Reducing NLRP3 Inflammasome Activation Research Articles

Nicardipine-chitosan nanoparticles alleviate thioacetamide-induced acute liver injury by targeting NFκB/NLRP3/IL-1β signaling in rats: Unraveling new roles beyond calcium channel blocking

Liver inflammatory diseases are marked by serious complications. Notably, nicardipine (NCD) has demonstrated anti-inflammatory properties, but its benefits in liver inflammation have not been studied yet. However, the therapeutic efficacy of NCD is limited by its short half-life and low bioavailability. Therefore, we aimed to evaluate the potential of NCD-loaded chitosan nanoparticles (ChNPs) to improve its pharmacokinetic profile and hepatic accumulation. Four formulations of NCD-ChNPs were synthesized and characterized. The optimal formulation (NP2) exhibited a mean particle diameter of 172.6 ± 1.94 nm, a surface charge of +25.66 ± 0.93 mV, and an encapsulation efficiency of 88.86 ± 1.17 %. NP2 showed good physical stability as a lyophilized powder over three months. It displayed pH-sensitive release characteristics, releasing 77.15 ± 5.09 % of NCD at pH 6 (mimicking the inflammatory microenvironment) and 52.15 ± 3.65 % at pH 7.4, indicating targeted release in inflamed liver tissues. Pharmacokinetic and biodistribution studies revealed that NCD-ChNPs significantly prolonged NCD circulation time and enhanced its concentration in liver tissues compared to plain NCD. Additionally, the study investigated the protective effects of NCD-ChNPs in thioacetamide-induced liver injury in rats by modulating the NFκB/NLRP3/IL-1β signaling axis. NCD-ChNPs effectively inhibited NFκB activation, reduced NLRP3 inflammasome activation, and subsequent release of IL-1β, which correlated with improved hepatic function and reduced inflammation and oxidative stress. These findings highlight the potential of NCD-ChNPs as a promising nanomedicine strategy for the treatment of liver inflammatory diseases, warranting further investigation into their clinical applications, particularly in hypertensive patients with liver inflammatory conditions.

Dental pulp mesenchymal stem cell-derived exosomes inhibit neuroinflammation and microglial pyroptosis in subarachnoid hemorrhage via the miRNA-197-3p/FOXO3 axis

BackgroundSubarachnoid hemorrhage (SAH) is a severe stroke subtype that lacks effective treatment. Exosomes derived from human dental pulp stem cells (DPSCs) are a promising acellular therapeutic strategy for neurological diseases. However, the therapeutic effects of DPSC-derived exosomes (DPSC-Exos) on SAH remain unknown. In this study, we investigated the therapeutic effects and mechanisms of action of DPSC-Exos in SAH.Materials and methodsSAH was established using 120 male Sprague-Dawley rats. One hour after SAH induction, DPSC-Exos were administered via tail vein injection. To investigate the effect of DPSC-Exos, SAH grading, short-term and long-term neurobehavioral assessments, brain water content, western blot (WB), immunofluorescence staining, Nissl staining, and HE staining were performed. The role of miR-197-3p/FOXO3 in regulating pyroptosis was demonstrated through miRNA sequencing, bioinformatics analysis, and rescue experiments. The SAH model in vitro was established by stimulating BV2 cells with hemoglobin (Hb) and the underlying mechanism of DPSC-Exos was investigated through WB and Hoechst/PI staining.ResultsThe expressions of pro-inflammatory cytokines (IL-1β, IL-6, and TNF-α) were increased after SAH. DPSC-Exos alleviated brain edema and neuroinflammation by inhibiting the expression of FOXO3 and reducing NLRP3 inflammasome activation, leading to improved neurobehavioral functions at 24 h after SAH. In vitro, the expression of the NLRP3 inflammasome components (NLRP3 and caspase1-p20), GSDMD-N, and IL-18 was inhibited in BV2 cells pretreated with DPSC-Exos. Importantly, DPSC-Exos overexpressing miR-197-3p had a more obvious protective effect than those from NC-transfected DPSCs, while those from DPSCs transfected with the miR-197-3p inhibitor had a weaker protective effect. Functional studies indicated that miR-197-3p bound to the 3ʹ-untranslated region of FOXO3, inhibiting its transcription. Furthermore, the overexpression of FOXO3 reversed the protective effects of miR-197-3p.ConclusionsDPSC-Exos inhibited activation of the NLRP3 inflammasome and related cytokine release via the miR-197-3p/FOXO3 pathway, alleviated neuroinflammation, and inhibited microglial pyroptosis. These findings suggest that using DPSC-Exos is a promising therapeutic strategy for SAH.Graphical

Resveratrol ameliorates glioblastoma inflammatory response by reducing NLRP3 inflammasome activation through inhibition of the JAK2/STAT3 pathway

ObjectivesThe aim of this study was to investigate the anti-tumor effect of resveratrol (RSV) on glioblastoma (GBM) and its specific mechanism in improving the inflammatory response of the tumor microenvironment. The tumor microenvironment of GBM is highly neuroinflammatory, inducing tumor immunosuppression. Therefore, ameliorating the inflammatory response is an important focus for anti-tumor research.MethodsThe anti-tumor effect of RSV on GBM was demonstrated through in vitro cellular assays, including CCK-8, EdU, PI staining, Transwell, wound healing assay, and flow cytometry. Potential mechanisms of RSV's anti-GBM effects were identified through network pharmacological analysis. In addition, the relationship of RSV with the JAK2/STAT3 signaling pathway and the inflammasome NLRP3 was verified using Western blot.ResultsRSV significantly inhibited cell viability in GBM cell lines LN-229 and U87-MG. Furthermore, it inhibited the proliferation and invasive migration ability of GBM cells, while promoting apoptosis. Network pharmacological analysis revealed a close association between the anti-GBM effects of RSV and the JAK/STAT signaling pathway, as well as inflammatory responses. Western blot analysis confirmed that RSV inhibited the over-activation of the inflammasome NLRP3 through the JAK2/STAT3 signaling pathway. Partial reversal of RSV's inhibition of inflammasome NLRP3 was observed with the addition of the JAK/STAT agonist RO8191.ConclusionsIn vitro, RSV can exert anti-tumor effects on GBM and improve the inflammatory response in the GBM microenvironment by inhibiting the activation of the JAK2/STAT3 signaling pathway. These findings provide new insights into potential therapeutic targets for GBM.

Antagonizing interleukin-5 receptor ameliorates dextran sulfate sodium-induced experimental colitis in mice through reducing NLRP3 inflammasome activation

Inflammatory bowel disease (IBD) is a condition characterized by inflammation in the gastrointestinal tract. Reducing intestinal inflammation is a promising approach for treating IBD. The nucleotide-binding oligomerization domain-like receptor family pyrin domain-containing protein 3 (NLRP3) inflammasome, a critical component of the innate immune system, is implicated in the pathogenesis of IBD. Therefore, inhibiting NLRP3 inflammasome activation is a potential therapeutic strategy for IBD. In this study, we investigated the effects of the interleukin-5 (IL-5) receptor antagonist YM-90709 on dextran sulfate sodium-induced experimental colitis in mice. We found that YM-90709 reduced the expressions of IL-1β and caspase-1 p20 in the colon and ameliorated colitis. Furthermore, we identified YM-90709 as an effective agent for inhibiting NLRP3 inflammasome activation. Knockdown of IL-5 receptor or using an inhibitor of STAT5, a key transcription factor downstream of the IL-5/IL-5 receptor signal pathway, also reduced NLRP3 inflammasome-dependent IL-1β release and ASC speck formation. Our study is the first to demonstrate that the NLRP3 inflammasome may be a downstream signal of IL-5/IL-5 receptor and that YM-90709 protects against IBD by inhibiting IL-5 receptor. These findings suggest a new strategy for regulating intestinal inflammation and managing IBD.

Roles of HDAC6 in the pathogenesis of Alzheimer’s disease

AbstractBackgroundIt has recently been demonstrated that HDAC6 depletion ameliorates the impairment of memory function and mitochondrial axonal transport induced by the accumulation of Aβ. However, very little is known about molecular mechanisms through which HDAC6 regulates APP processing. Also, it remains controversial whether HDAC6 induces inflammation or reduces it.MethodWe analyzed the expression of HDAC6 in AD patients and 5xFAD mice. We confirmed that HDAC6 regulates BACE1 in primary neurons and NLRP3 inflammasome in primary microglia. And we crossed HDAC6 knock‐out mice with 5xFAD mice and got 5xFAD/HDAC6 KO. We compared 5xFAD (n = 11) and 5xFAD/HDAC6 KO (n = 13). We did a nest building test, open field test, Morris water maze, Y‐maze, passive avoidance, and elevated plus maze. After finishing all these behaviour tests, we sacrificed the mouse and checked protein and mRNA. We did Immunohistochemistry using Aβ plaque, NLRP3 inflammasome and synapse‐related markers. We also did RNA‐seq analysis in these mice’s brains.ResultWe confirmed that the expression of HDAC6 was increased in the AD cortex compared to neuropathologically normal brains. The stability of BACE1 is regulated by its C‐terminal lysine, and HDAC6 deacetylates the C‐terminal lysine of BACE1 by direct binding. Also, HDAC6 deficiency reduced NLRP3 inflammasome activation‐, and subsequent inflammatory responses in microglia. By crossing HDAC6 knock‐out mice with 5xFAD mice, the role of HDAC6 was confirmed in an animal model of AD. HDAC6 deletion improved cognitive function and reduced the protein expression of BACE1 and inflammatory factors in 5xFAD mice. Also, HDAC6‐deficient 5xFAD mice developed fewer dense‐core plaques, IL‐1β, and ASC specks than 5xFAD mice. Furthermore, RNA‐sequencing was analyzed from the hippocampus of 5xFAD and 5xFAD/HDAC6 KO mice. Synapse and neuron development‐related genes were significantly up‐regulated, and type I interferon, immune response, and prion disease‐related genes were down‐regulated in HDAC6‐depleted mice. Finally, we confirmed that the ratio of PSD95 colocalized with C1q was significantly decreased in 5xFAD/HDAC6 KO compared to 5xFAD mice, which suggested that synapse loss was inhibited in AD mice.ConclusionThis study reveals distinct functions of HDAC6 and suggests HDAC6 as a significant regulator in the pathogenesis of AD that could be exploited therapeutically.

Glibenclamide-Loaded Nanoparticles Reduce NLRP3 Inflammasome Activation and Modulate miR-223-3p/miR-7-1-5p Expression in THP-1 Cells.

The anti-hyperglycemic drug glibenclamide (Glb) might represent an interesting therapeutic option in human neurodegenerative diseases because of its anti-inflammatory activity and its ability to downregulate activation of the NLRP3 inflammasome. Bi-functionalized liposomes that can cross the blood-brain barrier (BBB) may be used to release Glb into the central nervous system (CNS), overcoming its poor solubility and bioavailability. Here, we analyzed in vitro the effect of Glb-loaded nanovectors (GNVs) and Glb itself on NLRP3 inflammasome activation using a lipopolysaccharide- and nigericine-activated THP-1 cell model. Apoptosis-associated speck-like protein containing a CARD (ASC) aggregation and NLRP3-related cytokine (IL-1β, caspase 1, and IL-18) production and gene expression, as well as the concentration of miR-223-3p and miR-7-1-5p, known to modulate the NLRP3 inflammasome, were evaluated in all conditions. Results showed that both GNVs and Glb reduced significantly ASC-speck oligomerization, transcription and translation of NLRP3, as well as the secretion of caspase 1 and IL-1β (p < 0.05 for all). Unexpectedly, GNVs/Glb significantly suppressed miR-223-3p and upregulated miR-7-1-5p expression (p < 0.01). These preliminary results thus suggest that GNVs, similarly to Glb, are able to dampen NLRP3 inflammasome activation, inflammatory cytokine release, and modulate miR-223-3p/miR-7-1-5p. Although the mechanisms underlying the complex relation among these elements remain to be further investigated, these results can open new roads to the use of GNVs as a novel strategy to reduce inflammasome activation in disease and rehabilitation.

The STING inhibitor C-176 attenuates MPTP-induced neuroinflammation and neurodegeneration in mouse parkinsonian models

Recent emerging evidence reveals that cGAS-STING-mediated Type I interferon (IFN) signaling axis takes part in the microglial-associated neuroinflammation. However, the potential role of pharmacological inhibition of STING on neuroinflammation and dopaminergic neurodegeneration remains unknown. In the present study, we investigated whether pharmacological inhibition of STING attenuates neuroinflammation and neurodegeneration in experimental models of Parkinson’s disease. We report that therapeutic inhibition of STING with C-176 significantly inhibited the activation of downstream signaling pathway, suppressed neuroinflammation, and ameliorated MPTP-induced dopaminergic neurotoxicity and motor deficit. Furthermore, pharmacological inhibition of STING with C-176 attenuated proinflammatory response in BV2 microglial cells exposed to LPS/MPP+. More importantly, C-176 also reduced NLRP3 inflammasome activation both in vitro and in vivo. The results of our study suggest that pharmacologic inhibition of STING protects against dopaminergic neurodegeneration and neuroinflammation that may act at least in part through suppressing NLRP3 inflammasome activation. STING signaling may hold great promise for the development of new treatment strategy for PD.

Inhibitors of NLRP3 Inflammasome in Ischemic Heart Disease: Focus on Functional and Redox Aspects.

Myocardial ischemia-reperfusion injury (MIRI) is caused by several mechanisms, including the production of reactive oxygen species (ROS), altered cellular osmolarity, and inflammatory response. Calcium overload, altered oxygen levels, and mitochondrial ROS are also involved in these MIRI processes, resulting in the irreversible opening of the mitochondrial permeability transition pore (mPTP). These mechanisms and processes are associated with NLRP3 inflammasome priming and activation, which can also induce cell death by pyroptosis through the up-regulation of the caspase-1 pathway and IL-18 release. In addition, endothelial dysfunction, both in the presence and absence of MIRI, is also accompanied by altered oxygen levels, decreased nitric oxide production, and ROS overproduction, resulting in the expression of adhesion molecules and leukocyte infiltration in which the NLRP3 inflammasome plays a central role, thus contributing, through endothelial dysfunction, to the alteration of coronary flow, typical of ischemic heart disease. Given the intricate interrelationship between ROS and NLRP3, ROS inhibitors can reduce NLRP3 inflammasome activation, while NLRP3 inhibitors can reduce oxidative stress and inflammation. NLRP3 inhibitors have been intensively studied as anti-inflammatory agents in basic cardiovascular sciences. In this review, we analyze the interrelation between ROS and NLRP3 in ischemic heart disease and the effects of some NLRP3 inhibitors as possible therapeutic agents in this disease condition. All compounds considered in this review need larger studies to confirm their appropriate use in clinical scenarios as anti-ischemic drugs.

Characterization of Electrospun BDMC-Loaded PLA Nanofibers with Drug Delivery Function and Anti-Inflammatory Activity.

Controlled drug release systems are the subject of many investigations to achieve the therapeutic effect of drugs. They have numerous advantages, such as localized effects, lower side effects, and less onset of action. Among drug-delivery systems, electrospinning is a versatile and cost-effective method for biomedical applications. Furthermore, electrospun nanofibers are promising as drug carrier candidates due to their properties that mimic the extracellular matrix. In this work, electrospun fibers were made of Poly-L-lactic acid (PLA), one of the most widely tested materials, which has excellent biocompatible and biodegradable properties. A curcuminoid, bisdemethoxycurcumin (BDMC) was added in order to complete the drug delivery system. The PLA/BDMC membranes were characterized, and biological characteristics were examined in vitro. The results show that the average fiber diameter was reduced with the drug, which was mainly released during the first 24 h by a diffusion mechanism. It was seen that the use of our membranes loaded with BDMC enhanced the rate of proliferation in Schwann cells, the main peripheral neuroglial cells, and modulated inflammation by reducing NLRP3 inflammasome activation. Considering the results, the prepared PLA/BDMC membranes hold great potential for being used in tissue engineering applications.

FAM69C functions as a kinase for eIF2α and promotes stress granule assembly

Stress granules are dynamic cytoplasmic ribonucleoprotein granules that assemble in response to cellular stress. Aberrant formation of stress granules has been linked to neurodegenerative diseases. However, the molecular mechanisms underlying the initiation of stress granules remain elusive. Here we report that the brain-enriched protein kinase FAM69C promotes stress granule assembly through phosphorylation of eukaryotic translation initiation factor 2 (eIF2α). FAM69C physically interacts with eIF2α and functions as a stress-specific kinase for eIF2α, leading to stress-induced protein translation arrest and stress granule assembly. Primary microglia derived from Fam69c knockout mice exhibit aberrant stress granule assembly in response to oxidative stress and ATP. Defective stress granule assembly in microglia correlates with the formation of ASC specks and NLRP3 inflammasome activation, whereas induction of stress granule precludes inflammasome formation. Consistently, increased NLRP3 levels, caspase-1 cleavage and Il18 expression corroborate microglia-associated neuroinflammation in aged Fam69c knockout mice. Our study demonstrates that FAM69C is critical for stress granule assembly and suggests its role in the regulation of microglia function.

MCC950 Regulates Stem Cells Destiny Through Modulating SIRT3-NLRP3 Inflammasome Dynamics During Oxygen Glucose Deprivation.

Ischemic stroke is the major cause of death and morbidity worldwide. Stem cell treatment is at the forefront of ischemic therapeutic interventions. However, the fate of these cells following transplantation is mostly unknown. The current study examines the influence of oxidative and inflammatory pathological events associated with experimental ischemic stroke (oxygen glucose deprivation (OGD)) on the stem cell population (human Dental Pulp Stem Cells, and human Mesenchymal Stem Cells) through the involvement of the NLRP3 inflammasome. We explored the destiny of the above-mentioned stem cells in the stressed micro (-environment) and the ability of MCC950 to reverse the magnitudes. An enhanced expression of NLRP3, ASC, cleaved caspase1, active IL-1β and active IL-18 in OGD-treated DPSC and MSC was observed. The MCC950 significantly reduced NLRP3 inflammasome activation in the aforementioned cells. Further, in OGD groups, oxidative stress markers were shown to be alleviated in the stem cells under stress, which was effectively relieved by MCC950 supplementation. Interestingly, whereas OGD increased NLRP3 expression, it decreased SIRT3 levels, implying that these two processes are intertwined. In brief, we discovered that MCC950 inhibits NLRP3-mediated inflammation by inhibiting the NLRP3 inflammasome and increasing SIRT3. To conclude, according to our findings, inhibiting NLRP3 activation while enhancing SIRT3 levels with MCC950 reduces oxidative and inflammatory stress in stem cells under OGD-induced stress. These findings shed light on the causes of hDPSC and hMSC demise following transplantation and point to strategies to lessen therapeutic cell loss under ischemic-reperfusion stress.

Piceatannol and 3'-Hydroxypterostilbene Alleviate Inflammatory Bowel Disease by Maintaining Intestinal Epithelial Integrity and Regulating Gut Microbiota in Mice.

Inflammatory bowel disease has become a significant health concern across the globe, causing frequent and long-term harm to the digestive system. This study evaluated the effect of piceatannol (PIC) and 3'-hydroxypterostilbene (HPSB) on dextran sulfate sodium (DSS)-induced colitis in mice and investigated whether their effects are exerted through the amelioration of gut barrier dysfunction to reduce the severity of colitis. The findings showed that both PIC and HPSB attenuated inflammation by inhibiting the TNF-α/NF-κB/MLC pathway and reducing NLRP3 inflammasome activation. However, PIC was comparably effective in modulating tight junctions. The results may be attributed to the effect of PIC on reducing cell apoptosis-associated protein expression, including Bax/Bcl-2 and caspase-3 activation. Furthermore, microbiota analysis revealed that both PIC and HPSB increased representative probiotic species, including Akkermansiaceae and Lactobacillus intestinalis, and exhibited inhibitory effects on several bacterial species (Spiroplasmataceae and Acholeplasmataceae). Based on linear discriminant analysis effect size, butyrate-producing bacteria were identified as a biomarker in the PIC group. Overall, the results demonstrated that PIC repressed inflammation, inhibited cell apoptosis, and regulated microbiota composition. Consequently, PIC is more effective in maintaining gut barrier integrity than HPSB, and it is a promising ingredient in the development of functional food for colitis prevention.

Neuroprotective Effect of Taohong Siwu Decoction on Cerebral Ischemia/Reperfusion Injury via Mitophagy-NLRP3 Inflammasome Pathway.

Objective: Globally, cerebral ischemia has been shown to be the second leading cause of death. Our previous studies have shown that Taohong Siwu Decoction (THSWD) exhibits obvious neuroprotective effects on cerebral ischemia/reperfusion (I/R) injury (CIRI). In this study, we further explored the modulatory effect of THSWD on mitochondrial autophagy in CIRI and the relationship between modulatory effect and NLRP3 inflammatory vesicle activation, so as to further explain the mechanism of neuroprotective effect of THSWD. Methods: Middle cerebral artery occlusion reperfusion (MCAO/R) model in rats was built to simulate I/R. Adult male SD rats (220–270 g) were randomly divided into the following four groups: the sham group, the MCAO/R group, the MCAO/R + THSWD group, and the MCAO/R + THSWD + Mitochondrial division inhibitor 1 (Mdivi-1) group. Neurological defect scores were used to evaluate neurological function. 2,3,5-Triphenyltetrazolium chloride (TTC) staining was conducted to measure cerebral infarct volume. Nissl staining, H&E staining and TUNEL staining were executed to detect ischemic cortical neuronal cell viability and apoptosis. Electron microscopy was used to observe the ultrastructural changes of mitochondria. Total Reactive Oxygen Species (ROS) in tissue were measured by fluorescence spectrophotometry, and the activation status of microglia was evaluated by Iba-1/CD16 immunofluorescence staining. The levels of mitophagy-related proteins (LC3, Parkin, PINK1), NLRP3 inflammasome-related proteins (NLRP3, ASC, Pro-caspase-1, Cleaved-caspase-1), and inflammatory cytokines (Pro-IL-18, Pro-IL-1β, IL-18, IL-1β) were evaluated by western blotting. Results: The studies showed that THSWD treatment alleviated cerebral infarction and neurological deficiencies. THSWD upregulated the expressions of autophagy markers (LC3-II/LC3-I and Beclin1) mitochondrial autophagy markers (Parkin and PINK1) after CIRI. Furthermore, THSWD treatment attenuated microglia activation and damage to mitochondrial structures, thereby reducing ROS production and NLRP3 inflammasome activation. In contrast, the mitochondrial autophagy inhibitor Mdivi-1 inhibited the above beneficial effects of THSWD. Conclusions: THSWD exhibits neuroprotective effects against MCAO/R in rats by enhancing mitochondrial autophagy and reducing NLRP3 inflammasome activation.

A Novel Role for Glucocerebrosidase 1 (GBA1) in Parkinson’s Disease

Parkinson’s disease (PD) is the second most common neurodegenerative disorder, yet the pathological mechanisms driving disease onset and progression are not well understood. Glucocerebrosidase 1 (GBA1), the gene encoding the lysosomal enzyme glucoslyceramidase (GCase), has been identified as the most important genetic risk factor contributing to the development of PD. In this study, we aimed to investigate the relationship between inflammasome activation and GCase dysfunction in immune cells in PD. Primary microglial was generated from brains of P0‐ P2 mouse pups. After overnight serum cells was primed with 200 ng/mL of ultrapure LPS for 3 hr followed by treatment with either 40 µM of LTI‐291 or 200 µM of Condruitol β‐epoxide (CBE) for an hour. The cells was activated with 5 mM of adenosine triphosphate (ATP). The supernatant and cells were collected for IL‐1β and GBA assay, respectively. For in vivostudies, 8‐ 12 weeks old male C57BL/6J mice were injected with 20 μg of 6‐hydroxydopamine (6‐OHDA). Mice were treated with 12 mg/kg daily doses of LTI‐291 via oral gavage commencing 24 hours prior to surgery. On day 3 post‐surgery, brains were collected for analysis of protein expression. Our findings demonstrate that NLRP3 inflammasome activation increases IL‐1β production which significantly decreased in the presence of GCase activator, LTI‐291. This indicates the decreased inflammasome activation in conjunction with increased GCase activity. Similarly, GCase activity in LTI‐291 treated cells was significantly increased when compared to control and LPS primed samples. In vivo,6‐ OHDA mice had significant loss of tyrosine hydroxylase (TH), indicative of the substantial dopaminergic neuronal death which was rescued in LTI‐291 treated mice. Overall, these findings identify the potential role of NLRP3 inflammasome activation in driving GCase dysfunction, which could contribute to PD pathogenesis. Furthermore, we demonstrated that pharmacological augmentation of GCase activity was protective, reducing NLRP3 inflammasome activation in vitro and dopaminergic neuronal loss in vivo.

Metformin Protects against Spinal Cord Injury and Cell Pyroptosis via AMPK/NLRP3 Inflammasome Pathway.

Spinal cord injury (SCI) is an extreme neurological impairment with few effective drug treatments. Pyroptosis is a recently found and proven type of programmed cell death that is characterized by a reliance on inflammatory caspases and the release of a large number of proinflammatory chemicals. Pyroptosis differs from other cell death mechanisms such as apoptosis and necrosis in terms of morphological traits, incidence, and regulatory mechanism. Pyroptosis is widely involved in the occurrence and development of SCI. In-depth research on pyroptosis will help researchers better understand its involvement in the onset, progression, and prognosis of SCI, as well as provide new therapeutic prevention and treatment options. Herein, we investigated the role of AMPK-mediated activation of the NLRP3 inflammasome in the neuroprotection of MET-regulated pyroptosis. We found that MET treatment reduced NLRP3 inflammasome activation by activating phosphorylated AMPK and reduced proinflammatory cytokine (IL-1β, IL-6, and TNF-α) release. At the same time, MET improved motor function recovery in rats after SCI by reducing motor neuron loss in the anterior horn of the spinal cord. Taken together, our study confirmed that MET inhibits neuronal pyroptosis after SCI via the AMPK/NLRP3 signaling pathway, which is mostly dependent on the AMPK pathway increase, hence decreasing NLRP3 inflammasome activation.

Urolithin A promotes mitophagy and suppresses NLRP3 inflammasome activation in lipopolysaccharide-induced BV2 microglial cells and MPTP-induced Parkinson's disease model

Microglia-mediated neuroinflammation and mitochondrial dysfunction play critical role in the pathogenic process of Parkinson's disease (PD). Mitophagy plays central role in mitochondrial quality control. Hence, regulation of microglial activation through mitophagy could be a valuable strategy in controlling microglia-mediated neurodegeneration and neuroinflammation. Urolithin A (UA) is a natural compound produced by gut bacteria from ingested ellagitannins (ETs) and ellagic acid (EA). Several preclinical studies have reported the beneficial effects of UA on age-related conditions by increasing mitophagy and blunting excessive inflammatory responses. However, the specific role of UA in pathology of PD remains unknown. In this study, we showed that treatment with UA reduced the loss of dopaminergic neurons, ameliorated behavioral deficits and neuroinflammation in MPTP mouse model of PD. Further study revealed that UA promotes mitophagy, restores mitochondrial function and attenuate proinflammatory response in BV2 microglial cells exposed to LPS. Moreover, UA also reduced NLRP3 inflammasome activation both in vitro and in vivo. Importantly, disruption of microglial mitophagy with pharmacological or genetic approach partly blunted the neuroprotective effects of UA in MPTP mouse model of PD. Collectively, these results provide strong evidence that UA protects against dopaminergic neurodegeneration and neuroinflammation. The mechanism may be related with its inhibition of NLRP3 inflammasome activation via promoting mitophagy in microglia.

Dl-3-n-Butylphthalide Rescues Dopaminergic Neurons in Parkinson's Disease Models by Inhibiting the NLRP3 Inflammasome and Ameliorating Mitochondrial Impairment.

BackgroundNeuroinflammation and mitochondrial impairment play important roles in the neuropathogenesis of Parkinson’s disease (PD). The activation of NLRP3 inflammasome and the accumulation of α-synuclein (α-Syn) are strictly correlated to neuroinflammation. Therefore, the regulation of NLRP3 inflammasome activation and α-Syn aggregation might have therapeutic potential. It has been indicated that Dl-3-n-butylphthalide (NBP) produces neuroprotection against some neurological diseases such as ischemic stroke. We here intended to explore whether NBP suppressed NLRP3 inflammasome activation and reduced α-Syn aggregation, thus protecting dopaminergic neurons against neuroinflammation.MethodsIn our study, we established a MPTP-induced mouse model and 6-OHDA-induced SH-SY5Y cell model to examine the neuroprotective actions of NBP. We then performed behavioral tests to examine motor dysfunction in MPTP-exposed mice after NBP treatment. Western blotting, immunofluorescence staining, flow cytometry and RT-qPCR were conducted to investigate the expression of NLRP3 inflammasomes, neuroinflammatory cytokines, PARP1, p-α-Syn, and markers of microgliosis and astrogliosis.ResultsThe results showed that NBP exerts a neuroprotective effect on experimental PD models. In vivo, NBP ameliorated behavioral impairments and reduced dopaminergic neuron loss in MPTP-induced mice. In vitro, treatment of SH-SY5Y cells with 6-OHDA (100uM,24 h) significantly decreased cell viability, increased intracellular ROS production, and induced apoptosis, while pretreatment with 5uM NBP could alleviated 6-OHDA-induced cytotoxicity, ROS production and cell apoptosis to some extent. Importantly, both in vivo and in vitro, NBP suppressed the activation of the NLRP3 inflammasome and the aggregation of α-Syn, thus inhibited neuroinflammation ameliorated mitochondrial impairments.ConclusionsIn summary, NBP rescued dopaminergic neurons by reducing NLRP3 inflammasome activation and ameliorating mitochondrial impairments and increases in p-α-Syn levels. This current study may provide novel neuroprotective mechanisms of NBP as a potential therapeutic agent.

Attenuating Effect of Chlorella Extract on NLRP3 Inflammasome Activation by Mitochondrial Reactive Oxygen Species.

The NOD-like receptor family, pyrin domain-containing protein 3 (NLRP3) inflammasome has been linked to the pathogenesis of a wide variety of human diseases. Although many drugs and inhibitors have been developed to treat NLRP3-associated diseases, only limited clinical data support their efficacy and safety. Chlorella, a unicellular green alga that is widely and safely used as a food supplement, contains various antioxidants. In this study, we obtained a fat-soluble extract from Chlorella (CE) and demonstrated that it reduced NLRP3 inflammasome activation by inhibiting mitochondrial reactive oxygen species and caspase-1 activation. In addition, CE supplementation attenuated lipopolysaccharide-induced interleukin 1β transcription through activation of hypoxia-inducible factor 1α in vitro and in vivo. As Chlorella is a safe and useful food supplement, it may be a practical pharmacological approach for treating NLRP3-driven diseases.

The role of HMGB1 on TDI-induced NLPR3 inflammasome activation via ROS/NF-κB pathway in HBE cells

To explore the potential role of HMGB1 on TDI-induced NLRP3 inflammasome activation, HBE cells were treated with TDI-HSA conjugate to observe the changes of HMGB1, TLR4, NF-κB, Nrf2 and NLRP3 inflammasome related proteins expressions, ROS release and MMP. NAC, TPCA-1 and Resatorvid pre-treatments were applied to explore the effects of ROS, NF-κB and TLR4 on TDI-induced NLRP3 inflammasome activation. The CRISPR/Cas9 system was used to construct HMGB1 gene knockout HBE cell line and then to explore the role of HMGB1 on TDI-HSA induced NLRP3 inflammasome activation. GL pre-treatment was applied to further confirm the role of HMGB1. Results showed that TDI increased HMGB1, TLR4, P-p65, Nrf2 proteins expressions and ROS release, decreased MMP level and activated NLRP3 inflammasome in HBE cells in a dose dependent manner. NAC, TPCA-1 and Resatorvid pre-treatments decreased the expression of P-p65 and inhibited NLRP3 inflammasome activation. Inhibition of HMGB1 decreased Nrf2 expression and ROS release, improved MMP level and reduced NLRP3 inflammasome activation. GL ameliorated NLRP3 inflammasome activation via inhibiting HMGB1 regulated ROS/NF-κB pathway. These results indicated that HMGB1 was involved in TDI-induced NLRP3 inflammasome activation as a positive regulatory mechanism. The study provided a potential target for early prevention and treatment of TDI-OA.

Cichoric Acid Ameliorates Monosodium Urate-Induced Inflammatory Response by Reducing NLRP3 Inflammasome Activation via Inhibition of NF-kB Signaling Pathway.

Gouty arthritis is characterized by the deposition of monosodium urate (MSU) within synovial joints and tissues due to increased urate concentrations. Here, we elucidated the role of the natural compound cichoric acid (CA) on the MSU crystal-stimulated inflammatory response. The THP-1-derived macrophages (THP-Ms) were pretreated with CA and then stimulated with MSU suspensions. The protein levels of p65 and IκBα, the activation of the NF-κB signaling pathway by measuring the expression of its downstream inflammatory cytokines, and the activity of NLRP3 inflammasome were measured by western blotting and ELISA. CA treatment markedly inhibited the degradation of IκBα and the activation of NF-κB signaling pathway and reduced the levels of its downstream inflammatory genes such as IL-1β, TNF-α, COX-2, and PGE2 in the MSU-stimulated THP-M cells. Therefore, we infer that CA effectively alleviated MSU-induced inflammation by suppressing the degradation of IκBα, thereby reducing the activation of the NF-κB signaling pathway and the NLRP3 inflammasome. These results suggest that CA could be a novel therapeutic strategy in averting acute episodes of gout.