The Protective Effect of Melatonin on LPS-Induced Myocardial Injury via the Caspase-11/GSDMD Pathway.

BACKGROUND: Melatonin (MT) has been demonstrated to have cardioprotective effects. Nevertheless, the precise mechanism through which MT provides protection against the etiology of LPS-induced myocardial injury remains uncertain. In this investigation, our objective was to explore the impact of MT on LPS-induced myocardial injury in an in vitro setting. METHODS: H9C2 cells were categorized into four groups: a control group (H9C2 group), an MT group, an LPS group, and an MT + LPS group. The H9C2 group received treatment with sterile saline solution, the LPS group was exposed to 5 µg/mL LPS for 24 hours, the MT + LPS group underwent pretreatment with 150 µmol/L MT for 2 hours, followed by exposure to 5 µg/mL LPS for 24 hours, and the MT group received only 150 µmol/L MT for 2 hours. Cell viability and lactate dehydrogenase (LDH) release were assessed using the CCK-8 assay and LDH activity assay, respectively. The levels of reactive oxygen species (ROS) were quantified in each group of cells, and the percentage of propidium iodide (PI)-stained apoptotic cells was determined by flow cytometry. The mRNA levels of caspase11, GSDMD, and IL-18 in each group of cells were quantified. RESULTS: MT treatment significantly protected H9C2 cells from LPS-induced damage, as evidenced by decreased LDH release. LPS treatment markedly increased ROS levels in H9C2 cells, which were subsequently reduced by MT. LPS caused a substantial decrease in superoxide dismutase (SOD) activity and a significant increase in malondialdehyde (MDA) levels, while MT treatment significantly reversed these effects. Additionally, MT markedly enhanced the proportion of viable H9C2 cells compared to LPS-treated controls, as evidenced by the PI staining assay. LPS upregulated both mRNA levels and protein levels of IL-18 in H9C2 cells. However, MT treatment effectively mitigated this LPS-induced increase. Furthermore, MT significantly decreased LPS-induced protein levels of cleaved-caspase 11 and GSDMD-N in H9C2 cells. CONCLUSION: Overall, our findings suggest that MT inhibits the Caspase11-GSDMD signaling pathway via pyroptosis-related proteins (caspase-11 and GSDMD-N) and reduces the expression of inflammation-related cytokines (IL-18), thereby exerting a protective effect on H9C2 cells after LPS injury.

Gasdermin D Mediated Mitochondrial Metabolism Orchestrate Neurogenesis Through LDHA During Embryonic Development.

Regulatory cell death is an important way to eliminate the DNA damage that accompanies the rapid proliferation of neural stem cells during cortical development, including pyroptosis, apoptosis, and so on. Here, the study reports that the absence of GSDMD-mediated pyroptosis results in defective DNA damage sensor pathways accompanied by aberrant neurogenesis and autism-like behaviors in adult mice. Furthermore, GSDMD is involved in organizing the mitochondrial electron transport chain by regulating the AMPK/PGC-1α pathway to target Aifm3. This process promotes a switch from oxidative phosphorylation to glycolysis. The perturbation of metabolic homeostasis in neural progenitor cells increases lactate production which acts as a signaling molecule to regulate the p38MAPK pathway. And activates NF-𝜿B transcription to disrupt cortex development. This abnormal proliferation of neural progenitor cells can be rescued by inhibiting glycolysis and lactate production. Taken together, the study proposes a metabolic axis regulated by GSDMD that links pyroptosis with metabolic reprogramming. It provides a flexible perspective for the treatment of neurological disorders caused by genotoxic stress and neurodevelopmental disorders such as autism.

Atramacronoid A induces the PANoptosis-like cell death of human breast cancer cells through the CASP-3/PARP-GSDMD-MLKL pathways.

Breast cancer is the most common malignant tumor and a major cause of mortality among women worldwide. Atramacronoid A (AM-A) is a unique natural sesquiterpene lactone isolated from the rhizome of Atractylodes macrocephala Koidz (known as Baizhu in Chinese). Our study demonstrated that AM-A triggers a specific form of cell death resembling PANoptosis-like cell death. Further analysis indicated that AM-A-induced PANoptosis-like cell death is associated with the CASP-3/PARP-GSDMD-MLKL pathways, which are mediated by mitochondrial dysfunction. These results suggest the potential of AM-A as a lead compound and offer insights for the development of therapeutic agents for breast cancer from natural products.

The role and mechanism of pyroptosis and potential therapeutic targets in non-alcoholic fatty liver disease (NAFLD).

Non-alcoholic fatty liver disease (NAFLD) is a clinical pathological syndrome characterized by the excessive accumulation of fat within liver cells, which can progress to end-stage liver disease in severe cases, posing a threat to life. Pyroptosis is a distinct, pro-inflammatory form of cell death, differing from traditional apoptosis. In recent years, there has been growing research interest in the association between pyroptosis and NAFLD, encompassing the mechanisms and functions of pyroptosis in the progression of NAFLD, as well as potential therapeutic targets. Controlled pyroptosis can activate immune cells, eliciting host immune responses to shield the body from harm. However, undue activation of pyroptosis may worsen inflammatory responses, induce cellular or tissue damage, disrupt immune responses, and potentially impact liver function. This review elucidates the involvement of pyroptosis and key molecular players, including NOD-like receptor thermal protein domain associated protein 3(NLRP3) inflammasome, gasdermin D (GSDMD), and the caspase family, in the pathogenesis and progression of NAFLD. It emphasizes the promising prospects of targeting pyroptosis as a therapeutic approach for NAFLD and offers valuable insights into future directions in the field of NAFLD treatment.

Inducers and Inhibitors of Pyroptotic Death of Granulosa Cells in Models of Premature Ovarian Insufficiency and Polycystic Ovary Syndrome.

Granulosa cells (GCs), the largest cell population and primary source of steroid hormones in the ovary, are the important somatic ovarian components. They have critical roles in folliculogenesis by supporting oocyte, facilitating its growth, and providing a microenvironment suitable for follicular development and oocyte maturation, thus having essential functions in maintaining female fertility and in reproductive health in general. Pyroptotic death of GCs and associated inflammation have been implicated in the pathogenesis of several reproductive disorders in females including Premature Ovarian Insufficiency (POI) and Polycystic Ovary Syndrome (PCOS). Here, I reviewed factors, either intrinsic or extrinsic, that induce or inhibit pyroptosis in GCs in various models of these disorders, both in vitro and in vivo, and also covered associated molecular mechanisms. Most of these studied factors influence NLRP3 inflammasome- and GSDMD (Gasdermin D)-mediated pyroptosis in GCs, compared to other inflammasomes and gasdermins (GSDMs). I conclude that a more complete mechanistic understanding of these factors in terms of GC pyroptosis is required to be able to develop novel strategies targeting inflammatory cell death in the ovary.

Inhibition of GSDMD-dependent pyroptosis decreased methamphetamine self-administration in rats.

It is widely believed that the activation of the central dopamine (DA) system is crucial to the rewarding effects of methamphetamine (METH) and to the behavioral outcomes of METH use disorder. It was reported that METH exposure induced gasdermin D (GSDMD)-dependent pyroptosis in rats. The membrane pore formation caused by METH-induced pyroptosis may also contribute to the overflow of DA into the extracellular space and subsequently increase the DA levels in the brain. The present study firstly investigated whether the membrane pore information induced by GSDMD-dependent pyroptosis was associated with the increased DA levels in the ventral tegmental area (VAT) and nucleus accumbens (NAc) of rats self-administering METH and SY-SH5Y cells treated by METH. Subsequently, the effect of pore formation blockade or genetic inhibition of GSDMD on the reinforcing and motivational effect of METH was determined in rats, using the animal model of METH self-administration (SA). METH exposure significantly increased the activity of NLRP1/Cas-1/GSDMD pathway and the presence of pyroptosis, accompanied by the significantly increased DA levels in VTA and NAc. Moreover, intraperitoneal injections of disulfiram (DSF) or microinjection of rAAV-shGSDMD into VTA/NAc significantly reduced the reinforcing and motivational effect of METH, accompanied by the decreased level of DA in VTA and NAc. The results provided novel evidence that METH-induced pyroptosis could increase DA release in VTA and NAc via the NLRP1/Cas-1/GSDMD pathway. Additionally, membrane pores or GSDMD blockade could significantly reduce the reinforcing and motivational effect of METH. In conclusion, blocking GSDMD and membrane pore formation could be a promising potential target for the development of agents to treat METH use disorder.

Oridonin ameliorates ocular surface inflammatory responses by inhibiting the NLRP3/caspase-1/GSDMD pyroptosis pathway in dry eye.

Chronic inflammation is one of the central drivers in the development of dry eye disease (DED), in which pyroptosis induced by the NLRP3/caspase-1/gasdermin D (GSDMD) pathway plays a key role. This pathway has become a major target for the treatment of a variety of inflammatory disorders. Oridonin (Ori) is a naturally occurring substance with anti-inflammatory properties obtained from Rabdosia rubescens. Whether Ori can exert an anti-inflammatory effect on DED, and its anti-inflammatory mechanism of action, are still unknown. This experiment is intended to investigate the impact of Ori on the hyperosmolarity-induced NLRP3/caspase-1/GSDMD pyroptosis pathway in immortalized human corneal epithelial (HCE-T) cells, as well as its efficacy and mechanism of action on ocular surface injury in DED mice. Our study showed that Ori could inhibit hyperosmotic-induced pyroptosis through the NLRP3/caspase-1/GSDMD pathway in HCE-T cells, and similarly, Ori inhibited the expression of this pathway in DED mice. Moreover, Ori was protective against hyperosmolarity-induced HCE-T cell damage. In addition, we found that the morphology and number of HCE-T cells were altered under culture conditions of various osmolarities. With increasing osmolarity, the proliferation, migration, and healing ability of HCE-T cells decreased significantly, and the expression of N-GSDMD was elevated. In a mouse model of DED, Ori application inhibited the expression of the NLRP3/caspase-1/GSDMD pyroptosis pathway, improved DED signs and injury, decreased corneal sodium fluorescein staining scores, and increased tear volume. Thus, our study suggests that Ori has potential applications for the treatment of DED, provides potential novel therapeutic approaches to treat DED, and provides a theoretical foundation for treating DED using Ori.

Cadmium activates the innate immune system through the AIM2 inflammasome.

Cadmium (Cd) is a widely used heavy metal and has recently been recognized as a possible source of human toxicity due to its ability to accumulate in organs. Accumulation of heavy metals has several adverse effects, including inducing inflammation, in multiple organs, such as the testis. However, how Cd ions are sensed by host cells and how tissue inflammation eventually occurs remains unclear. Here, we show that Cd activates the AIM2 inflammasome by mediating genomic DNA release into the cytoplasm after DNA damage via oxidative stress, to trigger IL-1ß secretion and pyroptosis. Specifically, the toxicity effects induced by Cd in cells were prevented by melatonin, which served as an antagonist of oxidative stress. Accordingly, in a mouse model, Cd-induced inflammation in the testis and consequential male reproductive dysfunction were effectively reversed by melatonin. Thus, our results suggest a function of AIM2 in Cd-mediated testis inflammation and identify AIM2 as a major pattern recognition receptor in response to heavy metal Cd ions.

Ginsenoside Rg1 Suppresses Pyroptosis via the NF-κB/NLRP3/GSDMD Pathway to Alleviate Chronic Atrophic Gastritis In Vitro and In Vivo.

Chronic atrophic gastritis (CAG) is characterized by the loss of gastric glandular cells, which are replaced by the intestinal-type epithelium and fibrous tissue. Ginsenoside Rg1 (Rg1) is the prevalent ginsenoside in ginseng, with a variety of biological activities, and is usually added to functional foods. As a novel form of programmed cell death (PCD), pyroptosis has received substantial attention in recent years. Despite the numerous beneficial effects, the curative impact of Rg1 on CAG and whether its putative mechanism is partially via inhibiting pyroptosis still remain unknown. To address this gap, we conducted a study to explore the mechanisms underlying the potential anti-CAG effect of Rg1. We constructed a CAG rat model using a multifactor comprehensive method. A cellular model was developed by using 1-methyl-3-nitro-1-nitrosoguanidine (MNNG) combined with Nigericin as a stimulus applied to GES-1 cells. After Rg1 intervention, the levels of inflammatory indicators in the gastric tissue/cell supernatant were reduced. Rg1 relieved oxidative stress via reducing the myeloperoxidase (MPO) and malonaldehyde (MDA) levels in the gastric tissue and increasing the level of superoxide dismutase (SOD). Additionally, Rg1 improved MNNG+Nigericin-induced pyroptosis in the morphology and plasma membrane of the cells. Further research supported novel evidence for Rg1 in the regulation of the NF-κB/NLRP3/GSDMD pathway and the resulting pyroptosis underlying its therapeutic effect. Moreover, by overexpression and knockout of GSDMD in GES-1 cells, our findings suggested that GSDMD might serve as the key target in the effect of Rg1 on suppressing pyroptosis. All of these offer a potential theoretical foundation for applying Rg1 in ameliorating CAG.

Oxymatrine Attenuates Ulcerative Colitis through Inhibiting Pyroptosis Mediated by the NLRP3 Inflammasome.

Ulcerative colitis (UC) is difficult to cure and easy to relapse, leading to poor quality of life for patients. Oxymatrine (OMT) is one of the main alkaloids of Sophora flavescens Aiton, which has many effects, such as anti-inflammation, anti-oxidative stress, and immunosuppression. This study aimed to investigate whether OMT could attenuate ulcerative colitis by inhibiting the NOD-like receptor family pyrin domain containing three (NLRP3) inflammasome-mediated pyroptosis. In this study, the UC rat models were established by 2,4,6-Trinitrobenzenesulfonic acid (TNBS) in vivo, while RAW264.7 cells and peritoneal macrophages were stimulated with Lipopolysaccharides/Adenosine Triphosphate (LPS/ATP) in vitro to simulate pyroptosis models, and Western blotting (WB) and other detection techniques were applied to analyze proteins involved in the NLRP3 inflammasome pathway. Our results showed that OMT alleviated colitis ulcers and pathological damage in the TNBS-induced UC rats and exhibited an inhibitory effect on pyroptosis at the early stage of UC. In the model group, the pyroptosis reached the peak at 24 h after modeling with the contents of active-cysteine-aspartic proteases-1 (caspase-1), Gasdermin D (GSDMD)-N, and cleaved-interleukin-1 beta (IL-1ß) to the highest expression level. Meanwhile, we found that OMT (80 mg kg-1) remarkably decreased the expression levels of NLRP3, active-caspase-1, and cleaved-IL-1ß at 24 h in the lesion tissue from UC rats. Further experiments on cells demonstrated that OMT at concentrations of 100 and 250 µM significantly inhibited cell death caused by NLRP3 inflammasome activation (p < 0.05), downregulated caspase-1, GSDMD, and decreased the levels of active-caspase-1, GSDMD-N, cleaved-IL-1ß in RAW326.7 cells, and peritoneal macrophages. In summary, these results indicated that OMT could attenuate ulcerative colitis through inhibiting pyroptosis mediated by the NLRP3 inflammasome. The inhibition of the NLRP3 inflammasome may be a potential strategy for UC.

Human Coronavirus 229E Infection Inactivates Pyroptosis Executioner Gasdermin D but Ultimately Leads to Lytic Cell Death Partly Mediated by Gasdermin E.

Human coronavirus 229E (HCoV-229E) is associated with upper respiratory tract infections and generally causes mild respiratory symptoms. HCoV-229E infection can cause cell death, but the molecular pathways that lead to virus-induced cell death as well as the interplay between viral proteins and cellular cell death effectors remain poorly characterized for HCoV-229E. Studying how HCoV-229E and other common cold coronaviruses interact with and affect cell death pathways may help to understand its pathogenesis and compare it to that of highly pathogenic coronaviruses. Here, we report that the main protease (Mpro) of HCoV-229E can cleave gasdermin D (GSDMD) at two different sites (Q29 and Q193) within its active N-terminal domain to generate fragments that are now unable to cause pyroptosis, a form of lytic cell death normally executed by this protein. Despite GSDMD cleavage by HCoV-229E Mpro, we show that HCoV-229E infection still leads to lytic cell death. We demonstrate that during virus infection caspase-3 cleaves and activates gasdermin E (GSDME), another key executioner of pyroptosis. Accordingly, GSDME knockout cells show a significant decrease in lytic cell death upon virus infection. Finally, we show that HCoV-229E infection leads to increased lytic cell death levels in cells expressing a GSDMD mutant uncleavable by Mpro (GSDMD Q29A+Q193A). We conclude that GSDMD is inactivated by Mpro during HCoV-229E infection, preventing GSDMD-mediated cell death, and point to the caspase-3/GSDME axis as an important player in the execution of virus-induced cell death. In the context of similar reported findings for highly pathogenic coronaviruses, our results suggest that these mechanisms do not contribute to differences in pathogenicity among coronaviruses. Nonetheless, understanding the interactions of common cold-associated coronaviruses and their proteins with the programmed cell death machineries may lead to new clues for coronavirus control strategies.

Pyroptosis in Diabetic Peripheral Neuropathy and its Therapeutic Regulation.

Pyroptosis is a pro-inflammatory form of cell death resulting from the activation of gasdermins (GSDMs) pore-forming proteins and the release of several pro-inflammatory factors. However, inflammasomes are the intracellular protein complexes that cleave gasdermin D (GSDMD), leading to the formation of robust cell membrane pores and the initiation of pyroptosis. Inflammasome activation and gasdermin-mediated membrane pore formation are the important intrinsic processes in the classical pyroptotic signaling pathway. Overactivation of the NOD-like receptor thermal protein domain associated protein 3 (NLRP3) inflammasome triggers pyroptosis and amplifies inflammation. Current evidence suggests that the overactivation of inflammasomes and pyroptosis may further induce the progression of cancers, nerve injury, inflammatory disorders and metabolic dysfunctions. Current evidence also indicates that pyroptosis-dependent cell death accelerates the progression of diabetes and its frequent consequences including diabetic peripheral neuropathy (DPN). Pyroptosis-mediated inflammatory reaction further exacerbates DPN-mediated CNS injury. Accumulating evidence shows that several molecular signaling mechanisms trigger pyroptosis in insulin-producing cells, further leading to the development of DPN. Numerous studies have suggested that certain natural compounds or drugs may possess promising pharmacological properties by modulating inflammasomes and pyroptosis, thereby offering potential preventive and practical therapeutic approaches for the treatment and management of DPN. This review elaborates on the underlying molecular mechanisms of pyroptosis and explores possible therapeutic strategies for regulating pyroptosis-regulated cell death in the pharmacological treatment of DPN.

Rabies Virus Infection Causes Pyroptosis of Neuronal Cells.

Rabies virus (RABV) is a neurotropic virus that causes fatal neurological disease, raising serious public health issues and attracting extensive attention in society. To elucidate the molecular mechanism of RABV-induced neuronal damage, we used hematoxylin-eosin staining, transmission electron microscopy, transcriptomics analysis, and immune response factor testing to investigate RABV-infected neurons. We successfully isolated the neurons from murine brains. The specificity of the isolated neurons was identified by a monoclonal antibody, and the viability of the neurons was 83.53-95.0%. We confirmed that RABV infection induced serious damage to the neurons according to histochemistry and transmission electron microscope (TEM) scanning. In addition, the transcriptomics analysis suggested that multiple genes related to the pyroptosis pathway were significantly upregulated, including gasdermin D (Gsdmd), Nlrp3, caspase-1, and IL-1ß, as well as the chemokine genes Ccl2, Ccl3, Ccl4, Ccl5, Ccl7, Ccl12, and Cxcl10. We next verified this finding in the brains of mice infected with the rRC-HL, GX074, and challenge virus standard strain-24 (CVS-24) strains of RABV. Importantly, we found that the expression level of the Gsdmd protein was significantly upregulated in the neurons infected with different RABV strains and ranged from 691.1 to 5764.96 pg/mL, while the basal level of mock-infected neurons was less than 100 pg/mL. Taken together, our findings suggest that Gsdmd-induced pyroptosis is involved in the neuron damage caused by RABV infection.

Dexmedetomidine enhances Mitophagy via PINK1 to alleviate hippocampal neuronal Pyroptosis and improve postoperative cognitive dysfunction in elderly rat.

Postoperative cognitive dysfunction (POCD) is a common complication in elderly surgical patients, significantly affecting their quality of life. Dexmedetomidine (Dex), an anesthetic, has shown promise in alleviating POCD, but its underlying mechanism remains unclear. This study aims to explore how Dex improves POCD in aged rats by targeting the PINK1-mediated mitochondrial autophagy pathway, reducing caspase-1/11-GSDMD-induced hippocampal neuronal pyroptosis. Transcriptome sequencing identified 300 differentially expressed genes enriched in the mitochondrial autophagy pathway in Dex-treated POCD rat hippocampal tissue, with Pink1 as a key candidate. In a POCD rat model, Dex treatment upregulated hippocampal PINK1 expression. In vitro experiments using H19-7 rat hippocampal neurons revealed that Dex enhanced mitochondrial autophagy and suppressed neuronal pyroptosis by upregulating PINK1. Further mechanistic validation demonstrated that Dex activated PINK1-mediated mitochondrial autophagy, inhibiting caspase-1/11-GSDMD-induced neuronal pyroptosis. In vivo experiments confirmed Dex's ability to reduce caspase-1/11-GSDMD-dependent hippocampal neuronal pyroptosis and improve postoperative cognitive function in aged rats. Dexmedetomidine improves postoperative cognitive dysfunction in elderly rats by enhancing mitochondrial autophagy via PINK1 upregulation, mitigating caspase-1/11-GSDMD-induced neuronal pyroptosis.

Co-exposure of microcystin-LR and nitrite induced kidney injury through TLR4/NLRP3/GSDMD-mediated pyroptosis.

The degradation of cyanobacterial blooms releases hazardous contaminants such as microcystin-LR (MC-LR) and nitrite, which may collectively exert toxicity on various bodily systems. To evaluate their individual and combined toxicity in the kidney, mice were subjected to different concentrations of MC-LR and/or nitrite over a 6-month period in this study. The results revealed that combined exposure to MC-LR and nitrite exacerbated renal pathological alterations and dysfunction compared to exposure to either compound alone. Specifically, the protein and mRNA expression of kidney injury biomarkers, such as kidney injury molecule 1 (KIM-1) and neutrophil gelatinase-associated lipocalin (NGAL), were notably increased in combined exposure group. Concurrently, co-exposure to MC-LR and nitrite remarkedly upregulated levels of proinflammatory cytokines TNF-α, IL-6 and IL-1ß, while decreasing the anti-inflammatory cytokine IL-10. Notably, MC-LR and nitrite exhibited synergistic effects on the upregulation of renal IL-1ß levels. Moreover, MC-LR combined with nitrite not only elevated mRNA levels of proinflammatory cytokines but also increased protein levels of pyroptosis biomarkers such as IL-1ß, Gasdermin D (GSDMD), and Cleaved-GSDMD. Mechanistic investigations revealed that co-exposure to MC-LR and nitrite promoted pyroptosis both in vivo and in vitro, possibly through the activation of the TLR4/NLRP3/GSDMD pathway. Pretreatment with TLR4 inhibitor and NLRP3 inhibitor effectively suppressed pyroptosis induced by the co-exposure of these two toxins in HEK293T cells. These findings provide compelling evidence that MC-LR combined with nitrite synergistically induces pyroptosis in the kidney by activating the TLR4/NLRP3/GSDMD pathway. Overall, this study significantly enhances our comprehension of how environmental toxins interact and induce harm to the kidneys, offering promising avenues for identifying therapeutic targets to alleviate their toxic effects on renal health.

Effect of IL-9 neutralising antibody on pyroptosis via SGK1/NF-κB/NLRP3/GSDMD in allergic rhinitis mice.

Allergic rhinitis is a common non-infectious inflammatory disease that affects approximately 15 % of people worldwide and has a complex and unclear aetiology. In recent years, pyroptosis has been found to play a role in the development of allergic rhinitis. IL-9, pyroptosis, serum and glucocorticoid-induced protein kinase 1 (SGK1), NOD-like receptor 3 (NLRP3), and nuclear factor kappa B (NF-κB) have been shown to influence each other. Herein, we aimed to explore the role of IL-9 neutralising antibody in pyroptosis involving IL-9, SGK1, NF-κB, and NLRP3 in allergic rhinitis. We observed a decrease in cytokines involved in pyroptosis and gasdermin D (GSDMD) compared with those in mice with allergic rhinitis. Further, phosphorylation of NF-κB/p65 decreased compared with that in mice with allergic rhinitis; NLRP3 and ASC also decreased, although the levels were higher than those in controls. SGK1 levels decreased compared with that in mice with allergic rhinitis and increased after using IL-9 neutralising antibodies, thus demonstrating its negative regulatory effects. The IL-9 neutralising antibody reduced the inflammatory and pyroptosis responses via SGK1 and NF-κB/NLRP3/GSDMD pathway. Our research results indicate that IL-9 regulates allergic rhinitis via the influence of SGK1 and NF-κB/NLRP3/GSDMD signalling pathway, providing new insights for developing novel drugs to treat allergic rhinitis.

Staphylococcus Aureus Membrane Vesicles Kill Tumor Cells Through a Caspase-1-Dependent Pyroptosis Pathway.

Introduction: Nanosized outer membrane vesicles (OMVs) from Gram-negative bacteria have attracted increasing interest because of their antitumor activity. However, the antitumor effects of MVs isolated from Gram-positive bacteria have rarely been investigated. Methods: MVs of Staphylococcus aureus USA300 were prepared and their antitumor efficacy was evaluated using tumor-bearing mouse models. A gene knock-in assay was performed to generate luciferase Antares2-MVs for bioluminescent detection. Cell counting kit-8 and lactic dehydrogenase release assays were used to detect the toxicity of the MVs against tumor cells in vitro. Active caspase-1 and gasdermin D (GSDMD) levels were determined using Western blot, and the tumor inhibition ability of MVs was determined in B16F10 cells treated with a caspase-1 inhibitor. Results: The vesicular particles of S. aureus USA300 MVs were 55.23 ± 8.17 nm in diameter, and 5 µg of MVs remarkably inhibited the growth of B16F10 melanoma in C57BL/6 mice and CT26 colon adenocarcinoma in BALB/c mice. The bioluminescent signals correlated well with the concentrations of the engineered Antares2-MVs (R2 = 0.999), and the sensitivity for bioluminescence imaging was 4 × 10-3 µg. Antares2-MVs can directly target tumor tissues in vivo, and 20 µg/mL Antares2-MVs considerably reduced the growth of B16F10 and CT26 tumor cells, but not non-carcinomatous bEnd.3 cells. MV treatment substantially increased the level of active caspase-1, which processes GSDMD to trigger pyroptosis in tumor cells. Blocking caspase-1 activation with VX-765 significantly protected tumor cells from MV killing in vitro and in vivo. Conclusion: S. aureus MVs can kill tumor cells by activating the pyroptosis pathway, and the induction of pyroptosis in tumor cells is a promising strategy for cancer treatment.

SzM protein of Streptococcus equi ssp. zooepidemicus triggers the release of neutrophil extracellular traps depending on GSDMD.

Streptococcus equissp.zooepidemicus (SEZ) is a crucial pathogen and contributes to various infections in numerous animal species. Swine streptococcicosis outbreak caused by SEZ has been reported in several countries in recent years. SzM protein is a cell membrane-anchored protein, which exhibits as an important virulence factor of SEZ. Effects of SzM protein on host innate immune need further study. Here, recombinant SzM (rSzM) protein of the SEZ was obtained, and mice were intraperitoneally injected with rSzM protein. We discovered that rSzM protein can recruit neutrophils into the injected site. In further study, neutrophils were isolated and treated with rSzM protein, NETs release were triggered by rSzM protein independently, and GSDMD protein was promoted-expressed and activated. In order to investigate the role of GSDMD in NETs formation, neutrophils isolated from WT mice and GSDMD-/- mice were treated with rSzM protein. The results showed that GSDMD deficiency suppressed the NETs release. In conclusion, SzM protein of SEZ can trigger the NETs release in a GSDMD-depending manner.

GSDMD in regulated cell death: A novel therapeutic target for sepsis.

Sepsis is a life-threatening multi-organ dysfunction syndrome caused by an abnormal host response to infection. Regulated cell death is essential for maintaining tissue homeostasis and eliminating damaged, infected, or aging cells in multicellular organisms. Gasdermin D, as a member of the gasdermin family, plays a crucial role in the formation of cytoplasmic membrane pores. Research has found that GSDMD plays important roles in various forms of regulated cell death such as pyroptosis, NETosis, and necroptosis. Therefore, through mediating regulated cell death, GSDMD regulates different stages of disease pathophysiology. This article mainly summarizes the concept of GSDMD, its role in regulated cell death, its involvement in organ damage associated with sepsis-related injuries mediated by regulated cell death via GSDMD activation and introduces potential drugs targeting GSDMD that may provide more effective treatment options for sepsis patients through drug modification.

[Regulation of electroacupuncture on non-canonical pathway of hepatocellular pyroptosis in rats with nonalcoholic fatty liver disease].

OBJECTIVE: To observe the effect and mechanism of electroacupuncture (EA) on non-canonical pathway of hepatocellular pyroptosis in nonalcoholic fatty liver disease (NAFLD). METHODS: Sixty male SD rats were randomly divided into a normal diet group (n=15) and a high fat modeling group (n=45). The rats in the high fat modeling group were fed with customized high fat diet for 8 weeks to establish NAFLD model. Thirty successfully modeled rats were selected and randomly divided into a model group (n=10), an EA group (n=10) and a non-acupoint with shallow needling group (n=10), and 10 rats were randomly selected from the normal diet group as the control group additionally. In the EA group, EA was applied at bilateral "Fenglong" (ST 40) and "Ganshu" (BL 18), with disperse-dense wave, in frequency of 4 Hz/20 Hz and in intensity of 3 mA. In the non-acupoint with shallow needling group, shallow needling was delivered at points 5 mm from bilateral "Fenglong" (ST 40) and "Ganshu" (BL 18), the EA stimulation parameters were same as the EA group. The intervention was given once a day, 20 min a time, 5 days a week for 4 weeks in the two groups. After intervention, the liver morphology was observed by oil red "O" staining, the serum levels of lipopolysaccharide (LPS), interleukin (IL)-1ß, IL-18 and tumor necrosis factor-α (TNF-α) were detected by ELISA, the protein expression of gasdermin D (GSDMD), GSDMD-N, cysteine aspartic acid specific protease-11 (Caspase-11), IL-1ß, IL-18 and TNF-α in liver tissue were detected by Western blot, the mRNA expression of GSDMD, Caspase-11, IL-1ß, IL-18 and TNF-α in liver tissue was detected by real-time PCR in rats of each group. RESULTS: In the model group, vacuoles in different size were found in the hepatocellular cytoplasm, and the fat droplets were in schistose accumulation. Compared with the model group, the hepatocellular fat droplets and the degree of hepatic steatosis were reduced in the EA group and the non-acupoint with shallow needling group. Compared with the control group, the serum levels of LPS, IL-1ß, IL-18 and TNF-α were increased (P<0.01), the protein and mRNA expression of GSDMD, Caspase-11, IL-1ß, IL-18, TNF-α as well as the protein expression of GSDMD-N in the liver tissue were increased (P<0.01) in the model group. Compared with the model group, the serum levels of LPS, IL-1ß, IL-18 and TNF-α were decreased (P<0.01), the protein and mRNA expression of GSDMD, IL-1ß, IL-18 and TNF-α in the liver tissue were decreased (P<0.01), the protein expression of GSDMD-N and the mRNA expression of Caspase-11 in the liver tissue were decreased (P<0.01) in the EA group and the non-acupoint with shallow needling group. Compared with the model group, the protein expression of Caspase-11 in the liver tissue was decreased (P<0.01) in the EA group. Compared with the non-acupoint with shallow needling group, the serum levels of LPS, IL-1ß, IL-18 and TNF-α were decreased (P<0.01), the protein and mRNA expression of GSDMD, Caspase-11, IL-1ß and IL-18 in the liver tissue were decreased (P<0.01), the protein expression of GSDMD-N and the mRNA expression of TNF-α in the liver tissue were decreased (P<0.01) in the EA group. CONCLUSION: EA can inhibit hepatocellular pyroptosis in NAFLD rats, and its mechanism may be related to reducing the serum level of LPS, and down-regulating the expression of the non-canonical pathway related factors i.e. GSDMD, GSDMD-N, Caspase-11, IL-1ß, IL-18 and TNF-α.