Alveolar hypoxia induces organ-specific inflammasome-related inflammation in male mouse lungs.

Inflammation through activation of caspase-1, seems to play a role in pulmonary hypertension induced by alveolar hypoxia. Whether alveolar hypoxia induces caspase-1-mediated inflammation and influx of leukocytes in other organs than the lungs, is not known. Our aim was to explore sites of caspase-1-related inflammation in alveolar hypoxia. Wild type (WT) mice were exposed to environmental hypoxia or room-air, and organs were analyzed. Right heart catheterization was performed after 14 days of alveolar hypoxia in WT mice and mice transplanted with WT or caspase-1-/- bone marrow. Hypoxia induced leukocyte accumulation and increased caspase-1 protein in the lungs, not in other organs. WT mice transplanted with WT or caspase-1-/- bone marrow showed no difference in pulmonary leukocyte accumulation or development of pulmonary hypertension after alveolar hypoxia. Caspase-1 and IL-18 were detected in bronchial epithelium in WT mice, and hypoxia induced IL-18 secretion from bronchial epithelial cells. IL-18 stimulation generated IL-6 mRNA in monocytes. Phosphorylated STAT3 was increased in hypoxic lungs, not in other organs. Alveolar hypoxia induces caspase-1 activation and leukocyte accumulation specific to the lungs, not in other organs. Caspase-1 activation and IL-18 secretion from bronchial epithelial cells might initiate hypoxia-induced inflammation, leading to pulmonary hypertension.

Investigating the involvement of the NLRP3/ASC/caspase-1 and NF-κb/MAPK pathways in the pathogenesis of gouty arthritis: Insights from irradiated and non-irradiated Trifolium alexandrium L. extracts and some metabolites.

ETHNOPHARMACOLOGICAL RELEVANCE: Trifolium alexandrinum L. (TA), has traditionally been used in folk medicine for its anti-inflammatory properties against hyperuricemia and gout. However, the specific mechanisms of action of TA have not been thoroughly studied. AIM OF THE WORK: This study aimed to evaluate the protective effects of irradiated (TR25) and non-irradiated (TR0) Trifolium alexandrinum L. aqueous extract (TAAE), along with two isolated compounds, caffeine (CAF) and saponin (SAP), in a rat model of acute gouty arthritis (GA). MATERIALS AND METHODS: The GA model was established by injecting a monosodium urate (MSU) suspension into the knee joint. Synovial tissue pathology was assessed, and levels of TNF-α, IL-6, IL-1ß, NF-κB, mTOR, AKT1, PI3K, NLRP3, and ASC were measured by ELISA. mRNA expression of ERK1, JNK, and p-38 MAPK was detected using qRT-PCR, and Caspase-1 protein expression was assessed by immunohistochemical analysis. Knee swelling, uric acid levels, liver and kidney function, and oxidative stress markers were also evaluated. RESULTS: TAAE analysis identified 170 compounds, with 73 successfully identified using LC-HR-MS/MS, including caffeine citrate and theasapogenol B glycoside as the main constituents. The studied materials demonstrated significant protective effects against GA. TR25 administration significantly mitigated knee joint circumference compared to other treatments. It demonstrated potential in alleviating hyperuricemia, renal and hepatic impairments induced by MSU crystals. TR25 also alleviated oxidative stress and reduced levels of IL1ß, IL-6, TNF-α, and NF-κB. Weak Caspase-1 immune-positive staining was observed in the TR25 group. TR25 decreased NLRP3 and ASC expression, reducing inflammatory cytokine levels in GA. It effectively inhibited the PI3K, AKT, and mTOR signaling pathways, promoting autophagy. Additionally, TR25 suppressed ERK1, JNK, and p-38 MAPK gene expression in synovial tissue. These effects were attributed to various components in TAAE, such as flavonoids, phenolic acids, tannins, alkaloids, and triterpenes. CONCLUSION: Importantly, irradiation (25 KGy) enhanced the antioxidant effects and phtchemical contents of TAAE. Additionally, TR0, TR25, CAF, and SAP exhibited promising protective effects against GA, suggesting their therapeutic potential for managing this condition. These effects were likely mediated through modulation of the NLRP3/ASC/Caspase-1 and ERK/JNK/p-38 MAPK signaling pathways, as well as regulation of the PI3K/AKT/mTOR pathway. Further research is warranted to fully elucidate the underlying mechanisms and optimize their clinical applications.

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.

Wogonin ameliorates the proliferation, inflammatory response, and pyroptosis in keratinocytes via NOD-like receptor family pyrin domain containing 3/Caspase-1/Gasdermin-D pathway.

BACKGROUND: Psoriasis refers to a highly prevalent and immunologically mediated dermatosis with considerable deterioration in life quality. Wogonin, a sort of flavonoid, has been mentioned to elicit protective activities in skin diseases. However, whether Wogonin is implicated in the treatment of psoriasis and its specific mechanisms are not fully understood. AIM: The present work attempted to elaborate the role of Wogonin during the process of psoriasis and to concentrate on the associated action mechanism. METHODS: Cell counting kit-8 (CCK-8) method was initially applied to assay the viability of human keratinocyte HaCaT cells treated by varying concentrations of Wogonin. To mimic psoriasis in vitro, HaCaT cells were exposed to M5 cytokines. CCK-8 and 5-Ethynyl-2'-deoxyuridine  assays were adopted for the measurement of cell proliferation. Inflammatory levels were examined with enzyme-linked immunosorbent assay. Immunofluorescence staining tested nucleotide-binding oligomerization domain (NOD)-like receptor family pyrin domain containing 3 (NLRP3) and Caspase-1 expressions. Western blot examined the protein expressions of proliferation-, inflammation-, pyroptosis-associated factors, and NLRP3. RESULTS: Wogonin treatment antagonized the proliferation, inflammatory response, and NLRP3/caspase-1/Gasdermin-D (GSDMD)-mediated pyroptosis in M5-challenged HaCaT cells. Besides, NLRP3 elevation partially abrogated the effects of Wogonin on M5-induced proliferation, inflammatory response, and NLRP3/caspase-1/GSDMD-mediated pyroptosis in HaCaT cells. CONCLUSION: In a word, Wogonin might exert anti-proliferation, anti-inflammatory and anti-pyroptosis activities in M5-induced cell model of psoriasis and the blockade of NLRP3/Caspase-1/GSDMD pathway might be recognized as a potential mechanism underlying the protective mechanism of Wogonin in psoriasis, suggesting Wogonin as a prospective anti-psoriasis drug.

Brief Pain Inventory (BPI) Pain Survey Versus Cysteine Protease Caspase-1 (Casp1) Blood Levels Following Midline Laparotomy: A Prospective Randomized Study of Patients With Benign Disease and Patients With Cancer.

BACKGROUND/AIM: There is lack of studies assessing the correlation between pain scales and acute phase immune response (APR) following surgery. The purpose of this work was to assess the correlation between cysteine protease caspase-1 (Casp1) blood levels and two pain scales in a cohort of 56 midline laparotomy (MLa) patients and to assess their link with other cytokines (CYTs). PATIENTS AND METHODS: Blood levels of Casp1 and other CYTs (IL-18, IL-18BP, IL-1ra, IL-6, IL-8, IL-10, IL-1ß) were measured before operation and following surgery in patients with MLa. Pain levels were assessed using the Numerical Rating Scale (NRS) and Brief Pain Inventory (BPI) scale, both preoperatively and postoperatively. RESULTS: Casp1 blood levels showed an increasing trend at postoperative day 1 (POP1) and this increase was almost significant in a linear mixed effect model (LME) analysis (p=0.06). Additionally, Casp1 blood levels were higher in patients with cancer than those with benign disease and correlated with IL-18 blood levels (r=0.24, p=0.007). Furthermore, Casp1 blood levels correlated with BPIsev (severity) score values in MLa patients (r=-0.49, p=0.048). A significant correlation was also observed between Casp1 blood levels and NRS scores in patients with MLa. CONCLUSION: This is the first report to evaluate two pain surveys (NRS and BPI) in MLa patients in relation to blood levels of Casp1 and eight CYTs. This analysis is important in confirming the significant correlation between NRS and BPI pain scales and Casp1 blood levels. Our study is also the first to demonstrate that adequate postoperative analgesia in patients with MLa provides better functional ability and improved patient satisfaction.

Anti-Inflammatory Effect of Atorvastatin and Rosuvastatin on Monosodium Urate-Induced Inflammation through IL-37/Smad3-Complex Activation in an In Vitro Study Using THP-1 Macrophages.

Objective: The pleiotropic effect of hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors (statins) is responsible for potent defense against inflammatory response. This study evaluated the inhibitory effects of HMG-CoA reductase inhibitors on the monosodium urate (MSU)-induced inflammatory response through the regulation of interleukin-37 (IL-37) expression. Methods: Serum was collected from patients with gout (n = 40) and from healthy controls (n = 30). The mRNA and protein expression of the target molecules IL-1ß, IL-37, caspase-1, and Smad3 were measured in THP-1 macrophages stimulated with MSU, atorvastatin, or rosuvastatin using a real-time quantitative polymerase chain reaction and Western blot assay. Transfection with IL-1ß or Smad3 siRNA in THP-1 macrophages was used to verify the pharmaceutical effect of statins in uric-acid-induced inflammation. Results: Serum IL-37 levels in gout patients were significantly higher than in controls (p < 0.001) and was associated with the serum uric acid level (r = 0.382, p = 0.008). THP-1 cells stimulated with MSU markedly induced IL-37 mRNA expression and the transition of IL-37 from the cytoplasm to the nucleus. Recombinant IL-37 treatment dose-dependently inhibited activation of caspase-1 and IL-1ß in MSU-induced inflammation. Atorvastatin and rosuvastatin attenuated caspase-1 activation and mature IL-1ß expression but augmented translocation of IL-37 from the cytoplasm to the nucleus. Atorvastatin and rosuvastatin induced phosphorylation of Smad3 in THP-1 cells treated with MSU crystals. Statins potently attenuated translocation of IL-37 from the cytoplasm to the nucleus in THP-1 macrophages transfected with Smad3 siRNA compared to cells with negative control siRNA. Conclusions: This study revealed that statins inhibit the MSU-induced inflammatory response through phosphorylated Smad3-mediated IL-37 expression in THP-1 macrophages.

Caspase-1 inhibitor CZL80 protects against acute seizures via amplifying the inhibitory neural transmission.

Current anti-seizure medications (ASDs) primarily target ion channels or neurotransmissions; however, their practicability is limited by unwanted side-effects and pharmacoresistance. Cumulative evidence has proposed pro-inflammatory caspase-1 as a potential target for developing ASDs. In this study, we showed that the small-molecular caspase-1 inhibitor CZL80 can prevent seizures in various models including the maximal electroshock (MES), the pentylenetetrazol (PTZ), and the amygdaloid kindled models. Specifically, we discovered that CZL80 prevented death, reduced the duration of generalized seizures, and increased the threshold of generalized seizures in a dose-dependent manner in the MES model. In the PTZ model, CZL80 decreased the seizure stages, prolonged the latency to stage 4 seizures, and decreased the death rate. And in amygdaloid kindled rats, CZL80 inhibited the seizure stages, shortened the durations of both generalized seizures and after-discharges. And the anti-seizure efficacy of CZL80 was diminished in caspase-1 knockout mice. In vitro electrophysiology recordings revealed that CZL80 was able to decreased the excitability of glutamatergic pyramidal neurons, as denoted by reducing the spontaneous neuronal firings and increasing the rheobase injected currents to elicit action potentials. Furthermore, CZL80 was able to increase the amplitudes of inhibitory post-synaptic currents (IPSC), while the excitatory post-synaptic currents (EPSC) were not influenced. Lastly, daily administration of CZL80 for 3 weeks did not influence the normal locomotor functions in mice. In sum, our results highlighted CZL80 as a potential anti-seizure therapy with therapeutic significance.

Mode of cell death in the penile cavernous tissue of type 1 diabetes mellitus rats.

BACKGROUND: Diabetes mellitus commonly causes endothelial cell and smooth muscle cell death in penile cavernous tissue. AIM: The study sought to study the mode of cell death in the penile cavernous tissue in type 1 diabetic rats. METHODS: A total of 36 Sprague Dawley rats 10 weeks of age were randomly divided into 2 groups: a normoglycemic group and type 1 diabetic group (intraperitoneal injection of Streptozotocin (STZ), 60 mg/kg). We randomly selected 6 rats from each group for tests at the end of 11, 14, and 18 weeks of age, respectively. All rats were able to eat and drink freely. The ratio of maximum intracavernous pressure to mean arterial pressure, concentration of serum testosterone, level of nitric oxide in the penile cavernosum, and expression of active caspase-1 (pyroptosis) and active caspase-3 (apoptosis) were determined. OUTCOMES: At the end of weeks 4 and 8 of type 1 diabetes, the proportions of endothelial cells and smooth muscle cells undergoing apoptosis and pyroptosis in penile cavernous tissue are different. RESULTS: The ratio of maximum intracavernous pressure to mean arterial pressure and nitric oxide levels were significantly lower in the 4- and 8-week diabetic groups than in the normoglycemic group (P < .01). Penile endothelial cell pyroptosis (5.67 ± 0.81%), smooth muscle cell apoptosis (23.72 ± 0.48%), total cell pyroptosis (9.67 ± 0.73%), and total apoptosis (10.52 ± 1.45%) were significantly greater in the 4-week diabetic group than in the normoglycemic group (P < .01). The proportion of endothelial cell pyroptosis (24.4 ± 3.69%), endothelial cell apoptosis (22.13 ± 2.43%), total cell pyroptosis (14.75 ± 0.93%), and total apoptosis (14.82 ± 1.08%) in the penile tissues of the 8-week diabetic group were significantly greater than those in the normoglycemic group (P < .01).The 8-week survival proportions of diabetic endothelial cells (38.86 ± 8.85%) and smooth muscle cells (44.46 ± 2.94%) was significantly lower than the 4-week survival proportions of endothelial cells (93.17 ± 8.07%) and smooth muscle cells (75.12 ± 4.76%) (P < .05). CLINICAL TRANSLATION: Inhibition of cell death by different methods at different stages may be the key to the treatment of type 1 diabetes-induced erectile dysfunction. STRENGTHS AND LIMITATIONS: The effect of type 1 diabetes on other types of cell death in penile cavernous tissue needs further study. CONCLUSION: The mode of death of endothelial cells in the cavernous tissue of the penis in the early stage in diabetic rats is dominated by pyroptosis, and the death of smooth muscle cells is dominated by apoptosis. Endothelial cell and smooth muscle cell death are not consistent at different stages of diabetes progression.

Syringic acid suppresses Cutibacterium acnes-induced inflammation in human keratinocytes via regulating the NLRP3/caspase-1/IL-1ß signaling axis by activating PPARγ/Nrf2-antioxidant pathway.

BACKGROUND: Our previous studies have demonstrated a strong relationship betweenCutibacterium acnes(C. acnes), oxidative stress, and acne inflammation. Syringic acid (SA) is a plant widely used for its antimicrobial, anti-inflammatory, and antioxidant activities, but lacking data on acne. This study aims to investigate the effect and mechanism of SA on acne inflammation induced by C. acnes in vitro and in vivo. METHODS: After using the SA to expose HaCaT keratinocytes, we reevaluated the effect of the SA on cell viability, cell apoptosis, ROS, CAT, SOD, and other inflammatory variables in the heat-killed C. acnes-treated HaCaT cells. Next, to induce mice with acne inflammation, ICR mice were given an intradermal injection of live C. acnes into their right ears. The effect of SA on this inflammation was then examined. Moreover, we explored the mechanism of SA on PPARγ/Nrf2 and NLRP3/caspase-1/IL-1ß pathways by ELISA, immunofluorescence microscopy, and western blot assay. RESULTS: Heat-killed C. acnes triggered remarkable cell apoptosis, ROS production, interleukin (IL)-1ß, IL-18, IL-6, and TNF-α release, reduced SOD and CAT activity, and upregulated the expression of proteins in HaCaT cells, including up-regulating IL-1ß, PPARγ, Nrf2, HO-1, NQO1, NLRP3, and caspase-1, whereas SA inhibited these effects by partially impairing PPARγ activation. In addition, PPARγ silencing decreased C. acnes-induced IL-1ß secretion and the production of intracellular ROS, down-regulating the expression of Nrf2. Nrf2 activator (SFN) enhanced anti-inflammatory activity through antioxidant mechanisms, boosting intracellular ROS production, reducing SOD and CAT activity, and promoting the increase in ROS, HO-1, NQO1, and IL-1ß levels, while PPARγ inhibitor (GW662) effectively inhibited this effect in heat-killed C. acnes-treated cells. Finally, SA also exhibited notable improvements in ear redness, swelling, and the expression of PPARγ, NLRP3, and IL-1ß in vivo. CONCLUSIONS: SA inhibited C. acnes-induced inflammation via regulating the NLRP3/caspase-1/IL-1ß signaling axis by activating the PPARγ/Nrf2-antioxidant pathway, suggesting a new treatment possibility for acne vulgaris.

Hypericum perforatum L. attenuates depression by regulating Akkermansia muciniphila, tryptophan metabolism and NFκB-NLRP2-Caspase1-IL1ß pathway.

BACKGROUND: Gut microbiota dysbiosis significantly contributes to progression of depression. Hypericum perforatum L. (HPL) is traditionally used in Europe for treating depression. However, its mechanism remains largely underexplored. PURPOSE: This study aims to investigate the pivotal gut microbiota species and microbial signaling metabolites associated with the antidepressant effects of HPL. METHODS: Fecal microbiota transplantation was used to assess whether HPL mitigates depression through alterations in gut microbiota. Microbiota and metabolic profiling of control, chronic restraint stress (CRS)-induced depression, and HPL-treated CRS mice were examined using 16S rRNA gene sequencing and metabolomics analysis. The influence of gut microbiota on HPL's antidepressant effects was assessed by metabolite and bacterial intervention experiments. RESULTS: HPL significantly alleviated depression symptoms in a manner dependent on gut microbiota and restored gut microbial composition by enriching Akkermansia muciniphila (AKK). Metabolomic analysis indicated that HPL regulated tryptophan metabolism, reducing kynurenine (KYN) levels derived from microbiota and increasing 5-hydroxytryptophan (5-HTP) levels. Notably, supplementation with KYN activated the NFκB-NLRP2-Caspase1-IL1ß pathway and increased proinflammatory IL1ß in the hippocampus of mice with depression. Interestingly, mono-colonization with AKK notably increased 5-hydroxytryptamine (5-HT) and decreased KYN levels, ameliorating depression symptoms through modulation of the NFκB-NLRP2-Caspase1-IL1ß pathway. CONCLUSIONS: The promising therapeutic role of HPL in treating depression is primarily attributed to its regulation of the NFκB-NLRP2-Caspase1-IL1ß pathway, specifically by targeting AKK and tryptophan metabolites.

Tongqiao Yizhi granule repress the nuclear factor kappa-b/nucleotide oligomerization domain-like receptors 3/caspase-1 pyroptosis pathway in the hippocampus to counter vascular dementia in rats.

OBJECTIVE: To explore the mechanism by which Tongqiao Yizhi granule (, TQYZKL) intervenes pyroptosis to treat vascular dementia (VaD) in a rat model. METHODS: The rat model of VaD was established by two-vessel occlusion (2VO). The rats were randomly divided into Sham group, Model group, Nimodipine group, TQYZKL (6.2 g?kg-1?d-1), TQYZKL (12.4 g?kg-1?d-1), TQYZKL (24.8 g?kg-1?d-1). The Morris water maze (MWM) test was carried out to test the learning and memory function; Hematoxylin-eosin staining and transmission electron microscopy (TEM) to observe the pathological damage in the hippocampus; Tunel fluorescence staining to detect neuronal pyroptosis in the hippocampus. The expression levels of pyroptosis-related proteins, namely Golgi peripheral membrane protein p65 (P65), nucleotide oligomerization domain-like receptors 3 (NLRP3), caspase-1 and Gasdermin D (GSDMD), were detected using Western blotting and reverse transcription polymerase chain reaction. Moreover, the serum levels of interleukin-1ß (IL-1ß) and interleukin-18 (IL-18) were determined through the enzyme-linked immunosorbent assay. RESULTS: The study revealed that TQYZKL effectively improved the ability of VaD ratsto learn and memorize, relieved the pathological damage in the hippocampus, restored neuronal morphology, and reduced the expression of pyroptosis-related proteins P65, NLRP3, caspase-1, GSDMD-N, IL-18 and IL-1ß (P < 0.05). CONCLUSION: TQYZKL inhibits neuronal pyroptosis in the hippocampus of VaD rats by regulating nuclear factor kappa-B/NLRP3/caspase-1 signaling pathway, thus exerting a therapeutic effect on VaD in the rats.

Immunohistochemical Expression of Caspase1 and Epidermal Growth Factor Receptor in Invasive Breast Carcinoma and Their Biological and Prognostic Associations.

BACKGROUND: Despite advances in breast carcinoma therapies, drug resistance mechanisms as anti-apoptosis and anti-pyroptosis limit the application of these therapies. This work assesses the immunohistochemical (IHC) expression of Caspase1 and EGFR in breast carcinoma and analyzes their clinicopathological associations as prognostic markers and potential therapeutic targets. Caspase1/EGFR expression patterns are utilized to specify breast carcinoma patients who may benefit from these therapies. METHODS: After reviewing the hematoxylin and eosin-stained slides and the routine breast carcinoma IHC stains (estrogen receptors, progesterone receptors, HER2/NEU, Ki-67) by two pathologists and preparation of tissue microarray blocks, anti-Caspase-1 and EGFR IHC staining was performed using Horseradish Peroxidase (HRP) technique. Intensity and percentage-based scoring was applied dividing the 153 included breast carcinomas into Caspase1-negative and positive expression groups; and EGFR low and overexpression groups. Groups were statistically analyzed in relation to age, tumor size, histological and molecular subtype, grade, nodal status, metastasis/recurrence, TNM stage and Ki-67 proliferation index. Kaplan-Meier's analysis was used to compare disease-free survival (DFS) and overall survival (OS). Combined patterns based on Caspase1 and EGFR expression status were created to stratify patients into prognostic groups. RESULTS: Caspase1 was positive in 54.2% of breast carcinomas and its positivity was significantly associated with smaller tumor size, absence of metastasis/recurrence, luminal A and B molecular subtypes and longer OS (p<0.05). EGFR overexpression was detected in 32.7% of carcinomas and was significantly associated with larger tumor size, TNBLBC and a shorter OS (p<0.05). Caspase1-negative/EGFR-overexpression pattern comprised 14.4% of carcinomas and had the worst prognostic associations including larger tumor size, metastasis/recurrence, TNBLBC subtype and shortest OS (p=0.002, 0.002, 0.004 and ≤0.001 respectively).  Conclusions: Combined Caspase1/EGFR IHC expression may provide a tool for selection of patients who benefit from combined EGFR-inhibitors with miR-155-5p down-regulators or photodynamic therapy via induction of apoptosis/pyroptosis in EGFR-overexpression carcinomas through enhanced Caspase1 signaling.

Release of IL-1ß and IL-18 in human primary bronchial epithelial cells exposed to cigarette smoke is independent of NLRP3.

Cigarette smoke (CS) is a major risk factor for chronic lung diseases and promotes activation of pattern recognition receptors in the bronchial epithelium. NOD-like receptor family, pyrin domain-containing 3 (NLRP3) is a pattern recognition receptor whose activation leads to caspase-1 cleavage, maturation/release of IL-1ß and IL-18, and eventually pyroptosis. Whether the NLRP3 inflammasome participates in CS-induced inflammation in bronchial epithelial cells is still unclear. Herein, we evaluated the involvement of NLRP3 in CS-induced inflammatory responses in human primary bronchial epithelial cells. To this purpose, human primary bronchial epithelial cells were stimulated with CS extracts (CSE) and lytic cell death, caspase activation (-1, -8, -3/7), cytokine release (IL-1ß, IL-18, and IL-8), NLRP3, pro-IL-1ß/pro-IL-18 mRNA, and protein expression were measured. The impact of inhibitors of NLRP3 (MCC950), caspases, and the effect of the antioxidant N-acetyl cysteine were evaluated. We found that CSE increased pro-IL-1ß expression and induced activation of caspase-1 and release of IL-1ß and IL-18. These events were independent of NLRP3 and we found that NLRP3 was not expressed. N-acetyl cysteine reverted CSE-induced caspase-1 activation. Overall, our findings support that the bronchial epithelium may play a central role in the release of IL-1 family cytokines independently of NLRP3 in the lungs of smokers.

Baicalein suppresses Coxsackievirus B3 replication by inhibiting caspase-1 and viral protease 2A.

Myocarditis is an inflammatory disease of the cardiac muscle and one of the primary causes of dilated cardiomyopathy. Group B coxsackievirus (CVB) is one of the leading causative pathogens of viral myocarditis, which primarily affects children and young adults. Due to the lack of vaccines, the development of antiviral medicines is crucial to controlling CVB infection and the progression of myocarditis. In this study, we investigated the antiviral effect of baicalein, a flavonoid extracted from Scutellaria baicaleinsis. Our results demonstrated that baicalein treatment significantly reduced cytopathic effect and increased cell viability in CVB3-infected cells. In addition, significant reductions in viral protein 3D, viral RNA, and viral particles were observed in CVB3-infected cells treated with baicalein. We found that baicalein exerted its inhibitory effect in the early stages of CVB3 infection. Baicalein also suppressed viral replication in the myocardium and effectively alleviated myocarditis induced by CVB3 infection. Our study revealed that baicalein exerts its antiviral effect by inhibiting the activity of caspase-1 and viral protease 2A. Taken together, our findings demonstrate that baicalein has antiviral activity against CVB3 infection and may serve as a potential therapeutic option for the myocarditis caused by enterovirus infection.

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.

The Therapeutic Potential of Neutrophil Extracellular Traps and NLRP3 Inflammasomes in Mycoplasma pneumoniae Pneumonia.

INTRODUCTION: Mycoplasma pneumoniae (MP) is the most common pathogen of community-acquired pneumonia in children. However, the role of neutrophil extracellular traps (NETs) in the pathogenesis of MP is unclear. METHODS: Both the level of NETs were detected between the 60 MP pneumonia patients and 20 healthy controls, whose the clinical characteristics were compared. Additionally, NETs formation induced by community-acquired respiratory distress syndrome (CARDS) toxin was also analyzed through transcriptome sequencing. RESULTS: The levels of cell-free DNA, Cit-H3, and MPO-DNA complexes were significantly increased in the patients with MP pneumonia. Importantly, both cell-free DNA and LDH were higher in hospitalized patients with severity than those without severity. In addition, CARDS toxin induced the NETs formation in vitro and in vivo. Transcriptomics GO and KEGG pathway analysis indicate that NOD like receptor signaling pathway and Toll-like receptor signaling pathway are significantly enriched. Finally, we found that DNase I significantly attenuated the higher levels of Cit-H3, and up-regulation of interleukin-1ß (IL-1ß) and interleukin-18 (IL-18) by down-regulating the expression of NLRP3 and Caspase1(p20) in the lung tissues. DISCUSSION: These results indicate that inhibiting excessive activation of NLRP3 inflammasomes, and NETs formation may alleviate MP pneumonia.

Acute Lung Injury and the NLRP3 Inflammasome.

Acute lung injury (ALI) manifests through harm to the capillary endothelium and alveolar epithelial cells, arising from a multitude of factors, leading to scattered interstitial alterations, pulmonary edema, and subsequent acute hypoxic respiratory insufficiency. Acute lung injury (ALI), along with its more serious counterpart, acute respiratory distress syndrome (ARDS), carry a fatality rate that hovers around 30-40%. Its principal pathological characteristic lies in the unchecked inflammatory reaction. Currently, the main strategies for treating ALI are alleviation of inflammation and prevention of respiratory failure. Concerning the etiology of ALI, NLRP3 Inflammasome is essential to the body's innate immune response. The composition of this inflammasome complex includes NLRP3, the pyroptosis mediator ASC, and pro-caspase-1. Recent research has reported that the inflammatory response centered on NLRP3 inflammasomes plays a key part in inflammation in ALI, and may hence be a prospective candidate for therapeutic intervention. In the review, we present an overview of the ailment characteristics of acute lung injury along with the constitution and operation of the NLRP3 inflammasome within this framework. We also explore therapeutic strategies targeting the NLRP3 inflammasome to combat acute lung injury.

NLRP3 Inflammasome Inhibitors for Antiepileptogenic Drug Discovery and Development.

Epilepsy is one of the most prevalent and serious brain disorders and affects over 70 million people globally. Antiseizure medications (ASMs) relieve symptoms and prevent the occurrence of future seizures in epileptic patients but have a limited effect on epileptogenesis. Addressing the multifaceted nature of epileptogenesis and its association with the Nod-like receptor family pyrin domain containing 3 (NLRP3) inflammasome-mediated neuroinflammation requires a comprehensive understanding of the underlying mechanisms of these medications for the development of targeted therapeutic strategies beyond conventional antiseizure treatments. Several types of NLRP3 inhibitors have been developed and their effect has been validated both in in vitro and in vivo models of epileptogenesis. In this review, we discuss the advances in understanding the regulatory mechanisms of NLRP3 activation as well as progress made, and challenges faced in the development of NLRP3 inhibitors for the treatment of epilepsy.

Hydrogen sulfide mitigates ox­LDL­induced NLRP3/caspase­1/GSDMD dependent macrophage pyroptosis by S­sulfhydrating caspase­1.

Macrophage pyroptosis mediates vascular inflammation and atherosclerosis (AS). Hydrogen sulfide (H2S) exerts a protective role in preventing inflammation and AS. However, its molecular mechanisms of regulating the pyroptosis signaling pathway and inhibiting macrophage pyroptosis remain unexplored. The present study aimed to determine whether H2S mitigates macrophage pyroptosis by downregulating the pyroptosis signaling pathway and S­sulfhydrating caspase­1 under the stimulation of oxidized low­density lipoprotein (ox­LDL), a pro­atherosclerotic factor. Macrophages derived from THP­1 monocytes were pre­treated using exogenous H2S donors sodium hydrosulfide (NaHS) and D,L­propargylglycine (PAG), a pharmacological inhibitor of endogenous H2S­producing enzymes, alone or in combination. Subsequently, cells were stimulated with ox­LDL or the desulfhydration reagent dithiothreitol (DTT) in the presence or absence of NaHS and/or PAG. Following treatment, the levels of H2S in THP­1 derived macrophages were measured by a methylene blue colorimetric assay. The pyroptotic phenotype of THP­1 cells was observed and evaluated by light microscopy, Hoechst 33342/propidium iodide fluorescent staining and lactate dehydrogenase (LDH) release assay. Caspase­1 activity in THP­1 cells was assayed by caspase­1 activity assay kit. Immunofluorescence staining was used to assess the accumulation of active caspase­1. Western blotting and ELISA were performed to determine the expression of pyroptosis­specific markers (NLRP3, pro­caspase­1, caspase­1, GSDMD and GSDMD­N) in cells and the secretion of pyroptosis­related cytokines [interleukin (IL)­1ß and IL­18] in the cell­free media, respectively. The S­sulfhydration of pro­caspase­1 in cells was assessed using a biotin switch assay. ox­LDL significantly induced macrophage pyroptosis by activating the pyroptosis signaling pathway. Inhibition of endogenous H2S synthesis by PAG augmented the pro­pyroptotic effects of ox­LDL. Conversely, exogenous H2S (NaHS) ameliorated ox­LDL­and ox­LDL + PAG­induced macrophage pyroptosis by suppressing the activation of the pyroptosis signaling pathway. Mechanistically, ox­LDL and the DTT increased caspase­1 activity and downstream events (IL­1ß and IL­18 secretion) of the caspase­1­dependent pyroptosis pathway by reducing S­sulfhydration of pro­caspase­1. Conversely, NaHS increased S­sulfhydration of pro­caspase­1, reducing caspase­1 activity and caspase­1­dependent macrophage pyroptosis. The present study demonstrated the molecular mechanism by which H2S ameliorates macrophage pyroptosis by suppressing the pyroptosis signaling pathway and S­sulfhydration of pro­caspase­1, thereby suppressing the generation of active caspase-1 and activity of caspase-1.

Reactive Oxygen Species Modulate Th17/Treg Balance in Chlamydia psittaci Pneumonia via NLRP3/IL-1ß/Caspase-1 Pathway Differentiation.

Chlamydia psittaci pneumonia (CPP) is a lung disease caused by the infection with the Chla-mydia psittaci bacterium, which can lead to severe acute respiratory distress syndrome and systemic symptoms. This study explored the specific mechanisms underlying the impact of reactive oxygen species (ROS) on the Th17/Treg balance in CPP. The levels of ROS and the differentiation ratio of Th17/Treg in the peripheral blood of healthy individuals and CPP patients were measured using ELISA and flow cytometry, respectively. The association between the ROS levels and Th17/Treg was assessed using Pearson correlation analysis. The ROS levels and the Th17/Treg ratio were measured in CD4+ T cells following H2O2 treatment and NLRP3 inhibition. The effects of H2O2 treatment and NLRP3 inhibition on the NLRP3/IL-1ß/caspase-1 pathway were observed using immunoblotting. Compared to the healthy group, the CPP group exhibited increased levels of ROS in the peripheral blood, an elevated ratio of Th17 differentiation, and a decreased ratio of Treg differentiation. ROS levels were positively correlated with the Th17 cell proportion but negatively correlated with the Treg cell proportion. The ROS levels and NLRP3/IL-1ß/caspase-1 expression were up-regulated in CD4+ T cells after H2O2 treatment. Furthermore, there was an increase in Th17 differentiation and a decrease in Treg differentiation. Conversely, the NLRP3/IL-1ß/caspase-1 pathway inhibition reversed the effects of H2O2 treatment, with no significant change in the ROS levels. ROS regulates the Th17/Treg balance in CPP, possibly through the NLRP3/IL-1ß/caspase-1 pathway. This study provides a new perspective on the development of immunotherapy for CPP.