reversed the effect of AS-IV on inhibiting pyroptosis and reducing inflammatory injury in mice after ICH. AS-IV inhibited NLRP3-mediated pyroptosis by promoting KLF2 expression and alleviated inflammatory injury in mice after ICH.

Ginsenoside Rb1 alleviates 3-MCPD-induced renal cell pyroptosis by activating mitophagy

Ginsenoside Rb1 (Gs-Rb1) is among the most significant effective pharmacological components in ginseng. 3-monochloropropane-1,2-diol (3-MCPD), a chloropropanol-like contaminant, is produced in the production of refined oils and thermal processing of food. Pyroptosis is a type of programmed cell death triggered by inflammasomes. Excessive pyroptosis causes kidney injury and inflammation. Previous studies have revealed that 3-MCPD induced pyroptosis in mice and NRK-52E cells. In the present study, we find that Gs-Rb1 attenuates 3-MCPD-induced renal cell pyroptosis by assaying GSDMD-N, caspase-1, IL-18, and IL-1β in mice and NRK-52E cells. In further mechanistic studies, we show that Gs-Rb1 removes damaged mitochondria via mitophagy and reduces intracellular reactive oxygen species (ROS) generation, therefore alleviating 3-MCPD-induced NOD-like receptor family pyrin domain containing 3 (NLRP3) activation and pyroptosis. The above results are further validated by the addition of autophagy inhibitor Chloroquine (CQ) and mitophagy inhibitor Cyclosporin A (CsA). Afterward, we explore how Gs-Rb1 activated mitophagy in vitro. We determine that Gs-Rb1 enhances the protein expression and nuclear translocation of Transcription factor EB (TFEB). However, silencing of the TFEB gene by small interfering RNA technology reverses the role of Gs-Rb1 in activating mitophagy. Therefore, we conclude that 3-MCPD damages mitochondria and leads to ROS accumulation, which causes NLRP3 activation and pyroptosis in ICR mice and NRK-52E cells, while Gs-Rb1 mitigates this phenomenon via the TFEB-mitophagy pathway. Our findings may provide new insights for understanding the molecular mechanisms by which Gs-Rb1 mitigates renal injury.

Emerging role of exosomes in ulcerative colitis: Targeting NOD-like receptor family pyrin domain containing 3 inflammasome.

Ulcerative colitis (UC) is a chronic recurrent inflammatory bowel disease. Despite ongoing advances in our understanding of UC, its pathogenesis is yet unelucidated, underscoring the urgent need for novel treatment strategies for patients with UC. Exosomes are nanoscale membrane particles that mediate intercellular communication by carrying various bioactive molecules, such as proteins, RNAs, DNA, and metabolites. The NOD-like receptor family pyrin domain containing 3 (NLRP3) inflammasome is a cytosolic tripartite protein complex whose activation induces the maturation and secretion of proinflammatory cytokines interleukin-1β (IL-1β) and IL-18, triggering the inflammatory response to a pathogenic agent or injury. Growing evidence suggests that exosomes are new modulators of the NLRP3 inflammasome, with vital roles in the pathological process of UC. Here, recent evidence is reviewed on the role of exosomes and NLRP3 inflammasome in UC. First, the dual role of exosomes on NLRP3 inflammasome and the effect of NLRP3 inflammasome on exosome secretion are summarized. Finally, an outlook on the directions of exosome-NLRP3 inflammasome crosstalk research in the context of UC is proposed and areas of further research on this topic are highlighted.

Imeglimin Exhibits Novel Anti-Inflammatory Effects on High-Glucose-Stimulated Mouse Microglia through ULK1-Mediated Suppression of the TXNIP-NLRP3 Axis.

Type 2 diabetes mellitus (T2DM) is an epidemiological risk factor for dementia and has been implicated in multifactorial pathologies, including neuroinflammation. In the present study, we aimed to elucidate the potential anti-inflammatory effects of imeglimin, a novel antidiabetic agent, on high-glucose (HG)-stimulated microglia. Mouse microglial BV2 cells were stimulated with HG in the presence or absence of imeglimin. We examined the effects of imeglimin on the levels of proinflammatory cytokines, intracellular reactive oxygen species (ROS), mitochondrial integrity, and components related to the inflammasome or autophagy pathways in these cells. Our results showed that imeglimin suppressed the HG-induced production of interleukin-1beta (IL-1β) by reducing the intracellular ROS levels, ameliorating mitochondrial dysfunction, and inhibiting the activation of the thioredoxin-interacting protein (TXNIP)-NOD-like receptor family pyrin domain containing 3 (NLRP3) axis. Moreover, the inhibitory effects of imeglimin on the TXNIP-NLRP3 axis depended on the imeglimin-induced activation of ULK1, which also exhibited novel anti-inflammatory effects without autophagy induction. These findings suggest that imeglimin exerted novel suppressive effects on HG-stimulated microglia through the ULK1-TXNIP-NLRP3 axis, and may, thereby, contribute to the development of innovative strategies to prevent T2DM-associated cognitive impairment.

Effect of tissue factor pathway inhibitor on the pyroptosis of vascular smooth muscle cells induced by angiotensin II.

In recent years, a mass of studies have shown that pyroptosis plays an important role in the proliferation of vascular smooth muscle cells (VSMCs). We investigated whether angiotensin II (Ang II) induces the pyroptosis of rat aortic VSMCs and the role of NOD-like receptor family pyrin domain containing 3 (NLRP3) in this process. Additionally, we explored the effect and related mechanism of recombinant tissue factor pathway inhibitor (rTFPI) in Ang II-induced VSMC pyroptosis. Cultured VSMCs were divided into five groups: control group, Ang II group (1×10-5 mol/L), MCC950 group (NLRP3 inhibitor, 15 nmol/L), Ang II + MCC950 group and Ang II + rTFPI (50 µg/L) group. Cell viability was measured by cell counting kit-8 (CCK8) assays and 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assays. Propidium iodide (PI) staining and immunofluorescence were performed to determine the pyroptosis of VSMCs. Changes in VSMC ultrastructure were evaluated through transmission electron microscopy. The expression levels of NLRP3, pro-caspase-1, gasdermin D-N (GSDMD-N), and interleukin-1β (IL-1β) were determined by western blot analysis. The cell viability, the positive rate of PI staining, and the expression level of GSDMD detected by immunofluorescence in the Ang II group were higher than that in the control group, whereas they all decreased in Ang II + MCC950 group and Ang II + rTFPI group compared with Ang II group (P<0.05). Electron microscopy analysis revealed less extracellular matrix, increased myofilaments, and decreased endoplasmic reticulum, Golgi complex, and mitochondria in Ang II + rTFPI-treated VSMCs than in Ang II-treated VSMCs. The protein expression levels of the pyroptosis-related molecules NLRP3, pro-caspase-1, GSDMD-N, and IL-1β in Ang II group showed an increasing trend compared with those in control group (P<0.05); however, these expression levels in Ang II + MCC950 and Ang II + rTFPI groups were significantly lower than those in Ang II group (P<0.05). Ang II may induce pyroptosis in VSMCs by activating NLRP3. rTFPI can inhibit Ang II-induced VSMC pyroptosis. Furthermore, rTFPI might exert this effect by inhibiting the NLRP3 pathway and therefore play an important role in the treatment of vascular remodeling induced by hypertension.

TRIM59 suppresses the brain ischaemia/reperfusion injury and pyroptosis of microglial through mediating the ubiquitination of NLRP3

Cerebral ischaemia/reperfusion (I/R) injury induces irreversible brain injury and causes functional impairment. Ubiquitination plays a crucial role in protein degradation, but its role in cerebral I/R injury remains unclear. Differentially expressed genes in stroke were identified by analysing the microarray dataset GSE119121. Cerebral I/R was simulated in vitro by treating human microglial HMC3 cells with oxygen–glucose deprivation/reperfusion (OGD/R). Cell viability was tested by Cell Counting Kit 8 (CCK-8) assays, and pyroptosis was examined by flow cytometry. Lactate dehydrogenase (LDH) and inflammatory cytokine secretion were measured by LDH cytotoxicity assays and enzyme-linked immunosorbent assay (ELISA), respectively. The cerebral I/R animal model was established by middle cerebral artery occlusion (MCAO) surgery in rats. Bioinformatic analysis indicated that tripartite motif-containing protein 59 (TRIM59) is downregulated in stroke, which was verified in cerebral I/R models. The upregulation of TRIM59 promoted viability and inhibited pyroptosis in OGD/R-treated microglia and alleviated cerebral I/R injury in vivo. TRIM59 attenuated NOD-like receptor family pyrin domain containing 3 (NLRP3) protein expression through ubiquitination, thus degrading NLRP3 and alleviating OGD/R-induced injury. TRIM59 relieves cerebral I/R injury in vivo and in vivo. Mechanistically, TRIM59 directly interacts with NLRP3 and inhibits NLRP3 through ubiquitination. Targeting the TRIM59/NLRP3 signalling axis may be an effective therapeutic strategy for cerebral I/R.

A mouse model to distinguish NLRP6-mediated inflammasome-dependent and -independent functions

The NOD-like receptor (NLR) family pyrin domain containing 6 (NLRP6) serves as a sensor for microbial dsRNA or lipoteichoic acid (LTA) in intestinal epithelial cells (IECs), and initiating multiple pathways including inflammasome pathway and type I interferon (IFN) pathway, or regulating nuclear factor-κB (NF-κB) and mitogen-activated protein kinase (MAPK) pathways. NLRP6 can exert its function in both inflammasome-dependent and inflammasome-independent manners. However, there is no tool to distinguish the contribution of individual NLRP6-mediated pathway to the physiology and pathology in vivo. Here, we validated that Arg39 and Trp50 residues in the pyrin domain (PYD) of murine NLRP6 are required for ASC recruitment and inflammasome activation, but are not important for the RNA binding and PYD-independent NLRP6 oligomerization. We further generated the Nlrp6R39E&W50E mutant mice, which showed reduced inflammasome activation in either steady state intestine or during viral infection. However, the type I IFN production in cells or intestine tissue from Nlrp6R39E&W50E mutant mice remain normal. Interestingly, NLRP6-mediated inflammasome activation or the IFN-I production seems to play distinct roles in the defense responses against different types of RNA viruses. Our work generated a useful tool to study the inflammasome-dependent role of NLRP6 in vivo, which might help to understand the complexity of multiple pathways mediated by NLRP6 in response to the complicated and dynamic environmental cues in the intestine.

3'-Deoxyadenosin alleviates methamphetamine-induced aberrant synaptic plasticity and seeking behavior by inhibiting the NLRP3 inflammasome.

JOURNAL/nrgr/04.03/01300535-202410000-00028/figure1/v/2024-02-06T055622Z/r/image-tiff Methamphetamine addiction is a brain disorder characterized by persistent drug-seeking behavior, which has been linked with aberrant synaptic plasticity. An increasing body of evidence suggests that aberrant synaptic plasticity is associated with the activation of the NOD-like receptor family pyrin domain containing-3 (NLRP3) inflammasome. 3'-Deoxyadenosin, an active component of the Chinese fungus Cordyceps militaris, has strong anti-inflammatory effects. However, whether 3'-deoxyadenosin attenuates methamphetamine-induced aberrant synaptic plasticity via an NLRP3-mediated inflammatory mechanism remains unclear. We first observed that 3'-deoxyadenosin attenuated conditioned place preference scores in methamphetamine-treated mice and decreased the expression of c-fos in hippocampal neurons. Furthermore, we found that 3'-deoxyadenosin reduced the aberrant potentiation of glutamatergic transmission and restored the methamphetamine-induced impairment of synaptic plasticity. We also found that 3'-deoxyadenosin decreased the expression of NLRP3 and neuronal injury. Importantly, a direct NLRP3 deficiency reduced methamphetamine-induced seeking behavior, attenuated the impaired synaptic plasticity, and prevented neuronal damage. Finally, NLRP3 activation reversed the effect of 3'-deoxyadenosin on behavior and synaptic plasticity, suggesting that the anti-neuroinflammatory mechanism of 3'-deoxyadenosin on aberrant synaptic plasticity reduces methamphetamine-induced seeking behavior. Taken together, 3'-deoxyadenosin alleviates methamphetamine-induced aberrant synaptic plasticity and seeking behavior by inhibiting the NLRP3 inflammasome.