NLRP3 Inflammasome Activation Research Articles

Hypoxia and TNF-alpha modulate extracellular vesicle release from human induced pluripotent stem cell-derived cardiomyocytes.

Extracellular vesicles (EVs) have emerged as important mediators of intercellular communication in the heart under homeostatic and pathological conditions, such as myocardial infarction (MI). However, the basic mechanisms driving cardiomyocyte-derived EV (CM-EV) production following stress are poorly understood. In this study, we generated human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) that express NanoLuc-tetraspanin reporters. These modified hiPSC-CMs allow for quantification of tetraspanin-positive CM-EV secretion from small numbers of cells without the need for time-consuming EV isolation techniques. We subjected these cells to a panel of small molecules to study their effect on CM-EV biogenesis and secretion under basal and stress-associated conditions. We observed that EV biogenesis is context-dependent in hiPSC-CMs. Nutrient starvation decreases CM-EV secretion while hypoxia increases the production of CM-EVs in a nSmase2-dependent manner. Moreover, the inflammatory cytokine TNF-α increased CM-EV secretion through a process involving NLRP3 inflammasome activation and mTOR signalling. Here, we detailed for the first time the regulatory mechanisms of EV biogenesis in hiPSC-CMs upon MI-associated stressors.

Resveratrol improved mitochondrial biogenesis by activating SIRT1/PGC-1α signal pathway in SAP

NLRP3 inflammasomes- pyroptosis axis is activated by microcirculation dysfunction and touched off severe acute pancreatitis (SAP). Activation of PGC-1α can improve microcirculation dysfunction by promoting mitochondrial biogenesis. Resveratrol (RSV), one typical SIRT1 agonist, possesses the ability of alleviating SAP and activing PGC-1α. Therefore, the study was designated to explore whether the protective effect of RSV in SAP was though suppressing NLRP3 inflammasomes- pyroptosis axis via advancing SIRT1/PGC-1α-dependent mitochondrial biogenesis. The models of SAP were induced by treating with sodium taurodeoxycholate in rats and AR42J cells. The pathological injury, water content (dry/wet ratio) and microcirculation function of pancreas, activity of lipase and amylase were used to evaluate pancreatic damage. The expression of inflammatory cytokine was measured by ELISA and RT-PCR. The damage of mitochondrial was evaluated by measuring the changes in Mitochondrial Membrane Potential (ΔΨm), mitochondrial ROS, ATP content and MDA as well as relocation of mtDNA and the activity of SOD and GSH. The expressions of NLRP3 inflammasomes- pyroptosis axis proteins were detected by Western blotting as well as SIRT1/PGC-1α/NRF1/TFAM pathway protein. Moreover, the modification of PGC-1α was measured by co-immunoprecipitation. The results displayed that RSV can significantly improve the damage of pancreas and mitochondrial, decrease the expression of pro-inflammatory factor and the activation of NLRP3 inflammasomes- pyroptosis axis, promote the expression of an-inflammatory factor and the deacetylation of PGC-1α together with facilitating SIRT1/PGC-1α-mediating mitochondrial biogenesis. Therefore, the protective effect of RSV in SAP is though inactivation of NLRP3 inflammasomes- pyroptosis axis via promoting mitochondrial biogenesis in a SIRT1/PGC-1α-dependent manner.

A Novel Dipterocarpol Derivative That Targets Alpha-Glucosidase and NLRP3 Inflammasome Activity for Treatment of Diabetes Mellitus.

Type 2 diabetes mellitus is a chronic metabolic disorder characterized by persistent hyperglycemia, chronic inflammation, impaired insulin secretion, and/or peripheral insulin resistance. Current α-glucosidase inhibitors approved for clinical use exhibit limited efficacy compared to other glucose-lowering agents. In this study, a series of mono- and bis-benzylidene derivatives were synthesized via aldol condensation of 3-oxo-dammarane triterpenoids with terephthalic aldehyde. The target mono- and bis-benzylidene derivatives, based on the dammarane triterpenoids hollongdione 1, (20S)-23,24-epoxy-25,26,27-trinordammar-3,24-dione 2, and 24(R,S)-20(S)-epoxy-25-hydroxy-dammar-3-one 3, were successfully synthesized. Several of these inhibitors demonstrated significantly greater efficacy than the reference drug acarbose. Notably, compound 4 inhibited S. cerevisiae α-glucosidase with an IC50 of 2.67 μM. Furthermore, the target compounds effectively inhibited NLRP3 inflammasome activation, reducing IL-1β production in LPS+ATP-stimulated murine peritoneal macrophages without detectable cytotoxicity. Compound 8, which exhibited dual activity, was further characterized as an inhibitor of NLRP3 activation in peripheral blood mononuclear cells, leading to the prevention of pyroptosis and IL-1β release. Additionally, compound 8 was shown to promote neuronal survival in LPS+ATP-treated rat hippocampal slices, highlighting its potential as a promising antidiabetic agent that targets both postprandial hyperglycemia and metaflammation.

(Pro)renin receptor aggravates myocardial pyroptosis in diabetic cardiomyopathy through AMPK-NLRP3 pathway.

As one of the most common complications of diabetes, diabetic cardiomyopathy (DCM) is the main cause of heart failure in patients with diabetes. However, the lack of effective treatments for DCM remains a clinical challenge. (Pro) renin receptor (PRR) is a member of renin angiotensin aldosterone system (RAAS). Here, we aim to determine whether PRR is involved in myocardial pyroptosis in diabetic cardiomyopathy. We established diabetic rats model by intraperitoneal injection of streptozotocin (STZ). PRR overexpression adenovirus or PRR knockdown adenovirus was injected into the tail vein. Western blot, histopathology and immunohistochemistry staining, ELISA and Echocardiography were used to detect cardiac function changes and myocardial injury levels of DCM rats. Primary cardiomyocytes were stimulated with high glucose and PRR overexpression or PRR knockdown was achieved by adenovirus transfection, we also used the inhibitor of AMPK to decrease the activity of AMPK. Western blot, Real-time PCR, Immunofluorescence and ELISA were used to detect the level of PRR and pyroptosis in cardiomyocyte. We found that high glucose increased the expression of PRR in heart. After overexpression of PRR, the expression of the pyroptosis related proteins such as Caspase-1, IL-1β, IL-18, and NLRP3 was significantly increased, the phosphorylation level of AMPK was significantly decreased, and the fibrosis level was significantly increased, thus aggravating the cardiac function injury of DCM. On the contrary, PRR knockdown can alleviate the level of myocardial pyroptosis in DCM and improve cardiac function. The related mechanism was that PRR could inhibit AMPK phosphorylation and promote the activation of NLRP3 inflammasome. PRR aggravated pyroptosis of cardiomyocyte, increased the dysfunction of cardiomyocyte, and may be related to the decrease of AMPK phosphorylation and the overactivation of NLRP3. This may provide new ideas and targets for the treatment of DCM.

Outer Membrane Vesicles Derived From Fusobacterium nucleatum Trigger Periodontitis Through Host Overimmunity.

The virulent bacteria-induced host immune response dominates the occurrence and progression of periodontal diseases because of the roles of individual virulence factors from these pathogens in the initiation and spread of inflammation. Outer membrane vesicles (OMVs) as a pathogenic entity have recently attracted great attention as messenger bridges between bacteria and host tissues. Herein, the novel role of OMVs derived from Fusobacterium nucleatum in the occurrence of periodontitis is dissected. In a rat periodontitis model, it is found that OMVs derived from F. nucleatum caused deterioration of periodontitis by enhancing inflammation of the periodontium and absorption of alveolar bone, which is almost equivalent to the effect of F. nucleatum itself. Furthermore, that OMVs can independently induce periodontitis is shown. The pathogenicity of OMVs is attributed to multiple pathogenic components identified by omics. After entering human periodontal ligament stem cells (hPDLSCs) by endocytosis, OMVs activated NLRP3 inflammasomes and impaired the mineralization of hPDLSCs through NF-κB (p65) signaling, leading to the final injury of the periodontium and damage of alveolar bone in periodontitis. These results provide a new understanding of OMVs derived from pathogens and cues for the prevention of periodontitis.

Dihydro-resveratrol ameliorates NLRP3 inflammasome-mediated neuroinflammation via Bnip3-dependent mitophagy in Alzheimer's disease.

Dihydro-resveratrol (DHR), a polyphenol derivative, that has been demonstrated to suppress inflammation-mediated injury. However, it is still unknown whether it has anti-neuroinflammatory and neuroprotective effects, and a therapeutic action in Alzheimer's disease (AD). The anti-inflammatory and anti-Alzheimer's disease actions of dihydro-resveratrol were investigated using lipopolysaccharide (LPS) and AD mice models, and primary microglial cells. The changes in behaviour in mice were detected by the Morris water maze test and open-field test. Flow cytometry assay, western blotting, immunofluorescence assays and co-immunoprecipitation were used to investigate the changes in the NLRP3 inflammasome activation and mitophagy. In this study, in vivo observations indicated that the administration of dihydro-resveratrol (DHR) dramatically restored spatial learning, memory ability, autophagy and mitophagy, attenuated NLRP3 inflammasome activation, neuroinflammation and amyloid precursor protein pathology in LPS mice and AD mice. In addition, the inhibition of autophagy and mitophagy, or the activation of NLRP3 in vivo greatly abolished DHR-generated therapeutic efficacy on neuroinflammation, amyloid precursor protein pathology and cognitive loss. Further examination indicated that the application of DHR after the LPS and ATP exposure significantly inhibited the NLRP3 inflammasome activation, neuroinflammation and enhanced autophagic and mitophagic activation in microglia. Additionally, in vitro results show that DHR protects microglial cells against LPS and ATP-induced cytotoxicity by inhibiting NLRP3 inflammasome through activating Bnip3-dependent mitophagy and ULK phosphorylation. In summary, these findings suggest that dihydro-resveratrol (DHR) possesses potent anti-neuroinflammatory property and can act as a potential therapeutic agent for the treatment of AD.

Bidirectional roles of extracellular and intracellular HMGB2 in the development of atherosclerosis

Abstract Rationale: HMGB family plays an important role in homeostasis of immunity and regulation of inflammation. However, the role of HMGB2 in atherosclerosis remains uncharacterized. Objective The current study aimed to ascertain the relation of HMGB2 levels in macrophages of atherosclerotic plaques and in circulatory monocytes/macrophages with coronary artery disease (CAD), and to investigate the influence of HMGB2 on atherogenesis in mice and inflammatory reaction in vitro. Methods/Results We evaluated HMGB2 levels in macrophages of atherosclerotic plaques in coronary endarterectomy tissue from patients with angiographically documented severe coronary artery disease (CAD)(n=23), and in coronary artery specimens from autopsy cases with no or mild atherosclerosis (n=11) by immunohistochemistry. HMGB2 levels were significantly decreased in macrophages of progressive atherosclerotic plaques, as compared with those from mild plaques. Moreover, we also analyzed HMGB2 levels in peripheral blood monocytes from patients with CAD (n=44) or age- and sex-matched non-CAD controls (n=44). The results showed that decreased intracellular HMGB2 protein levels in monocytes were associated with CAD. In experiments, HMGB2 deficiency significantly promoted atherosclerosis in ApoE-/- mice, whereas such increment of atherosclerosis was markedly attenuated after transplantation of bone marrow derived macrophages from ApoE-/-mice. Mechanistically, intracellular HMGB2 repressed IL-1 transcription and induced autophagic degradation of NLRP3, thus suppressing NLRP3 inflammasome priming and activation. Extracellular HMGB2 levels increased in progressive atherosclerotic plaques with macrophage apoptosis. Recombinant HMGB2 protein promoted inflammation in vitro via RAGE and intraperitoneal administration of this protein exacerbated atherosclerosis in mice. Conclusion This study indicates that decreased HMGB2 level in macrophage is associated with inflammation and atherosclerosis in patients with CAD. Intracellular HMGB2 inhibits NLRP3 inflammasome and atherosclerosis in mice, whereas released HMGB2 promotes inflammation and atherosclerosis in mice.

Alda-1 Ameliorates Oxidative Stress-Induced Cardiomyocyte Damage by Inhibiting the Mitochondrial ROS/TXNIP/NLRP3 Pathway.

Alda-1 functions as an agonist of aldehyde dehydrogenase (ALDH2) within the mitochondria, contributing to the preservation of mitochondrial structure and function. This study aimed to determine whether Alda-1 inhibited the accumulation of mitochondrial reactive oxygen species (mtROS) and improved cardiomyocyte damage under oxidative stress. Cardiomyocytes derived from human induced pluripotent embryonic stem cells (iPSC-CMs) and human AC16 cardiomyocytes were chosen for the experiments. Oxidative stress was induced in both cardiomyocyte types using hydrogen peroxide (H2O2), a commonly employed agent. The mtROS accumulation and mitochondrial functional status were assessed by measuring the ROS content, mitochondrial membrane potential, and mitochondrial respiratory chain function. Co-IP experiments were used to analyze whether mtROS promoted protein interactions between TXNIP and NLRP3 and induced NLRP3 inflammasome activation. The results showed that Alda-1 mitigated damage to mitochondrial structure and function under oxidative stress, concurrently reducing the accumulation of mtROS. Co-IP experiments revealed that elevated mtROS levels attenuated the protein interaction between TXNIP and TRX while intensifying the interaction between TXNIP and NLRP3. Consequently, this triggers activation of the NLRP3 inflammasome, leading to cardiomyocyte damage. In contrast, TXNIP knockdown inhibited H2O2-induced myocardial injury and enhanced the therapeutic effects of Alda-1. Collectively, the results show that, in an H2O2 environment, Alda-1 increased the production of ALDH2 activity in cardiomyocytes, hindered the production of mtROS, disrupted the interaction between TXNIP and NLRP3, and alleviated myocardial damage during oxidative stress.

The role of Gαq in regulating NLRP3 inflammasome activation.

G proteins are a class of important signal transducers in mammalians. G proteins can corpoarated with G proteincoupled receptors (GPCRs) and transmit signals from extracellular stimuli into intracellular response, which will regulate a series of biological functions. G-proteins are heterotrimeric proteins composed of Gα, Gβ, and Gγ subunits. Based on structural and functional similarity of their α-subunits, G proteins are typically grouped into four classes (Gi, Gs, Gq/11, and G12/13). The Gq/11 subfamily consists of Gq, G11, G14, and G15/16 proteins. Gαq is the α-subunit of Gq protein and encoded by GNAQ. Our previous studies revealed that Gαq play an important role in regulating T cell survival and T cell differentiation. Inflammasomes are multiprotein complexes that play a critical role in modulating innate inflammatory response. NLRP3 inflammasome is currently the most extensively studied inflammasome. We found that Gαq suppressed NLRP3 inflammasome activation in macrophage, Gαq also suppressed NLRP3 inflammasome activation in a LPS-induced sepsis mouse model. Gαq can locate to mitochondria and Gαq was required for the maintenance of mitochondrial homeostasis. Gαq regulated NLRP3 inflammasome activation by modulating mitochondrial reactive oxygen species (mtROS). We found that Gαq suppressed NLRP3 inflammasome activation in macrophage, Gαq also suppressed NLRP3 inflammasome activation in a LPS-induced sepsis mouse model. Gαq can locate to mitochondria and Gαq was required for the maintenance of mitochondrial homeostasis. Gαq regulated NLRP3 inflammasome activation by modulating mitochondrial reactive oxygen species (mtROS). Our results indicate that Gαq regulates NLRP3 inflammasome activation by modulating mitochondrial ROS production. Our research provides new mechanistic insight into the activation of NLRP3 inflammasome. As it has been proved that NLRP3 inflammasome plays an important role in the pathogenesis many diseases such as Alzheimer's disease, cancer, and inflammatory bowel disease, Gαq might become a novel drug target for these diseases in future.

Aminoadipic acid aggravates atherosclerotic vascular inflammation through ROS/TXNIP/NLRP3 pathway, a harmful microbial metabolite reduced by paeonol

AimOur previous study has found a differential microbial metabolite in atherosclerosis (AS) mice, aminoadipic acid (AAA), which was considered as a potential harmful metabolite. However, whether it can promote AS vascular inflammation and its mechanisms remain unclear. Paeonol (Pae) plays an anti-AS role by regulating the metabolic profile, but whether Pae exerts its antiatherogenic effect by reducing serum AAA levels is unknown. ResultsThe clinical trial results showed that the AS patients’ serum AAA levels were higher than those healthy people’. Besides, AAA supplementation could increase aortic plaque size, serum inflammatory cytokines levels and liver malondialdehyde, superoxide dismutase levels in AS mice. Moreover, after AAA stimulation, the ROS levels and ASC, TXNIP, NLRP3 and caspase-1 proteins levels were increased in HUVECs, which could be reversed by antioxidant NAC and NLRP3 inhibitor. Pae significantly reduced the plaque size in the aorta, improved blood lipid levels and decreased serum inflammation factor levels in AS mice. Simultaneously, Pae could reduce the serum AAA levels of AS mice through the gut microbiota transmission. Finally, Pae inhibited NLRP3 inflammasome activation in aortas of AS mice. Broad-spectrum antibiotics could weaken the inhibitory effect of Pae on NLRP3 inflammasome. ConclusionOur study clarified that AAA could promote AS vascular inflammation via activating the ROS/TXNIP/NLRP3 pathway. Pae could inhibit AS development by reducing serum AAA levels in a microbiota-dependent manner. Taken together, we proposed that AAA could be served as a potential biomarker for AS clinical diagnosis and provided a new treatment strategy for AS.

Comparative Modulation of Immune Responses and Inflammation by n-6 and n-3 Polyunsaturated Fatty Acids in Oxylipin-Mediated Pathways

Our study investigates the comparative effects of n-6 and n-3 polyunsaturated fatty acids (PUFAs) on immune modulation and inflammation using a fat-1 transgenic mouse model capable of endogenously converting n-6 PUFAs to n-3 PUFAs. The results show that n-6 PUFAs, particularly arachidonic acid (AA), promote a pro-inflammatory environment by increasing the production of inflammatory mediators, including leukotrienes and prostaglandins, while upregulating NFκB signaling and NLRP3 inflammasome activation. In contrast, n-3 PUFAs, such as eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), exhibit anti-inflammatory and pro-resolving properties by enhancing the production of resolvins, protectins, and maresins, and upregulating PPARα expression. Quantitatively, n-3 PUFAs led to a 4-fold increase in resolvin levels compared to the n-6 group (p < 0.001), promoting a resolution of inflammation. This study underscores the critical importance of maintaining an optimal balance between n-6 and n-3 PUFAs in the diet to prevent chronic inflammation and suggests that increasing dietary n-3 PUFAs may mitigate inflammation-driven diseases. The findings highlight the need for further research into the optimal dietary ratios of n-6 and n-3 PUFAs for immune health and disease prevention.

The Protective Role of Baicalin in the Regulation of NLRP3 Inflammasome in Different Diseases.

The NACHT, LRR, and PYD domains-containing protein 3 (NLRP3) inflammasome consists of pro-caspase-1, NLRP3 and apoptosis-related speckle-like protein (ASC). It can detect multiple microorganisms, endogenous danger signals and environmental stimulus including adenosine triphosphate (ATP), urate, cholesterol crystals, and so on, thereby forming activated NLRP3 inflammasome. During the course of the activation of NLRP3 inflammasome, pro-caspase-1 is transformed into activated caspase-1 that results in the maturation and secretion of interleukin-1beta (IL-1β) and IL-18. The dysfunction of NLRP3 inflammasome participates in multiple diseases such as liver diseases, renal diseases, nervous system diseases and diabetes. Baicalin is the primary bioactive component of Scutellaria baicalensis, which has been used since ancient times. Baicalin has many types of biological functions, such as anti-bacterial, anti-tumor and antioxidant. More and more evidence suggests that baicalin regulation of NLRP3 inflammasome is involved in different diseases. However, the mechanism is still elusive. Here, we reviewed the progress of baicalin regulation of NLRP3 inflammasome in many kinds of diseases to lay a foundation for future researches.

Benzoylmesaconine mitigates NLRP3 inflammasome-related diseases by reducing intracellular K+ efflux and disrupting NLRP3 inflammasome assembly

BackgroundBenzoylmesaconine (BMA), a major alkaloid derived from the traditional Chinese medicine Aconitum carmichaeli Debx, exhibits potent anti-inflammatory properties. However, the precise mechanism underlying its action remains unclear. PurposeThis study aimed to investigate the inhibitory mechanism of BMA on the NLRP3 inflammasome and assess its therapeutic efficacy in NLRP3-related metabolic diseases. MethodsA classic NLRP3 inflammasome-activated bone marrow-derived macrophage (BMDM) model was established to evaluate BMA's effects on NLRP3 upstream and downstream protein expression, as well as pyroptosis. Two distinct animal disease models, MSU-induced gouty arthritis and DSS-induced colitis, were utilized to validate BMA's anti-inflammatory activity in vivo. ResultsIn vitro findings revealed that BMA can suppress NLRP3 inflammasome activation by inhibiting interleukin-1β (IL-1β) secretion and GSDMD-N protein expression. This mechanism involved blocking intracellular K+ efflux and interfering with the formation of NLRP3 inflammasomes. In vivo studies demonstrated that BMA significantly alleviated inflammatory symptoms in MSU-induced acute gout and DSS-induced colitis models. ConclusionThese findings suggest that BMA effectively inhibits the activation of the NLRP3 signaling pathway through dual mechanisms: reducing intracellular K+ efflux and disrupting NLRP3 inflammasome assembly. This multifaceted action highlights the therapeutic potential of BMA for NLRP3-related diseases.

Preventive effect of imperatorin against doxorubicin-induced cardiotoxicity through suppression of NLRP3 inflammasome activation.

Cardiotoxicity is one of the major obstacles to anthracycline chemotherapy. Anthracycline cardiotoxicity is closely associated with inflammation. Imperatorin (IMP), a furocoumarin ingredient extracted from Angelica dahurica, might have potential activity in preventing anthracycline cardiotoxicity due to its anti-cancer, anti-inflammatory, anti-oxidant, cardioprotective properties. This study aims to reveal the effect of IMP on doxorubicin (DOX)-induced cardiotoxicity and its underlying mechanism. We established a rat model of DOX-induced cardiotoxicity by intraperitoneal injection with DOX (1.25mg/kg twice weekly for 6weeks), and found that both IMP (25mg/kg and 12.5mg/kg) and dexrazoxane 12.5mg/kg relieved DOX-induced reductions in heart weight, change in cardiac histopathology, and elevated serum levels of LDH, AST and CK-MB. Moreover, DOX upregulated mRNA levels of NLRP3, CASP1, GSDMD, ASC, IL-1β and IL-18, elevated protein expressions of NLRP3, ASC, GSDMD-FL, GSDMD-N, pro‑caspase‑1, caspase‑1 p20, pro‑IL‑1β and IL‑1β in heart tissues, as well as increased serum levels of pro-inflammatory cytokines including IL-1β and IL-18, however both of IMP and dexrazoxane suppressed these alterations. In addition, we carried out neonatal rat cardiomyocytes experiments to confirm the results of the in vivo study. Consistently, pretreatment with IMP 25µg/mL relieved DOX (1μg/mL)-induced cardiomyocytes injury, including decreased cell viability and reduced supernatant LDH. IMP inhibited DOX-induced activation of NLRP3 inflammasome in cardiomyocytes. In conclusion, IMP had a protective effect against DOX-induced cardiotoxicity via repressing the activation of NLRP3 inflammasome. These findings suggest that IMP may be a promising alternative or adjunctive drug for the prevention of anthracycline cardiotoxicity.

Comprehensive evaluation of the nephrotoxicity of carbon quantum dots: Effects of the surface charge

Carbon quantum dots (CQDs) are garnering attention for their broad applications. This study offers a detailed evaluation of the biomedical safety and health risks of carbon quantum dots (CQDs) with different surface modifications, addressing a key gap in their safe application. It focuses on three CQD types: diammonium citrate-based (CQDs-A), spermidine trihydrochloride-based (CQDs-S), and a combination (CQDs-A/S), analyzing their physicochemical properties, cytotoxicity, oxidative stress, inflammatory responses, and nephrotoxicity. While all CQDs were under 10 nm, their biological impacts varied. Positively charged CQDs-S and CQDs-A/S showed significant cytotoxicity in HEK293 cells, inducing oxidative stress but not activating NLRP3 inflammasome, indicating a limited inflammatory response. Renal integrity remained unaffected, with stable zonula occludens 2 expression and unaltered renal markers. In vivo studies in BALB/c mice further supported the safety of CQDs, showing no organ damage or kidney pathology at high doses. The findings underscore the potential for safe biomedical use of CQDs, particularly when their retention time is minimized. This research makes a novel contribution by linking CQDs' surface charge to cytotoxic effects and oxidative stress, providing key insights into their safe use in biomedicine and filling a critical gap in nanoparticle toxicity studies.

Glibenclamide Prevents Inflammation by Targeting NLRP3 Inflammasome Activation In Vitro

The NLRP3 inflammasome is known to play a significant role in the development of neurodegeneration and physiological aging, as well as the development of metabolic inflammation, which has generated significant interest in the scientific community in finding effective inhibitors of the NLRP3 inflammasome and assessing their effects. The purpose of this study was to evaluate the effect of pharmacological modulation of NLRP3 activity using an indirect NLRP3 inflammasome inhibitor, glibenclamide, on the expression of metaflammasome components in in vitro brain cells obtained from middle-aged mice. The study revealed that glibenclamide reduces the expression of pro-inflammatory markers NLRP3 and IL18 in cell culture, which in turn leads to the prevention of phosphorylation of protein kinases of the metaflammasome complex – PKR and IKKβ. However, we did not observe changes in the expression of pathologically phosphorylated IRS, as well as in the number of senescent cells in cultures after the exposure to glibenclamide.

Inhibition of NLRP3 inflammasome contributes to paclitaxel efficacy in triple negative breast cancer treatment.

Chronic inflammation and NLRP3 inflammasome activation are among the determining factors of breast malignancies. Paclitaxel (PTX) is a drug used in breast cancer treatment which sustains prolonged inflammation, reducing the effectiveness of chemotherapy. Considering the impact of inflammatory processes in cancer progression, there is a strong concern to develop therapeutic strategy targeting NLRP3 inflammasome for triple-negative breast cancer (TNBC) treatment. Therefore, the aim of this study was to evaluate the potential of PTX and NLRP3 inflammasome modulation to counterbalance TNBC by inducing programmed cell death and inhibiting the activity of pro-inflammatory cytokines. The obtained results suggested the strong interaction between NLRP3 inflammasome and TNBC and revealed that pharmacological inhibition, using NLRP3-specific inhibitor MCC950, and genetic silencing of NLRP3 inflammasome using specific small interfering RNA, reduced inflammatory responses and facilitated PTX-determined tumor cell death. Thus, NLRP3 inflammasome manipulation in combination with anti-tumor drugs opens up new therapeutic perspectives for TNBC therapy.

Targeting S100A9-TLR2 axis controls macrophage NLRP3 inflammasome activation in fatty liver ischemia reperfusion injury

Abstract Liver ischemia reperfusion (IR) injury significantly impacts clinical outcomes by increasing the risk of hepatic dysfunction after liver surgery. Fatty livers are more susceptible to IR stress. Recent studies have demonstrated that S100A9 plays a crucial role in both IR injury and the progression of liver steatosis. Nevertheless, the precise mechanisms underlying these effects remain unclear. In our study, transcriptome analysis of fatty livers subjected to IR insult in mice identified S100A9 as an important mediator. Employing loss-of-function approaches, we investigated the immune regulatory function of S100A9 and its downstream signaling in fatty liver IR injury. As expected, S100A9 emerged as one of the most significantly upregulated genes during the reperfusion stage in fatty livers. Genetic knockdown of S100A9 markedly ameliorated liver pathological damage, evidenced by reduced macrophage/neutrophil infiltration as well as the decreased expression of proinflammatory factors. Transcriptome/functional studies revealed that S100A9 triggered liver inflammatory response via regulating Toll-like receptor 2 (TLR2)/Activating transcription factor 4 (ATF4) signaling. Additionally, TLR2 expression was notably increased in macrophages from ischemic fatty livers. In vitro, recombinant S100A9-stimulated macrophages exhibited the elevated production of proinflammatory factors and TLR2/ATF4 pathway activation. Intriguingly, S100A9 facilitated ATF4 nuclear translocation and enhanced NEK7/NLRP3 inflammasome activation in macrophages. In conclusion, our study identified S100A9 as a key regulator responsible for macrophage NLRP3 inflammasome activation and subsequent inflammatory injury in fatty liver IR process. Targeting TLR2/ATF4 signaling may offer a novel therapeutic strategy for mitigating S100A9-mediated liver injury.

GLP-1R activation attenuates the progression of pulmonary fibrosis via disrupting NLRP3 inflammasome/PFKFB3-driven glycolysis interaction and histone lactylation

BackgroundPulmonary fibrosis is a serious interstitial lung disease with no viable treatment except for lung transplantation. Glucagon-like peptide-1 receptor (GLP-1R), commonly regarded as an antidiabetic target, exerts antifibrotic effects on various types of organ fibrosis. However, whether GLP-1R modulates the development and progression of pulmonary fibrosis remains unclear. In this study, we investigated the antifibrotic effect of GLP-1R using in vitro and in vivo models of pulmonary fibrosis.MethodsA silica-induced pulmonary fibrosis mouse model was established to evaluate the protective effects of activating GLP-1R with liraglutide in vivo. Primary cultured lung fibroblasts treated with TGF-β1 combined with IL-1β (TGF-β1 + IL-1β) were used to explore the specific effects of liraglutide, MCC950, and 3PO on fibroblast activation in vitro. Cell metabolism assay was performed to determine the glycolytic rate and mitochondrial respiration. RNA sequencing was utilized to analyse the underlying molecular mechanisms by which liraglutide affects fibroblast activation. ChIP‒qPCR was used to evaluate histone lactylation at the promoters of profibrotic genes in TGF-β1 + IL-1β- or exogenous lactate-stimulated lung fibroblasts.ResultsActivating GLP-1R with liraglutide attenuated pulmonary inflammation and fibrosis in mice exposed to silica. Pharmacological inhibition of the NLRP3 inflammasome suppressed PFKFB3-driven glycolysis and vice versa, resulting in decreased lactate production in TGF-β1 + IL-1β-stimulated lung fibroblasts. Activating GLP-1R inhibited TGF-β1 + IL-1β-induced fibroblast activation by disrupting the interaction between the NLRP3 inflammasome and PFKFB3-driven glycolysis and subsequently prevented lactate-mediated histone lactylation to reduce pro-fibrotic gene expression. In addition, activating GLP-1R protected mitochondria against the TGF-β1 + IL-1β-induced increase in oxidative phosphorylation in fibroblasts. In exogenous lactate-treated lung fibroblasts, activating GLP-1R not only repressed NLRP3 inflammasome activation but also alleviated p300-mediated histone lactylation. Finally, GLP-1R activation blocked silica-treated macrophage-conditioned media-induced lung fibroblast activation.ConclusionsThe antifibrotic effects of GLP-1R activation on pulmonary fibrosis could be attributed to the inhibition of the interaction between NLRP3 inflammasome and PFKFB3-driven glycolysis, and histone lactylation in lung fibroblasts. Thus, GLP-1R is a specific therapeutic target for the treatment of pulmonary fibrosis.

Acylated Anthocyanins From Black Carrots and Their Related Phenolic Acids Diminish Priming and Activation of the NLRP3 Inflammasome in THP-1 Monocytes.

Excessive activation of the nucleotide-binding oligomerization domain-like receptor pyrin domain-containing protein 3 (NLRP3) inflammasome contributes to chronic inflammation. Thus, targeting NLRP3 inflammasome activation by anthocyanins may prevent inflammatory diseases. Therefore, the present study determines the influence of a black carrot extract (BCE) with high amounts of acylated anthocyanins and their related phenolic acids on the NLRP3 inflammasome. THP-1 monocytes are pretreated with a BCE, cyanidin-3-glucoside (C3G), or hydroxycinnamic acids. NLRP3 inflammasome assembly is initiated by priming THP-1 monocytes with lipopolysaccharide and/or activating the NLRP3 inflammasome with nigericin. Flow cytometry is used to assess apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC) speck formation, as well as ASC and NLRP3 protein expression. Caspase-1 activity is measured using a bioluminescent assay, and cytokine concentrations are determined by enzyme-linked immunosorbent assays (ELISA). C3G and phenolic acids diminish ASC and NLRP3 protein expression. In addition, C3G and phenolic acids attenuate ASC speck formation. Furthermore, the BCE and C3G decline caspase-1 activity. Consistently, IL-1β and IL-18 secretion are reduced upon NLRP3 inflammasome activation. The present study shows that a BCE with high amounts of acylated anthocyanins and their related phenolic acids diminish priming and activation of the NLRP3 inflammasome in THP-1 monocytes.