Chronic PM2.5 exposure disrupts intestinal barrier integrity via microbial dysbiosis-triggered TLR2/5-MyD88-NLRP3 inflammasome activation.

BACKGROUND: PM2.5, a known public health risk, is increasingly linked to intestinal disorders, however, the mechanisms of its impact are not fully understood. PURPOSE: This study aimed to explore the impact of chronic PM2.5 exposure on intestinal barrier integrity and to uncover the underlying molecular mechanisms. METHODS: C57BL/6 J mice were exposed to either concentrated ambient PM2.5 (CPM) or filtered air (FA) for six months to simulate urban pollution conditions. We evaluated intestinal barrier damage, microbial shifts, and metabolic changes through histopathology, metagenomics, and metabolomics. Analysis of the TLR signaling pathway was also conducted. RESULTS: The mean concentration of PM2.5 in the CPM exposure chamber was consistently measured at 70.9 ± 26.8 µg/m³ throughout the study period. Our findings show that chronic CPM exposure significantly compromises intestinal barrier integrity, as indicated by reduced expression of the key tight junction proteins Occludin and Tjp1/Zo-1. Metagenomic sequencing revealed significant shifts in the microbial landscape, identifying 35 differentially abundant species. Notably, there was an increase in pro-inflammatory nongastric Helicobacter species and a decrease in beneficial bacteria, such as Lactobacillus intestinalis, Lactobacillus sp. ASF360, and Eubacterium rectale. Metabolomic analysis further identified 26 significantly altered metabolites commonly associated with intestinal diseases. A strong correlation between altered bacterial species and metabolites was also observed. For example, 4 Helicobacter species all showed positive correlations with 13 metabolites, including Lactate, Bile acids, Pyruvate and Glutamate. Additionally, increased expression levels of TLR2, TLR5, Myd88, and NLRP3 proteins were noted, and their expression patterns showed a strong correlation, suggesting a possible involvement of the TLR2/5-MyD88-NLRP3 signaling pathway. CONCLUSIONS: Chronic CPM exposure induces intestinal barrier dysfunction, microbial dysbiosis, metabolic imbalance, and activation of the TLR2/5-MyD88-NLRP3 inflammasome. These findings highlight the urgent need for intervention strategies to mitigate the detrimental effects of air pollution on intestinal health and identify potential therapeutic targets.

Aloe-emodin alleviates cerebral ischemia-reperfusion injury by regulating microglial polarization and pyroptosis through inhibition of NLRP3 inflammasome activation.

BACKGROUND: Microglial activation plays a crucial role in injury and repair after cerebral ischemia, and microglial pyroptosis exacerbates ischemic injury. NOD-like receptor protein 3 (NLRP3) inflammasome activation has an important role in microglial polarization and pyroptosis. Aloe-emodin (AE) is a natural anthraquinone compound originated from rhubarb and aloe. It exerts antioxidative and anti-apoptotic effects during cerebral ischemia/reperfusion (I/R) injury. However, whether AE affects microglial polarization, pyroptosis, and NLRP3 inflammasome activation remains unknown. PURPOSE: This study aimed to explore the effects of AE on microglial polarization, pyroptosis, and NLRP3 inflammasome activation in the cerebral infarction area after I/R. METHODS: The transient middle cerebral artery occlusion (tMCAO) and oxygen-glucose deprivation/re-oxygenation (OGD/R) methods were used to create cerebral I/R models in vivo and in vitro, respectively. Neurological scores and triphenyl tetrazolium chloride and Nissl staining were used to assess the neuroprotective effects of AE. Immunofluorescence staining, quantitative polymerase chain reaction and western blot were applied to detect NLRP3 inflammasome activation and microglial polarization and pyroptosis levels after tMCAO or OGD/R. Cell viability and levels of interleukin (IL)-18 and IL-1ß were measured. Finally, MCC950 (an NLRP3-specific inhibitor) was used to evaluate whether AE affected microglial polarization and pyroptosis by regulating the activation of the NLRP3 inflammasome. RESULTS: AE improved neurological function scores and reduced the infarct area, brain edema rate, and Nissl-positive cell rate following I/R injury. It also showed a protective effect on BV-2 cells after OGD/R. AE inhibited microglial pyroptosis and induced M1 to M2 phenotype transformation and suppressed microglial NLRP3 inflammasome activation after tMCAO or OGD/R. The combined administration of AE and MCC950 had a synergistic effect on the inhibition of tMCAO- or OGD/R-induced NLRP3 inflammasome activation, which subsequently suppressed microglial pyroptosis and induced microglial phenotype transformation. CONCLUSION: AE exerts neuroprotective effects by regulating microglial polarization and pyroptosis through the inhibition of NLRP3 inflammasome activation after tMCAO or OGD/R. These findings provide new evidence of the molecular mechanisms underlying the neuroprotective effects of AE and may support the exploration of novel therapeutic strategies for cerebral ischemia.

SLC25A3 negatively regulates NLRP3 inflammasome activation by restricting the function of NLRP3.

The NACHT, leucine-rich repeat, and pyrin domains-containing protein 3 (collectively known as NLRP3) inflammasome activation plays a critical role in innate immune and pathogenic microorganism infections. However, excessive activation of NLRP3 inflammasome will lead to cellular inflammation and tissue damage, and naturally it must be precisely controlled in the host. Here, we discovered that solute carrier family 25 member 3 (SLC25A3), a mitochondrial phosphate carrier protein, plays an important role in negatively regulating NLRP3 inflammasome activation. We found that SLC25A3 could interact with NLRP3, overexpression of SLC25A3 and knockdown of SLC25A3 could regulate NLRP3 inflammasome activation, and the interaction of NLRP3 and SLC25A3 is significantly boosted in the mitochondria when the NLRP3 inflammasome is activated. Our detailed investigation demonstrated that the interaction between NLRP3 and SLC25A3 disrupted the interaction of NLRP3-NEK7, promoted ubiquitination of NLRP3, and negatively regulated NLRP3 inflammasome activation. Thus, these findings uncovered a new regulatory mechanism of NLRP3 inflammasome activation, which provides a new perspective for the therapy of NLRP3 inflammasome-associated inflammatory diseases.

SENP3 attenuates foam cell formation by deSUMOylating NLRP3 in macrophages stimulated with ox-LDL.

SUMO-specific protease 3 (SENP3) participates in the removal of SUMOylation and maintains the balance of the SUMO system, which ensures normal functioning of substrates and cellular activities. In the present study, we found that SENP3 expression was significantly reduced in ox-LDL-stimulated macrophages. SENP3 overexpression suppressed and SENP3 knockdown promoted macrophage foam cell formation. Moreover, SENP3 inhibited cholesterol uptake, CD36 expression, and NOD-like receptor thermal protein domain-associated protein 3 (NLRP3) inflammasome activation in ox-LDL-stimulated macrophages. Ox-LDL-stimulated NLRP3 SUMOylation was reduced by SENP3. Blocking NLRP3 SUMOylation inhibited foam cell formation and NLRP3 inflammasome activation. Thus, this study revealed that SENP3 inhibits macrophage foam cell formation by deSUMOylating NLRP3 and regulating NLRP3 inflammasome activation, which may provide a potentially innovative approach to treatment of atherosclerosis.

Desloratadine alleviates ALS-like pathology in hSOD1G93A mice via targeting 5HTR2A on activated spinal astrocytes.

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease with progressive loss of motor neurons in the spinal cord, cerebral cortex and brain stem. ALS is characterized by gradual muscle atrophy and dyskinesia. The limited knowledge on the pathology of ALS has impeded the development of therapeutics for the disease. Previous studies have shown that autophagy and astrocyte-mediated neuroinflammation are involved in the pathogenesis of ALS, while 5HTR2A participates in the early stage of astrocyte activation, and 5HTR2A antagonism may suppress astrocyte activation. In this study, we evaluated the therapeutic effects of desloratadine (DLT), a selective 5HTR2A antagonist, in human SOD1G93A (hSOD1G93A) ALS model mice, and elucidated the underlying mechanisms. HSOD1G93A mice were administered DLT (20 mg·kg-1·d-1, i.g.) from the age of 8 weeks for 10 weeks or until death. ALS onset time and lifespan were determined using rotarod and righting reflex tests, respectively. We found that astrocyte activation accompanying with serotonin receptor 2 A (5HTR2A) upregulation in the spinal cord was tightly associated with ALS-like pathology, which was effectively attenuated by DLT administration. We showed that DLT administration significantly delayed ALS symptom onset time, prolonged lifespan and ameliorated movement disorders, gastrocnemius injury and spinal motor neuronal loss in hSOD1G93A mice. Spinal cord-specific knockdown of 5HTR2A by intrathecal injection of adeno-associated virus9 (AAV9)-si-5Htr2a also ameliorated ALS pathology in hSOD1G93A mice, and occluded the therapeutic effects of DLT administration. Furthermore, we demonstrated that DLT administration promoted autophagy to reduce mutant hSOD1 levels through 5HTR2A/cAMP/AMPK pathway, suppressed oxidative stress through 5HTR2A/cAMP/AMPK/Nrf2-HO-1/NQO-1 pathway, and inhibited astrocyte neuroinflammation through 5HTR2A/cAMP/AMPK/NF-κB/NLRP3 pathway in the spinal cord of hSOD1G93A mice. In summary, 5HTR2A antagonism shows promise as a therapeutic strategy for ALS, highlighting the potential of DLT in the treatment of the disease. DLT as a 5HTR2A antagonist effectively promoted autophagy to reduce mutant hSOD1 level through 5HTR2A/cAMP/AMPK pathway, suppressed oxidative stress through 5HTR2A/cAMP/AMPK/Nrf2-HO-1/NQO-1 pathway, and inhibited astrocytic neuroinflammation through 5HTR2A/cAMP/AMPK/NF-κB/NLRP3 pathway in the spinal cord of hSOD1G93A mice.

Purple Sweet Potato Polysaccharide Exerting an Anti-inflammatory Effect via a TLR-Mediated Pathway by Regulating Polarization and Inhibiting the Inflammasome Activation.

Purple sweet potato polysaccharide (PSPP-1) is a novel glucan; this study aimed to examine the anti-inflammatory effect of PSPP-1 and elucidate its potential mechanisms. Lipopolysaccharide (LPS)-induced RAW264.7 was used as the model of inflammation, cell viability, and levels of nitric oxide (NO), reactive oxygen species (ROS), and calcium ion (Ca2+) were analyzed. ELISA and qPCR were used to assess the productions and mRNA expression of cytokines, and Western blotting was used to assess protein expressions in the TLR-mediated pathway, macrophage polarization, and inflammasome activation. The results demonstrated PSPP-1 inhibited cell proliferation and markedly decreased NO, ROS, and Ca2+ levels. Moreover, PSPP-1 suppressed the secretions and mRNA expressions of pro-inflammatory cytokines and increased those of anti-inflammatory cytokines. Furthermore, PSPP-1 could exert anti-inflammatory effects through different pathways mediated by both TLR2 and TLR4, which modulated the expressions of essential proteins in the myeloid differentiation factor 88 (MyD88)-dependent and toll/IL-1 receptor domain-containing adaptor-inducing interferon-ß (TRIF)-dependent signaling pathways. PSPP-1 even regulated the polarization of M1/M2 macrophages and inhibited the nucleotide oligomerization domain-like receptor protein 3 (NLRP3) inflammasome activation. These findings indicate that PSPP-1 can suppress LPS-induced inflammation via multiple pathways and may be a potential agent for therapeutic inflammation-related pathophysiological processes and disorders.

The Use of Schisandrin B to Combat Triple-Negative Breast Cancers by Inhibiting NLRP3-Induced Interleukin-1ß Production.

Triple-negative breast cancer (TNBC) is the most aggressive and fatal breast cancer subtype. Nowadays, chemotherapy remains the standard treatment of TNBC, and immunotherapy has emerged as an important alternative. However, the high rate of TNBC recurrence suggests that new treatment is desperately needed. Schisandrin B (Sch B) has recently revealed its anti-tumor effects in cancers such as cholangiocarcinoma, hepatoma, glioma, and multi-drug-resistant breast cancer. However, there is still a need to investigate using Sch B in TNBC treatment. Interleukin (IL)-1ß, an inflammatory cytokine that can be expressed and produced by the cancer cell itself, has been suggested to promote BC proliferation and progression. In the current study, we present evidence that Sch B can significantly suppress the growth, migration, and invasion of TNBC cell lines and patient-derived TNBC cells. Through inhibition of inflammasome activation, Sch B inhibits interleukin (IL)-1ß production of TNBC cells, hindering its progression. This was confirmed using an NLRP3 inhibitor, OLT1177, which revealed a similar beneficial effect in combating TNBC progression. Sch B treatment also inhibits IL-1ß-induced EMT expression of TNBC cells, which may contribute to the anti-tumor response.

Potassium ion efflux induces exaggerated mitochondrial damage and non-pyroptotic necrosis when energy metabolism is blocked.

Damage-associated molecular patterns (DAMPs) such as extracellular ATP and nigericin (a bacterial toxin) not only act as potassium ion (K+) efflux inducers to activate NLRP3 inflammasome, leading to pyroptosis, but also induce cell death independently of NLRP3 expression. However, the roles of energy metabolism in determining NLRP3-dependent pyroptosis and -independent necrosis upon K+ efflux are incompletely understood. Here we established cellular models by pharmacological blockade of energy metabolism, followed by stimulation with a K+ efflux inducer (ATP or nigericin). Two energy metabolic inhibitors, namely CPI-613 that targets α-ketoglutarate dehydrogenase and pyruvate dehydrogenase (a rate-limiting enzyme) and 2-deoxy-d-glucose (2-DG) that targets hexokinase, are recruited in this study, and Nlrp3 gene knockout macrophages were used. Our data showed that CPI-613 and 2-DG dose-dependently inhibited NLRP3 inflammasome activation, but profoundly increased cell death in the presence of ATP or nigericin. The cell death was K+ efflux-induced but NLRP3-independent, which was associated with abrupt reactive oxygen species (ROS) production, reduction of mitochondrial membrane potential, and oligomerization of mitochondrial proteins, all indicating mitochondrial damage. Notably, the cell death induced by K+ efflux and blockade of energy metabolism was distinct from pyroptosis, apoptosis, necroptosis or ferroptosis. Furthermore, fructose 1,6-bisphosphate, a high-energy intermediate of glycolysis, significantly suppressed CPI-613+nigericin-induced mitochondrial damage and cell death. Collectively, our data show that energy deficiency diverts NLRP3 inflammasome activation-dependent pyroptosis to Nlrp3-independent necrosis upon K+ efflux inducers, which can be dampened by high-energy intermediate, highlighting a critical role of energy metabolism in cell survival and death under inflammatory conditions.

ZDHHC5-mediated NLRP3 palmitoylation promotes NLRP3-NEK7 interaction and inflammasome activation.

The nucleotide-binding domain (NBD), leucine-rich repeat (LRR), and pyrin domain (PYD)-containing protein 3 (NLRP3) inflammasome is a critical mediator of the innate immune response. How NLRP3 responds to stimuli and initiates the assembly of the NLRP3 inflammasome is not fully understood. Here, we found that a cellular metabolite, palmitate, facilitates NLRP3 activation by enhancing its S-palmitoylation, in synergy with lipopolysaccharide stimulation. NLRP3 is post-translationally palmitoylated by zinc-finger and aspartate-histidine-histidine-cysteine 5 (ZDHHC5) at the LRR domain, which promotes NLRP3 inflammasome assembly and activation. Silencing ZDHHC5 blocks NLRP3 oligomerization, NLRP3-NEK7 interaction, and formation of large intracellular ASC aggregates, leading to abrogation of caspase-1 activation, IL-1ß/18 release, and GSDMD cleavage, both in human cells and in mice. ABHD17A depalmitoylates NLRP3, and one human-heritable disease-associated mutation in NLRP3 was found to be associated with defective ABHD17A binding and hyper-palmitoylation. Furthermore, Zdhhc5-/- mice showed defective NLRP3 inflammasome activation in vivo. Taken together, our data reveal an endogenous mechanism of inflammasome assembly and activation and suggest NLRP3 palmitoylation as a potential target for the treatment of NLRP3 inflammasome-driven diseases.

Altered insulin secretion dynamics relate to oxidative stress and inflammasome activation in children with obesity and insulin resistance.

BACKGROUND: Insulin resistance (IR) is considered the main driver of obesity related metabolic complications, and is related to oxidative stress and inflammation, which in turn promote each other. There is currently no specific definition of IR in children, rather, that for adult population is used by pediatric endocrinologists instead. Altered insulin secretion dynamics are associated with worse metabolic profiles and type 2 diabetes mellitus development, thus we aimed to test whether insulin response relates to oxidative stress and inflammation in children. METHODS: We conducted a case-control study, including 132 children classified as follows: 33 children without obesity (Lean); 42 with obesity but no IR according to the American Diabetes Association criteria for adults (OBIR-); 25 with obesity and IR and an early insulin response to an oral glucose tolerance test (OGTT) (EP-OBIR +); 32 with obesity, IR, and a late insulin peak (LP-OBIR +); and studied variables associated with lipid and carbohydrate metabolism, oxidative stress, inflammation and inflammasome activation. RESULTS: The measured parameters of children with obesity, IR, and an early insulin response were similar to those of children with obesity but without IR. It was late responders who presented an impaired antioxidant system and elevated oxidative damage in erythrocytes and plasma, and inflammasome activation at their white blood cells, despite lower classical inflammation markers. Increased uric acid levels seems to be one of the underlying mechanisms for inflammasome activation. CONCLUSIONS: It is insulin response to an OGTT that identifies children with obesity suffering oxidative stress and inflammasome activation more specifically. Uric acid could be mediating this pathological inflammatory response by activating NLRP3 in peripheral blood mononuclear cells.

Transient anxiety-and depression-like behaviors are linked to the depletion of Foxp3-expressing cells via inflammasome in the brain.

Forkhead box P3 (Foxp3) is a transcription factor that influences functioning of regulatory T cells (Tregs) that modulate peripheral immune response. Treg-mediated innate immunity and Treg-mediated adaptive immunity are receiving considerable attention for their implication in mechanisms associated with anxiety and depression. Here, we demonstrated that depletion of Foxp3-expressing cells causally promotes transient anxiety- and depression-like behaviors associated with inflammasome activation in "depletion of regulatory T cell" (DEREG) mice. We found that restoration of Foxp3-expressing cells causally reverses neurobehavioral changes through alteration of innate immune responses as assessed by caspase-1 activity and interleukin-1ß (IL-1ß) release in the hippocampal formation of DEREG mice. Moreover, we found that depletion of Foxp3-expressing cells induces a significant elevation of granulocytes, monocytes, and macrophages in the blood, which are associated with transient expression of the matrix metalloprotease-9. Similarly, we found that depletion of Foxp3-expressing cells in 5xFAD, a mouse model of Alzheimer's disease (AD), exhibits elevated activated caspase-1 and promotion of IL-1ß secretion and increased the level of amyloid-beta (Aß)1-42 and Aß plaque burden in the hippocampal formation that coincided with an acceleration of cognitive decline at a presymptomatic age in the 5xFAD mice. Thus, our study provides evidence supporting the idea that Foxp3 may have a causal influence on peripheral immune responses. This, in turn, can promote an innate immune response within the brain, potentially leading to anxiety- and depression-like behaviors or cognitive decline.

Air pollution induces pyroptosis of human monocytes through activation of inflammasomes and Caspase-3-dependent pathways.

According to the World Health Organization (WHO), air pollution is one of the most serious threats for our planet. Despite a growing public awareness of the harmful effects of air pollution on human health, the specific influence of particulate matter (PM) on human immune cells remains poorly understood. In this study, we investigated the effect of PM on peripheral blood monocytes in vitro. Monocytes from healthy donors (HD) were exposed to two types of PM: NIST (SRM 1648a, standard urban particulate matter from the US National Institute for Standards and Technology) and LAP (SRM 1648a with the organic fraction removed). The exposure to PM-induced mitochondrial ROS production followed by the decrease of mitochondrial membrane potential and activation of apoptotic protease activating factor 1 (Apaf-1), Caspase-9, and Caspase-3, leading to the cleavage of Gasdermin E (GSDME), and initiation of pyroptosis. Further analysis showed a simultaneous PM-dependent activation of inflammasomes, including NLRP3 (nucleotide-binding oligomerization domain-like receptor pyrin domain containing 3) and Caspase-1, followed by cleavage of Gasdermin D (GSDMD) and secretion of IL-1ß. These observations suggest that PM-treated monocytes die by pyroptosis activated by two parallel signaling pathways, related to the inorganic and organic PM components. The release of IL-1ß and expression of danger-associated molecular patterns (DAMPs) by pyroptotic cells further activated the remnant viable monocytes to produce inflammatory cytokines (TNF-α, IL-6, IL-8) and protected them from death induced by the second challenge with PM.In summary, our report shows that PM exposure significantly impacts monocyte function and induces their death by pyroptosis. Our observations indicate that the composition of PM plays a crucial role in this process-the inorganic fraction of PM is responsible for the induction of the Caspase-3-dependent pyroptotic pathway. At the same time, the canonical inflammasome path is activated by the organic components of PM, including LPS (Lipopolysaccharide/endotoxin). PM-induced pyroptosis of human monocytes. Particulate matter (PM) treatment affects monocytes viability already after 15 min of their exposure to NIST or LAP in vitro. The remnant viable monocytes in response to danger-associated molecular patterns (DAMPs) release pro-inflammatory cytokines and activate Th1 and Th17 cells. The mechanism of PM-induced cell death includes the increase of reactive oxygen species (ROS) production followed by collapse of mitochondrial membrane potential (ΔΨm), activation of Apaf-1, Caspase-9 and Caspase-3, leading to activation of Caspase-3-dependent pyroptotic pathway, where Caspase-3 cleaves Gasdermin E (GSDME) to produce a N-terminal fragment responsible for the switch from apoptosis to pyroptosis. At the same time, PM activates the canonical inflammasome pathway, where activated Caspase-1 cleaves the cytosolic Gasdermin D (GSDMD) to produce N-terminal domain allowing IL-1ß secretion. As a result, PM-treated monocytes die by pyroptosis activated by two parallel pathways-Caspase-3-dependent pathway related to the inorganic fraction of PM and the canonical inflammasome pathway dependent on the organic components of PM.

The Implication of Alu cDNA in the Pathogenesis of ARMD.

Age-related macular degeneration (ARMD or AMD) is a progressive, sight-threatening disease. The pathogenesis of ARMD is complex, involving many factors, such as metabolic, functional, genetic, and environmental factors. Recently, long interspersed nuclear element-1 (L1)- mediated reverse transcription (RT) of Alu RNA into cytoplasmic Alu complementary DNA (cDNA) has been associated with retinal pigment epithelium (RPE) destruction. These findings provide a strong input for a new direction in the management of ARMD, as certain human immunodeficiency virus (HIV) drugs, such as nucleoside reverse transcriptase inhibitors (NRTIs), were found to suppress inflammation and protect cells of the retina.

Activation of Inflammasome during Bluetongue Virus Infection.

Bluetongue virus (BTV), a double-stranded RNA virus belonging to the Sedoreoviridae family, provokes an economically important disease in ruminants. In this study, we show that the production of activated caspase-1 and interleukin 1 beta (IL-1ß) is induced in BTV-infected cells. This response seems to require virus replication since a UV-inactivated virus is unable to activate this pathway. In NLRP3-/- cells, BTV could not trigger further IL-1ß synthesis, indicating that it occurs through NLRP3 inflammasome activation. Interestingly, we observed differential activation levels in bovine endothelial cells depending on the tissue origin. In particular, inflammasome activation was stronger in umbilical cord cells, suggesting that these cells are more prone to induce the inflammasome upon BTV infection. Finally, the strength of the inflammasome activation also depends on the BTV strain, which points to the importance of viral origin in inflammasome modulation. This work reports the crucial role of BTV in the activation of the NLRP3 inflammasome and further shows that this activation relies on BTV replication, strains, and cell types, thus providing new insights into BTV pathogenesis.

A positive feedback cycle between the alarmin S100A8/A9 and NLRP3 inflammasome-GSDMD signalling reinforces the innate immune response in Candida albicans keratitis.

OBJECTIVE: Fungal keratitis is a severe sight-threatening ocular infection, without effective treatment strategies available now. Calprotectin S100A8/A9 has recently attracted great attention as a critical alarmin modulating the innate immune response against microbial challenges. However, the unique role of S100A8/A9 in fungal keratitis is poorly understood. METHODS: Experimental fungal keratitis was established in wild-type and gene knockout (TLR4-/- and GSDMD-/-) mice by infecting mouse corneas with Candida albicans. The degree of mouse cornea injuries was evaluated by clinical scoring. To interrogate the molecular mechanism in vitro, macrophage RAW264.7 cell line was challenged with Candida albicans or recombinant S100A8/A9 protein. Label-free quantitative proteomics, quantitative real-time PCR, Western blotting, and immunohistochemistry were conducted in this research. RESULTS: Herein, we characterized the proteome of mouse corneas infected with Candida albicans and found that S100A8/A9 was robustly expressed at the early stage of the disease. S100A8/A9 significantly enhanced disease progression by promoting NLRP3 inflammasome activation and Caspase-1 maturation, accompanied by increased accumulation of macrophages in infected corneas. In response to Candida albicans infection, toll-like receptor 4 (TLR4) sensed extracellular S100A8/A9 and acted as a bridge between S100A8/A9 and NLRP3 inflammasome activation in mouse corneas. Furthermore, the deletion of TLR4 resulted in noticeable improvement in fungal keratitis. Remarkably, NLRP3/GSDMD-mediated macrophage pyroptosis in turn facilitates S100A8/A9 secretion during Candida albicans keratitis, thus forming a positive feedback cycle that amplifies the proinflammatory response in corneas. CONCLUSIONS: The present study is the first to reveal the critical roles of the alarmin S100A8/A9 in the immunopathology of Candida albicans keratitis, highlighting a promising approach for therapeutic intervention in the future.

The cGAS-STING pathway in diabetic retinopathy and age-related macular degeneration.

Diabetic retinopathy and age-related macular degeneration are common retinal diseases with shared pathophysiology, including oxidative stress-induced inflammation. Cellular mechanisms responsible for converting oxidative stress into retinal damage are ill-defined but have begun to clarify. One common outcome of retinal oxidative stress is mitochondrial damage and subsequent release of mitochondrial DNA into the cytosol. This leads to activation of the cGAS-STING pathway, resulting in interferon release and disease-amplifying inflammation. This review summarizes the evolving link between aberrant cGAS-STING signaling and inflammation in common retinal diseases and provides prospective for targeting this system in diabetic retinopathy and age-related macular degeneration. Further defining the roles of this system in the retina is expected to reveal new disease pathology and novel therapeutic approaches.

Circular RNA XRCC5 aggravates glioma progression by activating CLC3/SGK1 axis via recruiting IGF2BP2.

BACKGROUND: Increasing evidences have reported the critical roles of circular RNA (circRNA) in gliomas. Whereas, the role of circXRCC5 in glioma and its underlying molecular mechanism has not been reported. METHODS: The RNA transcripts and protein levels were detected using qRT-PCR, immunohistochemistry (IHC) and in situ hybridization (ISH) assays. Cell proliferation was characterized by CCK-8 and clone formation assays. The formation of NLRP3-inflammasomes was identified using immunofluorescence (IF) and Western blot assays. The cytokines were determined using immunosorbent assay (ELISA) and Western blot assays. The molecular interactions were validated using RIP and pull-down assays. RESULTS: circXRCC5 was over-expressed in glioma and positively related to the shorter survival rate, advanced TNM stage and larger tumor volume. circXRCC5 knockdown inhibited cell proliferation and NLRP3-mediated inflammasome activation of glioma cells. Subsequently, we found that circXRCC5 maintained mRNA stability of CLC3 by binding to IGF2BP2. Furthermore, CLC3 accelerated SGK1 expression via PI3K/PDK1/AKT pathway. The rescue experiments showed that both overexpression of CLC3 or SGK1 dramatically alleviated circXRCC5 knockdown-induced inhibition of cell proliferation and NLRP3-mediated inflammasome activation of glioma cells. In vivo, our study proved that circXRCC5 accelerated glioma growth by regulating CLC3/SGK1 axis. CONCLUSION: Our data concluded that circXRCC5 formed a complex with IGF2BP2 to regulate inflammasome activation and tumor growth via CLC3/SGK1 axis.

Liposomes loaded with quercetin for resolution of lung inflammation in a lipopolysaccharide-induced mouse model of sepsis.

Quercetin (QU) has been widely used as a dietary supplement and proved useful to treat lung diseases. However, the therapeutic potential of QU may be restricted because of its low bioavailability and poor water solubility. In this study, we investigated the effects of developed QU-loaded liposomes on macrophage-mediated lung inflammation.In vivo, a mouse model of sepsis induced by lipopolysaccharide challenge was used to detect the anti-inflammatory effects of liposomal QU. Hematoxylin/eosin staining and immunostaining were utilized to reveal pathological damage and leukocyte infiltration into the lung tissues. Quantitative reverse transcription-polymerase chain reaction and immunoblotting were used to determine cytokine production in the mouse lungs.In vitro, mouse RAW 264.7 macrophages were treated with free QU and liposomal QU. Cell viability assay and immunostaining were utilized to detect cytotoxicity and distribution of QU in the cells. Thein vivoresults showed that liposomal encapsulation promoted the inhibitory effects of QU on lung inflammation. Liposomal QU decreased mortality in septic mice with no obvious toxicity on vital organs. Mechanistically, the anti-inflammatory effects of liposomal QU were associated with inhibition of nuclear factor-kappa B-dependent cytokine production and inflammasome activation in macrophages. Collectively, the results showed that QU liposomes mitigated lung inflammation in septic mice through inhibition of macrophage inflammatory signaling.

Advances in the regulation of inflammasome activation by GBP family in infectious diseases.

Guanylate-binding proteins (GBPs) are a subfamily of interferon-inducible proteins that undertake distinct roles in the the context of bacteria, virus, chlamydia and parasites infections. These proteins exert a notable influence on the progression and outcomes of infectious diseases. Within the realm of host cell-autonomous immunity against pathogens, GBPs have been identified as the regulators of pyroptosis through canonical and noncanonical inflammasome activation pathways. In this review, we summarize the structure and evolution of GBP family members, the canonical and noncanonical inflammasome activation pathways, the roles of GBPs in regulating inflammasome activation, and the mechanisms of GBPs affecting infections induced by different pathogens. We hope to provide new basic research clues for the pathogenesis and diagnosis and treatment of infectious diseases.

Hunting down NLRP3 inflammasome: An executioner of radiation-induced injury.

Radiotherapy is one of the mainstream treatment modalities for several malignancies. However, radiation-induced injury to surrounding normal tissues limits its efficacy. The NLRP3 inflammasome is an essential mechanism of innate immunity that reacts to challenges from endogenous danger signals and pathological microbes. A growing body of evidence has demonstrated a key role of NLRP3 inflammasome in the pathogenesis of radiation-induced tissue injury. Despite accumulating evidence, the potential value of the NLRP3 inflammasome in the management of radiation-induced tissue injury is not adequately recognized. We conducted a literature review to characterize the relationship between NLRP3 inflammasome and radiation injury. By analyzing recent evidence, we identify NLRP3 inflammasome as one of the executioners of radiation-induced injury, since it responds to the challenges of radiation, induces cell pyroptosis and tissue dysfunction, and initiates non-resolving inflammation and fibrosis. Based on these concepts, we propose early intervention/prevention strategies targeting NLRP3 inflammasome in a radiation context, which may help resolve imperative clinical problems.