Oligomerization Domain-like Receptor Family Pyrin Research Articles

Peran Penting Inflamasom NLRP3 pada Aterosklerosis

Cardiovascular diseases (CVDs) still contribute as the main cause of mortality and premature mortality worldwide. In Indonesia, CVDs contribute to 35% of the main cause of death in non-communicable diseases followed by diabetes at 6%. The ischemic heart disease and acute ischemic stroke is the main cause of death in Indonesia due to atherosclerosis. Atherosclerosis is a multifactorial cause, with chronic inflammation which causes myocardial infarction and acute ischemic stroke. Research demonstrated that one of the underlying mechanisms of atherosclerosis is inflammation. The current research suggested that inflammation could activate a complex of cytosol proteins, namely nucleotide-binding oligomerization domain-like receptor family pyrin domain containing 3 (NLRP3) inflammasome. The NLRP3 inflammasome is known as a protein sensor as a response to danger signals known as pathogen-associated molecular patterns (PAMPs) or damage/danger-associated molecular patterns (DAMPs). The NLRP3 inflammasome will convert the pro-IL1β and pro-IL-18 into their mature forms which initiates the cell death (pyroptosis) through the cleavage of Caspase-1. This NLRP3 inflammasome has a central role in the initiation and progression of atherosclerosis by affecting the sequences of its cellular and molecular targets. To date, several small molecules and drugs have been identified as NLRP3 inflammasome inhibitory pathways and potential therapeutic agents for atherosclerosis. In this review, we will discuss further the structure of NLRP3 inflammasome and its roles in atherosclerosis, and the potential candidates of the small molecules and drugs targeted in the inflammasome pathway.

Punicalagin attenuates ventricular remodeling after acute myocardial infarction via regulating the NLRP3/caspase-1 pathway

Context Punicalagin has myocardial protection; the mechanism of punicalagin on ventricular remodeling (VR) after acute myocardial infarction (AMI) remains unclear. Objective These studies explore the role and mechanism of punicalagin in preventing and treating VR after AMI. Materials and methods Molecular docking was used to predict the targets of punicalagin. After 2 weeks of AMI model, the SD rats were randomly divided into model, and punicalagin (200, 400 mg/kg, gavage) groups for 4 weeks. Thoracotomy with perforation but no ligature was performed on rats in control group. The protein expression of nucleotide-binding oligomerization domain-like receptor family pyrin domain-containing 3 (NLRP3), apoptosis speck-like protein (ASC), caspase-1, gasdermin D (GSDMD), and GSDMD-N, the mRNA expression of NLRP3, caspase-1, GSDMD, interleukin-1β (IL-1β) and IL-18 were evaluated. Results Punicalagin had binding activities with NLRP3 (Vina score, −5.8), caspase-1 (Vina score, −6.7), and GSDMD (Vina score, −6.7). Punicalagin could improve cardiac function, alleviate cardiac pathological changes, minimize the excessive accumulation of collagen in the left ventricular myocardium (p < 0.01), and inhibit cardiomyocyte apoptosis (p < 0.01). Furthermore, punicalagin could inhibit the overexpression of NLRP3, caspase-1, and GSDMD via immunohistochemistry (p < 0.01). Punicalagin inhibited the protein levels of NLRP3, caspase-1, ASC, GSDMD, and GSDMD-N (p < 0.05, p < 0.01). Punicalagin reduced the mRNA expression of NLRP3, caspase-1, GSDMD, IL-1β and IL-18 (p < 0.05, p < 0.01). Conclusions Punicalagin may provide a useful treatment for the future myocardial protection.

The potential effects of festidinol treatment against the NLRP3 inflammasome and pyroptosis in D-galactose and aluminum chloride-induced Alzheimer's-like pathology in mouse brain.

Alzheimer's disease (AD) is a progressive neurodegenerative disorder that causes cognitive and memory decline. Neuroinflammation is currently considered as being an important pathology in AD. NLRP3, the nucleotide-binding and oligomerization (NOD) domain-like receptor (NLR) family pyrin domain (PYD)-containing 3 (NLRP3) inflammasome is a critical component of the innate immune response, which plays a key role in the development and progression of AD. Therefore, the NLRP3 inflammasome is one of the target treatments for AD. This study aimed to investigate the effect of festidinol, a flavanol isolated from Dracaena conferta, against NLRP3 inflammasome and blood-brain barrier damage in D-galactose and aluminum chloride-induced mice. The induced mice received D-galactose (150mg/kg) and aluminum chloride (10mg/kg) intraperitoneally for 90days to generate cognitive impairment. Festidinol (30mg/kg) and donepezil (5mg/kg) were given by oral gavage for 90days along with the induction. Then, learning and memory behavior, and molecular and morphological changes in the brain, which related to NLRP3 inflammasome, pyroptosis and the blood-brain barrier were measured. The results indicated that festidinol markedly decreased the escape latency and increased the time in the target quadrant in the Morris water maze test. Furthermore, festidinol significantly decreased the ionized calcium-binding adapter molecule 1 (Iba-1) and glial fibrillary acidic protein (GFAP) expression. Festidinol also markedly decreased the NLRP3 inflammasome pathway, interleukin 1 beta (IL-1β), gasdermin-D, N-terminal (GSDMD-N) and caspase-3. Pertinent to the blood-brain barrier, festidinol only decreased tumor necrosis factor-α and matrix metallopeptidase-9, but did not restore the tight junction components. In conclusion, festidinol can restore learning and memory and provide a protective effect against the NLRP3 inflammasome and pyroptosis.

Antisense Oligonucleotide Therapy Decreases IL-1β Expression and Prolongs Survival in Mutant Nlrp3 Mice.

Antisense oligonucleotides (ASOs) are a novel therapeutic strategy that targets a specific gene and suppresses its expression. The cryopyrin-associated periodic syndromes (CAPS) are a spectrum of autoinflammatory diseases characterized by systemic and tissue inflammation that is caused by heterozygous gain-of-function mutations in the nucleotide-binding and oligomerization domain-like receptor (NLR) family pyrin domain containing 3 (NLRP3) gene. The aim of this study was to investigate the efficacy of an Nlrp3-specific ASO treatment in CAPS. An Nlrp3-specific ASO was designed and tested in murine cell lines and bone marrow-derived macrophages (BMDMs) from wild-type and CAPS mouse models. Nlrp3 knock-in mice were treated invivo with Nlrp3-specific ASO, survival was monitored, and expression of organ-specific Nlrp3 and IL-1β was measured. Nlrp3-specific ASO treatment of murine cell lines and BMDMs showed a significant downregulation of Nlrp3 and mature IL-1β protein expression. Ex vivo treatment of Nlrp3 mutant mouse-derived BMDMs with Nlrp3-specific ASO demonstrated significantly reduced IL-1β release. In vivo, Nlrp3-specific ASO treatment of Nlrp3 mutant mice prolonged survival, reduced systemic inflammation, and decreased tissue-specific expression of Nlrp3 and mature IL-1β protein. The results of this study demonstrate that Nlrp3-specific ASO treatment downregulates Nlrp3 expression and IL-1β release in CAPS models, suggesting ASO therapy as a potential treatment of CAPS and other NLRP3-mediated diseases.

Pharmacological Inhibition of Gasdermin D Suppresses Angiotensin II-Induced Experimental Abdominal Aortic Aneurysms.

Gasdermin D, a molecule downstream of the nucleotide-binding oligomerization domain-like receptor family pyrin domain containing inflammasome, forms the membrane pore for the secretion of interleukin (IL)-1β and IL-18, and also mediates pyroptosis. This study was to explore the influence of treatment with disulfiram, a small molecule inhibitor to gasdermin D, on the formation and progression of experimental abdominal aortic aneurysms (AAA). AAAs were induced in 10-week-old male apolipoprotein E deficient mice by subcutaneous infusion of angiotensin II (1000 ng/min/kg body weight) for 28 days via osmotic minipumps. Three days prior to angiotensin II infusion, disulfiram (50 mg/kg) or an equal volume of saline as the vehicle control was administered daily via oral gavage. The influence on experimental AAAs was analyzed by serial measurements of aortic diameters via ultrasonography, grading AAA severity and histopathology at sacrifice. Serum IL-1β and IL-18 levels, systolic blood pressure, total cholesterol, and triglyceride were also measured. Additional experiments assayed the influences on the cell viability and IL-1β secretion of in vitro activated macrophages. Disulfiram significantly reduced the enlargement, incidence, and severity of angiotensin II-induced experimental AAAs with attenuation of medial elastin breaks, mural macrophage accumulation, and systolic blood pressure. The AAA suppression was also associated with reduced systemic levels of IL-1β but not IL-18. However, disulfiram treatment had no impact on body weight gain and lipid levels in aneurysmal mice. Additionally, disulfiram treatment also markedly reduced the secretion of IL-1β from activated macrophages with a limited effect on cell viability in vitro. Gasdermin D inhibition by disulfiram attenuated angiotensin II-induced experimental AAAs with reduced systemic IL-1β levels and in vitro activated macrophage IL-1β secretion. Our study suggests that pharmacological gasdermin D inhibition may have translational potential for limiting clinical AAA progression.

Activation of nucleotide-binding oligomerization domain-like receptor family pyrin domain containing 6 by Porphyromonas gingivalis regulates programmed cell death in epithelium

Background/purposeGingival epithelial cells form a physiological barrier against bacterial invasion. Programmed cell death (PCD) regulated by pathogen precognition receptors (PRRs) lead to tissue destruction and is closely related to inflammatory diseases. The purpose of this study was to investigate whether nucleotide-binding oligomerization domain-like receptor family pyrin domain containing 6 (NLRP6) expresses in periodontal epithelium and induces PCD of epithelial cells infected by Porphyromonas gingivalis (P. gingivalis), therefore involves in periodontitis. Material and methodsThe expression of NLRP6 was detected in periodontal epithelium from human gingival sections and HaCaT cells stimulated by P. gingivalis. NLRP6 was over-expressed by adenovirus infection in HaCaT or knocked down by siRNA in P. gingivalis infected HaCaT, and the cell death was observed by transmission electron microscopy and flow cytometry analysis. In addition, qPCR and Western blot were performed to determine the expression of NLRP6 and the pyroptosis excutors, caspase-1 and gasdermin D. Enzyme-linked immunosorbent assay were performed to detect the secretion of IL-1β and IL-18. ResultsNLRP6 was up-regulated in both gingival epithelium of patients with periodontitis and P. gingivalis infected HaCaT. Over-expression of NLRP6 in HaCaT led to caspase-1 dependent pyroptosis. Interestingly, knockdown of NLRP6 with siRNA followed by P. gingivalis stimulation inhibited pyroptosis and induced apoptosis. ConclusionUp-regulation of NLRP6 by P. gingivalis in HaCaT led to pyroptosis, while knocking down NLRP6 inhibited pyroptosis and induced apoptosis, which indicated this PRR may play a crucial role in periodontitis by regulating PCD in periodontal epithelium.

Ginsenoside Rk2, a dehydroprotopanaxadiol saponin, alleviates alcoholic liver disease via regulating NLRP3 and NLRP6 inflammasome signaling pathways in mice

Heavy alcohol consumption results in alcoholic liver disease (ALD) with inadequate therapeutic options. Here, we first report the potential beneficial effects of ginsenoside Rk2 (Rk2), a rare dehydroprotopanaxadiol saponin isolated from streamed ginseng, against alcoholic liver injury in mice. Chronic-plus-single-binge ethanol feeding caused severe liver injury, as manifested by significantly elevated serum aminotransferase levels, hepatic histological changes, increased lipid accumulation, oxidative stress, and inflammation in the liver. These deleterious effects were alleviated by the treatment with Rk2 (5 and 30 mg/kg). Acting as an nucleotide-binding oligomerization domain-like receptor family pyrin domain-containing 3 (NLRP3) inhibitor, Rk2 ameliorates alcohol-induced liver inflammation by inhibiting NLRP3 inflammasome signaling in the liver. Meanwhile, the treatment with Rk2 alleviated the alcohol-induced intestinal barrier dysfunction via enhancing NLRP6 inflammasome in the intestine. Our findings indicate that Rk2 is a promising agent for the prevention and treatment of ALD and other NLPR3-driven diseases.

Portrayal of NLRP3 Inflammasome in Atherosclerosis: Current Knowledge and Therapeutic Targets.

We are witnessing the globalization of a specific type of arteriosclerosis with rising prevalence, incidence and an overall cardiovascular disease burden. Currently, atherosclerosis increasingly affects the younger generation as compared to previous decades. While early preventive medicine has seen improvements, research advances in laboratory and clinical investigation promise to provide us with novel diagnosis tools. Given the physio-pathological complexity and epigenetic patterns of atherosclerosis and the discovery of new molecules involved, the therapeutic field of atherosclerosis has room for substantial growth. Thus, the scientific community is currently investigating the role of nucleotide-binding and oligomerization domain-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome, a crucial component of the innate immune system in different inflammatory disorders. NLRP3 is activated by distinct factors and numerous cellular and molecular events which trigger NLRP3 inflammasome assembly with subsequent cleavage of pro-interleukin (IL)-1β and pro-IL-18 pathways via caspase-1 activation, eliciting endothelial dysfunction, promotion of oxidative stress and the inflammation process of atherosclerosis. In this review, we introduce the basic cellular and molecular mechanisms of NLRP3 inflammasome activation and its role in atherosclerosis. We also emphasize its promising therapeutic pharmaceutical potential.

Progress on Regulation of NLRP3 Inflammasome by Chinese Medicine in Treatment of Ulcerative Colitis.

Ulcerative colitis (UC) is a chronic, non-specific intestinal disease that not only affects the quality of life of patients and their families but also increases the risk of colorectal cancer. The nucleotide-binding oligomerization domain-like receptor family pyrin domain-containing protein 3 (NLRP3) inflammasome is an important component of inflammatory response system, and its activation induces an inflammatory cascade response that is involved in the development and progression of UC by releasing inflammatory cytokines, damaging intestinal epithelial cells, and disrupting the intestinal mucosal barrier. Chinese medicine (CM) plays a vital role in the prevention and treatment of UC and is able to regulate NLRP3 inflammasome. Many experimental studies on the regulation of NLRP3 inflammasome mediated by CM have been carried out, demonstrating that CM formulae with main effects of clearing heat, detoxifying toxicity, drying dampness, and activating blood circulation. Flavonoids and phenylpropanoids can effectively regulate NLRP3 inflammasome. Other active components of CM can interfere with the process of NLRP3 inflammasome assembly and activation, leading to a reduction in inflammation and UC symptoms. However, the reports are relatively scattered and lack systematic reviews. This paper reviews the latest findings regarding the NLRP3 inflammasome activation-related pathways associated with UC and the potential of CM in treating UC through modulation of NLRP3 inflammasome. The purpose of this review is to explore the possible pathological mechanisms of UC and suggest new directions for development of therapeutic tools.

The role of NLRP3 inflammasome-mediated pyroptosis in ischemic stroke and the intervention of traditional Chinese medicine.

Ischemic stroke (IS) is the second leading cause of death and disability in the world. Pyroptosis, a form of programmed cell death initiated by caspases, participates in the occurrence and development of IS. Because it can increase cell membrane permeability, mediate the release of inflammatory factors, and aggravate inflammation, inhibiting this process can significantly reduce the pathological injury of IS. The nucleotide binding oligomerization domain-like receptor family pyrin domain protein 3 (NLRP3) is a multiprotein complex whose activation is the core link of pyroptosis. In recent years, studies have reported that traditional Chinese medicine (TCM) could regulate pyroptosis mediated by NLRP3 inflammasome through multi-channel and multi-target networks and thus exert the effect against IS. This article reviews 107 papers published in recent years in PubMed, Chinese National Knowledge Infrastructure (CNKI), and WanFang Data in recent years. It has found that the activation factors of NLRP3 inflammasome include ROS, mitochondrial dysfunction, K+, Ca2+, lysosome rupture, and trans-Golgi breakdown. TLR4/NF-κB/NLRP3, ROS/TXNIP/NLRP3, AMPK/Nrf2/NLRP3, DRP1/NLRP3, TAK1/JNK/NLRP3 signaling pathways regulate the initiation and assembly of the NLRP3 inflammasome, subsequently induce pyroptosis, affecting the occurrence and development of IS. TCM can affect the above signaling pathways and regulate the pyroptosis mediated by NLRP3 inflammasome, so as to play a protective role against IS, which provides a new entry point for discussing the pathological mechanism of IS and a theoretical basis for developing TCM treasure house.

Tongxinluo promotes endothelium-dependent arteriogenesis to attenuate diabetic peripheral arterial disease.

Peripheral arterial disease (PAD) has become one of the leading causes of disa-bility and death in diabetic patients. Restoring blood supply to the hindlimbs, especially by promoting arteriogenesis, is currently the most effective strategy, in which endothelial cells play an important role. Tongxinluo (TXL) has been widely used for the treatment of cardio-cerebrovascular diseases and extended for diabetes-related vascular disease. To investigate the effect of TXL on diabetic PAD and its underlying mechanisms. An animal model of diabetic PAD was established by ligating the femoral artery of db/db mice. Laser Doppler imaging and micro-computed tomography (micro-CT) were performed to assess the recovery of blood flow and arteriogenesis. Endothelial cell function related to arteriogenesis and cellular pyroptosis was assessed using histopathology, Western blot analysis, enzyme-linked immuno-sorbent assay and real-time polymerase chain reaction assays. In vitro, human vascular endothelial cells (HUVECs) and human vascular smooth muscle cells (VSMCs) were pretreated with TXL for 4 h, followed by incubation in high glucose and hypoxia conditions to induce cell injury. Then, indicators of HUVEC pyroptosis and function, HUVEC-VSMC interactions and the migration of VSMCs were measured. Laser Doppler imaging and micro-CT showed that TXL restored blood flow to the hindlimbs and enhanced arteriogenesis. TXL also inhibited endothelial cell pyroptosis via the reactive oxygen species/nucleotide-binding oligomerization domain-like receptor family pyrin domain-containing 3/Caspase-1/GSDMD signaling pathway. In addition, TXL restored endothelial cell functions, including maintaining the balance of vasodilation, acting as a barrier to reduce inflammation, and enhancing endothelial-smooth muscle cell interactions through the Jagged-1/Notch-1/ephrin-B2 signaling pathway. Similar results were observed in vitro. TXL has a pro-arteriogenic effect in the treatment of diabetic PAD, and the mechanism may be related to the inhibition of endothelial cell pyroptosis, restoration of endothelial cell function and promotion of endothelial cell-smooth muscle cell interactions.

The Chinese herbal prescription JieZe-1 inhibits caspase-1-dependent pyroptosis induced by herpes simplex virus-2 infection in vitro

ObjectiveJieZe-1 (JZ-1), a Chinese herbal prescription, has an obvious effect on genital herpes, which is mainly caused by herpes simplex virus type 2 (HSV-2). Our study aimed to address whether HSV-2 induces pyroptosis of VK2/E6E7 cells and to investigate the anti-HSV-2 activity of JZ-1 and the effect of JZ-1 on caspase-1-dependent pyroptosis. MethodsHSV-2-infected VK2/E6E7 cells and culture supernate were harvested at different time points after the infection. Cells were co-treated with HSV-2 and penciclovir (0.078125 mg/mL) or caspase-1 inhibitor VX-765 (24 h pretreatment with 100 μmol/L) or JZ-1 (0.078125–50 mg/mL). Cell counting kit-8 assay and viral load analysis were used to evaluate the antiviral activity of JZ-1. Inflammasome activation and pyroptosis of VK2/E6E7 cells were analyzed using microscopy, Hoechst 33342/propidium iodide staining, lactate dehydrogenase release assay, gene and protein expression, co-immunoprecipitation, immunofluorescence, and enzyme-linked immunosorbent assay. ResultsHSV-2 induced pyroptosis of VK2/E6E7 cells, with the most significant increase observed 24 h after the infection. JZ-1 effectively inhibited HSV-2 (the 50% inhibitory concentration = 1.709 mg/mL), with the 6.25 mg/mL dose showing the highest efficacy (95.76%). JZ-1 (6.25 mg/mL) suppressed pyroptosis of VK2/E6E7 cells. It downregulated the inflammasome activation and pyroptosis via inhibiting the expression of nucleotide-binding oligomerization domain-like receptor family pyrin domain-containing protein 3 (P < 0.001) and interferon-γ-inducible protein 16 (P < 0.001), and their interactions with apoptosis-associated speck-like protein containing a caspase recruitment domain, and reducing cleaved caspase-1 p20 (P < 0.01), gasdermin D-N (P < 0.01), interleukin (IL)-1β (P < 0.001), and IL-18 levels (P < 0.001). ConclusionJZ-1 exerts an excellent anti-HSV-2 effect in VK2/E6E7 cells, and it inhibits caspase-1-dependent pyroptosis induced by HSV-2 infection. These data enrich our understanding of the pathologic basis of HSV-2 infection and provide experimental evidence for the anti-HSV-2 activity of JZ-1.Please cite this article as: Liu T, Shao QQ, Wang WJ, Liu TL, Jin XM, Xu LJ, Huang GY, Chen Z. The Chinese herbal prescription JieZe-1 inhibits caspase-1-dependent pyroptosis induced by herpes simplex virus-2 infection in vitro. J Integr Med. 2023; 21(3): 277–288.

Mechanism of Kruppel-Like Factor 4 in Pyroptosis of Nasal Mucosal Epithelial Cells in Mice With Allergic Rhinitis.

Allergic rhinitis (AR) is a chronic nasal inflammation, characterized by nasal epithelial dysfunction. Gene therapy targeting transcription factors is a promising strategy for quenching allergic inflammation, including AR. This study sought to probe the mechanism of Kruppel-like factor 4 (KLF4) in pyroptosis of nasal mucosal epithelial cells (NEpCs) in AR mice and provide targets for AR treatment. AR mouse models were established using sensitization with ovalbumin, followed by injection with short hairpin RNA KLF4 (sh-KLF4). AR symptoms were assessed by the times of sneezing and nose rubbing, hematoxylin-eosin, and periodic acid-Schiff staining. Levels of KLF4, nucleotide-binding oligomerization domain-like receptor family pyrin domain containing 3 (NLRP3), cleaved caspase-1, and N-terminal domain (GSDMD-N) in nasal mucosal tissues were determined by Western blot assay, and levels of interleukin (IL)-1β and IL-18 in nasal lavage fluid were determined by enzyme-linked immunosorbent assay. The binding of KLF4 to the NLRP3 promoter was verified using chromatin immunoprecipitation and dual-luciferase assays. The functional rescue experiment was performed with oe-NLRP3 and sh-KLF4 in AR mice. KLF4 was upregulated in nasal mucosal tissues of AR mice. KLF4 inhibition reduced the times of sneezing and nose rubbing, inflammatory cell infiltration, and goblet cell hyperplasia in nasal mucosal tissues, and levels of NLRP3, cleaved caspase-1, GSDMD-N, IL-1β, and IL-18. KLF4 was enriched on the NLRP3 promoter and improved NLRP3 expression. NLRP3 overexpression reversed the inhibition of sh-KLF4 on pyroptosis of NEpCs in AR mice. KLF4 bound to the NLRP3 promoter and promoted pyroptosis of NEpCs in AR mice via activating NLRP3.

Ethanol extract of Chrysanthemumzawadskii inhibits the NLRP3 inflammasome by suppressing ASC oligomerization in macrophages.

Chrysanthemum zawadskii (C. zawadskii) is used in traditional East Asian medicine for the treatment of various diseases, including inflammatory disease. However, it has remained unclear whether extracts of C. zawadskii inhibit inflammasome activation in macrophages. The present study assessed the inhibitory effect of an ethanol extract of C. zawadskii (CZE) on the activation of the inflammasome in macrophages and the underlying mechanism. Bone marrow-derived macrophages were obtained from wild-type C57BL/6 mice. The release of IL-1β and lactate dehydrogenase in response to nucleotide-binding oligomerization domain-like receptor (NLR) family pyrin domain containing 3 (NLRP3) inflammasome activators, such as ATP, nigericin and monosodium urate (MSU) crystals, was significantly decreased by CZE in lipopolysaccharide (LPS)-primed BMDMs. Western blotting revealed that CZE inhibited ATP-induced caspase-1 cleavage and IL-1β maturation. To investigate whether CZE inhibits the priming step of the NLRP3 inflammasome, we confirmed the role of CZE at the gene level using RT-qPCR. CZE also downregulated the gene expression of NLRP3 and pro-IL-1β as well as NF-κB activation in BMDMs in response to LPS. Apoptosis-associated speck-like protein containing a caspase-recruitment domain (CARD) oligomerization and speck formation by NLRP3 inflammasome activators were suppressed by CZE. By contrast, CZE did not affect NLR family CARD domain-containing protein 4 or absent in melanoma 2 inflammasome activation in response to Salmonella typhimurium and poly(dA:dT) in LPS-primed BMDMs, respectively. The results revealed that three key components of CZE, namely linarin, 3,5-dicaffeoylquinic acid and chlorogenic acid, decreased IL-1β secretion in response to ATP, nigericin and MSU. These findings suggest that CZE effectively inhibited activation of the NLRP3 inflammasome.

Bushen Huoxue Recipe Inhibits Renal Fibrosis by Regulating the Reactive Oxygen Species/Nucleotide-Binding Oligomerization Domain-Like Receptor Family Pyrin Domain-Containing 3-Induced Pyroptosis Pathway

To probe the role and specific mechanism of Bushen Huoxue Recipe in inhibiting renal fibrosis (RF), 150 healthy adult male Sprague-Dawley rats were randomized to sham operated group (SO group), model group (UUO group), conventional treatment group (UUO+EPL group), high-dose Bushen Huoxuefang group (UUO+H group), and low-dose Bushen Huoxue Recipe group (UUO+L group), with 30 mice in each group. The UUO, UUO+EPL, UUO+H, and UUO+L groups showed decreased pathological damage scores in rat renal tissue and increased blood urea nitrogen (BUN), serum creatinine (Cr), interleukin-1β (IL-1β), IL-18, high-sensitivity C-reactive protein (hs-CRP), and caspase-1 levels compared with the SO group. The expression of NLRP3, type III collagen (Col-III), matrix fibronectin (FN), and α-smooth actin (α-SMA) increased (P &lt;0.05). Inflammatory cells aggregated in rat kidney tissue, and some renal tubular cells atrophied, fell off, vacuolized, underwent pyknosis, and the shape of tubules was incomplete. The lumen was enlarged, and the DNA damage was greater. The UUO+EPL, UUO+H and UUO+L groups showed increased pathological damage score of rat renal tissue and decreased expression levels of Cr, BUN, hs-CRP, IL-1β, IL-18, caspase-1, NLRP3, FN, Col-III, and α-SMA than UUO group. After a longer period, the UUO+EPL and UUO+H groups decreased more significantly than the UUO+L group. We conclude that Bushen Huoxue Recipe inhibits RF by regulating reactive oxygen species (ROS)/NLRP3-induced pyroptosis pathway.

Experimental study of cardioprotective effects of Cinnamomi Ramulus and Cinnamomi Cortex formula granules on myocardial ischemia/reperfusion injury in rats based on efficacy of "warming and coordinating heart Yang"

This study aimed to parallelly investigate the cardioprotective activity of Cinnamomi Ramulus formula granules(CRFG) and Cinnamomi Cortex formula granules(CCFG) against acute myocardial ischemia/reperfusion injury(MI/RI) and the underlying mechanism based on the efficacy of &quot;warming and coordinating the heart Yang&quot;. Ninety male SD rats were randomly divided into a sham group, a model group, CRFG low and high-dose(0.5 and 1.0 g·kg~(-1)) groups, and CCFG low and high-dose(0.5 and 1.0 g·kg~(-1)) groups, with 15 rats in each group. The sham group and the model group were given equal volumes of normal saline by gavage. Before modeling, the drug was given by gavage once a day for 7 consecutive days. One hour after the last administration, the MI/RI rat model was established by ligating the left anterior descending artery(LAD) for 30 min ischemia followed by 2 h reperfusion except the sham group. The sham group underwent the same procedures without LAD ligation. Heart function, cardiac infarct size, cardiac patho-logy, cardiomyocyte apoptosis, cardiac injury enzymes, and inflammatory cytokines were determined to assess the protective effects of CRFG and CCFG against MI/RI. The gene expression levels of nucleotide-binding oligomerization domain-like receptor family pyrin domain protein 3(NLRP3) inflammasome, apoptosis-associated speck-like protein containing a CARD(ASC), cysteinyl aspartate specific proteinase-1(caspase-1), Gasdermin-D(GSDMD), interleukin-1β(IL-1β), and interleukin-18(IL-18) were determined by real-time quantitative polymerase chain reaction(RT-PCR). The protein expression levels of NLRP3, caspase-1, GSDMD, and N-GSDMD were determined by Western blot. The results showed that both CRFG and CCFG pretreatments significantly improved cardiac function, decreased the cardiac infarct size, inhibited cardiomyocyte apoptosis, and reduced the content of lactic dehydrogenase(LDH), creatine kinase MB isoenzyme(CK-MB), aspartate transaminase(AST), and cardiac troponin Ⅰ(cTnⅠ). In addition, CRFG and CCFG pretreatments significantly decreased the levels of IL-1β, IL-6, and tumor necrosis factor-α(TNF-α) in serum. RT-PCR results showed that CRFG and CCFG pretreatment down-regulated the mRNA expression levels of NLRP3, caspase-1, ASC, and downstream pyroptosis-related effector substances including GSDMD, IL-18, and IL-1β in cardiac tissues. Western blot revealed that CRFG and CCFG pretreatments significantly decreased the protein expression levels of NLRP3, caspase-1, GSDMD, and N-GSDMD in cardiac tissues. In conclusion, CRFG and CCFG pretreatments have obvious cardioprotective effects on MI/RI in rats, and the under-lying mechanism may be related to the inhibition of NLRP3/caspase-1/GSDMD signaling pathway to reduce the cardiac inflammatory response.

Pharmacological Inhibition of the NLRP3 Inflammasome: Structure, Molecular Activation, and Inhibitor-NLRP3 Interaction.

The nucleotide-binding, oligomerization domain-like receptor family pyrin domain containing 3 (NLRP3) inflammasome is a multiprotein complex that combines sensing, regulation, and effector functions to regulate inflammation in health and disease. NLRP3 is activated by a diverse range of inflammation-instigating signals known as pathogen associated molecular patterns and danger associated molecular patterns. Upon activation, NLRP3 oligomerizes and recruits partner proteins to form a supramolecular platform to process the maturation and release of interleukin (IL)-1β, IL-18, and gasdermin D, major mediators of inflammation and inflammatory cell death termed pyroptosis. The NLRP3 inflammasome has been implicated in the pathogenesis of a wide range of disease conditions, including chronic inflammatory disease that are associated with lifestyle and dietary changes, aging, and environmental exposures, and have become the leading cause of death worldwide. Pharmacological targeting of NLRP3 and signaling demonstrated promising efficacy in ameliorating a list of disease conditions in animal models. These findings underscore the potential and importance of NLRP3 as a druggable target for treating a range of diseases. In this review, recent progress in understanding the structure and mechanism of action of the NLRP3 inflammasome is discussed with focus on pharmacological inhibition of NLRP3 by small molecule inhibitors. New structural and mechanistic insights into NLRP3 activation and inhibitor-NLRP3 interactions would aid in the rational design and pharmacological evaluation of NLRP3 inhibitors for treatment of human disease. SIGNIFICANCE STATEMENT: The NLRP3 inflammasome plays central role in innate immune sensing and control of inflammation. Pharmacological inhibition of NLRP3 demonstrated promising efficacy in a range of diseases in animal models. Recent elucidation of the structure and inhibitor binding of NLRP3 generated new insights into its mode of action and inhibitor-NLRP3 interaction at molecular levels, providing new framework for developing small chemical inhibitors of NLRP3 with improved efficacy and specificity against chronic disease that has become major health concerns worldwide.

The NLRP3 Inflammasome in Age-Related Cerebral Small Vessel Disease Manifestations: Untying the Innate Immune Response Connection.

In this narrative review, we present the evidence on nucleotide-binding and oligomerization (NOD) domain-like receptor (NLR) family pyrin domain (PYD)-containing 3 (NLRP3) inflammasome activation for its putative roles in the elusive pathomechanism of aging-related cerebral small vessel disease (CSVD). Although NLRP3 inflammasome-interleukin (IL)-1β has been implicated in the pathophysiology of coronary artery disease, its roles in cerebral arteriothrombotic micro-circulation disease such as CSVD remains unexplored. Here, we elaborate on the current manifestations of CSVD and its' complex pathogenesis and relate the array of activators and aberrant activation involving NLRP3 inflammasome with this condition. These neuroinflammatory insights would expand on our current understanding of CSVD clinical (and subclinical) heterogenous manifestations whilst highlighting plausible NLRP3-linked therapeutic targets.

Regulatory functional role of NLRP3 inflammasome during Mycoplasma hyopneumoniae infection in swine.

Mycoplasma hyopneumoniae causes enzootic pneumonia, a highly contagious respiratory disease in swine that causes significant economic losses worldwide. It is unknown whether the nucleotide oligomerization domain-like receptor (NLR) family pyrin domain containing 3 (NLRP3) inflammasome regulates the immune response in swine during M. hyopneumoniae infection. The current study utilized an in vivo swine model of M. hyopneumoniae infection to investigate the regulatory functional role of the NLRP3 inflammasome during M. hyopneumoniae infection. Notable histopathological alterations were observed in M. hyopneumoniae-infected swine tissues, which were associated with an inflammatory response and disease progression. Swine M. hyopneumoniae infection was associated with an increase in the expression of the NLRP3 inflammasome, which stimulated pro-inflammatory cytokines such as tumor necrosis factor-alpha, interleukin 18, and interleukin 1 beta (IL-1β). The impact of the NLRP3 inhibitor, MCC950 on NLRP3 and pro-inflammatory cytokines in M. hyopneumoniae-infected swine was examined to investigate the relationship between the NLRP3 inflammasome and M. hyopneumoniae infection. Taken together, our findings provide strong evidence that the NLRP3 inflammasome plays a critical regulatory functional role in M. hyopneumoniae infection in swine.

Baicalin Ameliorates Lung Injury in Rats by Inhibiting NLRP3 Inflammasome Activation via NF-[Formula: see text]B Signaling Pathway.

Hemorrhagic shock (HS) is defined as a reduction in tissue oxygenation and organ dysfunction due to severe blood loss. Lung injury is a frequent complication of HS. Baicalin, isolated from Radix Scutellariae, has been reported to profile the antitumor, anti-oxidative, anti-inflammatory, and antibacterial roles in various pathological processes. Nevertheless, the effects of baicalin on HS-induced lung injury are unclear. This study aims to examine the therapeutic effects of baicalin on lung injury. We first established the lung injury rat models by withdrawing blood in the femoral artery followed by resuscitation. A pathological analysis showed that HS-administrated rats presented severe capillary leakage and pulmonary edema, while baicalin therapy alleviated the symptoms. Baicalin therapy reduced the number of macrophages and neutrophils in bronchoalveolar lavage fluid and decreased the expression and activity of myeloperoxidase (neutrophile infiltration marker) in the lung tissues of HS rats, indicating that baicalin alleviated HS-induced infiltration of inflammatory cells. The secretion of inflammatory cytokines, including interleukin (IL)-1[Formula: see text], IL-6, IL-18, and tumor necrosis factor [Formula: see text] (TNF-[Formula: see text]), as well as the activation of the nucleotide-binding oligomerization domain-like receptor family pyrin domain-containing-3 (NLRP3) inflammasome, were inhibited by baicalin administration. Furthermore, we found that the NF-[Formula: see text]B pathway, a canonical pro-inflammatory pathway, was also blocked after treatment with baicalin in HS-evoked rats, as indicated by the decreased expression of p65 and p65 phosphorylation in the lung tissues. In summary, we infer that baicalin may exert a protective role in HS-induced lung injury by suppressing the activation of NLRP3 inflammasome via the NF-[Formula: see text]B pathway.