NOD-like Receptor Pyrin Domain Containing 3 Research Articles

Effect of electroacupuncture on inflammatory signal expression in local tissues of rats with chronic pelvic pain syndrome based on purinergic 2X7 receptor/NOD-like receptor pyrin domain-containing 3 signal pathway.

To study the expression of inflammatory signal in local prostate tissue of chronic pelvic pain syndrome (CPPS) rats by electroacupuncture (EA) of Guanyuan (CV4), Zhongji (CV3), Huiyang (BL35) and Sanyinjiao (SP6), and to explore the possible mechanism of anti-inflammatory and analgesic effects of EA. METHODS：A total of 36 Sprague-Dawley male rats were randomly divided into three groups: control, model and EA (n=12 rats/group). The CPPS model was made by injection of CFA into ventral lobes of the prostate (0.1 mL). Electric acupuncture apparatus was applied to stimulate Guanyuan (CV4), Zhongji (CV3), bilateral Huiyang (BL35) and Sanyinjiao (SP6) acupoints in EA group. The general condition of rats was observed and the prostate index (PI) was calculated. The thermal pain threshold was collected after each therapeutic course. Histopathological changes of the prostate tissue were examined by hematoxylin-eosin staining method. The expression levels of tumor necrosis factor α (TNF-α), interleukin-1β (IL-1β) and prostaglandin E2 (PGE2) in prostatic homogenates were measured by enzyme linked immunosorbent assay (ELISA). Moreover, the expression levels of purinergic 2X7 receptor (P2X7R), NOD-like receptor pyrin domain-containing 3 (NLRP3), caspase-1 and interleukin-18 (IL-18) mRNA were quantified by quantitative real-time polymerase chain reaction. Compared with control group, the PI of rats increased, and the thermal pain threshold decreased significantly in model group. The morphological structure of prostate tissues of rats in model group was severely damaged with a large number of inflammatory cells infiltration. Additionally, the levels of TNF-α, IL-1β and PGE2 were higher, and the expressions of P2X7R, NLRP3, caspase-1 and IL-18 mRNA were higher than those in control group. After EA treatment, the PI was significantly decreased, the thermal pain threshold was significantly increased, and the tissue damage was significantly improved. The expressions of inflammatory cytokines were lower in EA group, and expression of P2X7R/NLRP3 pathway was down-regulated. The effect of EA at Guanyuan (CV4), Zhongji (CV3), Huiyang (BL35) and Sanyinjiao (SP6) can improve inflammation and pain symptoms of CPPS rats induced by Complete Freund's adjuvant (CFA). This suggests that EA at Guanyuan (CV4), Zhongji (CV3), Huiyang (BL35) and Sanyinjiao (SP6) can produce anti-inflammatory analgesia effect by preventing the activation of P2X7R/NLRP3 signal pathway, inhibit the release of inflammatory cytokines in CPPS rats, which may provide a putative novel target for the treatment of CPPS.

Pectic polysaccharides derived from Hainan Rauwolfia ameliorate NLR family pyrin domain-containing 3-mediated colonic epithelial cell pyroptosis in ulcerative colitis.

Pectic polysaccharides (PPs) could exert functions on ulcerative colitis (UC), which is classified as a nonspecific inflammatory disorder. This study investigated the molecular mechanism of PPs derived from Rauwolfia in UC. First, the dextran sodium sulfate (DSS)-induced mouse colitis models and lipopolysaccharide (LPS)-treated colonic epithelial cell (YAMC) models were established and treated with PP. Subsequently, the effects of PPs on mucosal damages in DSS mice were detected, and the levels of inflammatory cytokines, pyroptosis-related factors, oxidative stress-related markers, and the tight junction-related proteins in the tissues or cells were examined, and the results suggested that PPs ameliorated colonic mucosal damages and cell pyroptosis in DSS mice, and limited colonic epithelial cell pyroptosis in in vitro UC models. Subsequently, the binding relations of retinol-binding protein 4 (RBP4) to miR-124-3p and NLR pyrin domain-containing 3 (NLRP3) were analyzed. miR-124-3p targeted RBP4 and reduced the binding of RBP4 to NLRP3, thus inhibiting NLRP3-mediated pyroptosis. Finally, functional rescue experiments revealed that miR-124-3p suppression or RBP4 overexpression promoted colonic epithelial cell pyroptosis. Collectively, Rauwolfia-derived PPs limited miR-124-3p and targeted RBP4 and reduced the binding potency of RBP4 to NLRP3 to inhibit NLRP3-mediated pyroptosis, resulting in the alleviation of colonic epithelial cell pyroptosis and mucosal damages in UC.

Significance of elevated serum and hepatic NOD-like receptor pyrin domain containing 3 (NLRP3) in hepatitis C virus-related liver disease

NOD-like receptor pyrin domain containing 3 (NLRP3) is a microbial and danger signal sensor that acts as a regulator of inflammation via activation of Caspase-1 (CASP1) and has been identified as a major contributor to human liver diseases. The present study was conducted to investigate the association between NLRP3 and the progression of hepatitis C virus (HCV)-related liver disease. Serum NLRP3 levels were analyzed in 49 patients with chronic HCV infection and 18 healthy controls and liver tissues from 34 patients were examined to assess the protein expression of NLRP3 and its activation marker CASP1 using immunohistochemical staining. The results showed that the median serum NLRP3 levels was significantly higher in HCV-infected patients compared with healthy controls (1040 pg/ml vs 695 pg/ml respectively, P < 0.001) and were positively correlated with hepatic NLRP3 and CASP1 expression (r = 0.749, P < 0.001 and r = 0.557, P = 0.001 respectively). The NLRP3 levels in serum and the liver significantly increased with worsening liver pathology and showed positive correlations with serum aminotransferases levels, HCV viremia, and albumin-bilirubin score (P < 0.05). The receiver operating characteristic curve analysis revealed a high diagnostic performance of serum NLRP3 in determining the extent of liver necroinflammation, fibrosis, and steatosis (area under the curve = 0.951, 0.971, and 0.917 respectively, P < 0.001). In conclusion, NLRP3 plays an important role in liver disease progression during HCV infection via CASP1 activation and might be a promising therapeutic target. Serum NLRP3 could be an additional biomarker for liver inflammation and fibrosis.

Regulation of caries-induced pulp inflammation by NLRP3 inflammasome: A laboratory-based investigation.

To evaluate the expression and function of the nod-like receptor pyrin domain containing 3 (NLRP3) inflammasome in caries induced pulpitis. NLRP3 expression was determined with immunohistochemistry in the dental pulp and qPCR in dental pulp cells (DPCs). THP-1 macrophages expressing the apoptosis-related speck-like protein (ASC) and green fluorescent protein (GFP) fusion protein were used to assess NLRP3 inflammasome activation by live cell imaging, following treatment with lipopolysaccharide (LPS) and lipoteichoic acid (LTA). Caspase I inhibitor was used to confirm inflammasome activation. An ex-vivo pulpitis model in which the DPCs were co-cultured with THP-1 macrophages was used to study the effect of the NLRP3 inflammasome inhibitor (MCC950), and cytokines were measured using ELISA and multiplex array. Data were analysed using the t-test or anova followed by a Bonferroni post hoc test with the level of significance set at p ≤ .05. NLRP3 inflammasome was differentially expressed in dental pulp of sound and carious teeth. Treatment of DPCs with LTA significantly upregulates NLRP3 and IL-1 β-expression (p < .05) and in induces more ASC specks formation compared to LPS. IL-β release in response to LTA treatment is significantly reduced with Caspase I inhibitor suggesting inflammasome dependent mechanism (p < .01). NLRP3-specific inhibitor, MCC950, significantly reduced IL-1β and IL-6 in an ex-vivo pulpitis model (p < .01) but had no effect on IL-8 or matrix metalloproteinase-9 (MMP-9). Expression and upregulation of NLRP3 inflammasome with caries and LTA treatment suggest a role in caries-induced pulpitis. NLRP3 inhibitor attenuated the release of selective inflammatory cytokines and could be a potential treatment target that merit further investigation.

Management of Atrial Fibrillation Using Immunoglobulin Free Light Chains, Novel Biomarkers of Inflammation.

AF is the most common cardiac arrhythmia. There is growing evidence that inflammatory mechanisms play an important role in its pathogenesis; inflammasome activation contributes to the onset and progression of AF. An increase in NOD-like-receptor-pyrin domain-containing-3 (NLRP3) inflammasome activation releases proinflammatory cytokines that activate nuclear factor (NF)-κB, which regulates the production of immunoglobulin free light chains (FLCs). Serum FLC levels are increased in patients with AF, and FLCs are biomarkers of inflammation. Inflammasomes and NF-κB may be targets for anti-inflammatory strategies to prevent and treat AF when FLC levels are elevated. This review discusses the role of inflammation in the pathogenesis of AF, as well as FLCs as novel inflammatory biomarkers for the management of AF.

NLRP3 mediates trophoblastic inflammasome activation and protects against Listeria monocytogenes infection during pregnancy

Intrauterine Listeria monocytogenes (L. monocytogenes) infections pose a major threat during pregnancy via affecting placental immune responses. However, the underlying mechanisms of placental defense against this pathogen remain ill-defined. Therefore, this study aims to investigate the function and the mechanism of inflammasomes on against L. monocytogenes infection during pregnancy. A listeriosis murine model and cell culture system was used to investigate the role of trophoblastic nucleotide-binding oligomerization domain-like receptor pyrin domain-containing 3 (NLRP3) in orchestrating innate immune responses to L. monocytogenes infection. Caspase-1 activity was determined using a caspase-1 activity colorimetric kit. NLRP3 and apoptosis-associated speck-like protein containing a CARD (ASC) in placental tissue was detected by immunohistochemistry. NLRP3 in HTR-8/SVneo cells was also detected by immunofluorescence. The expression of interleukin 1β (IL-1β), NLRP3, ASC, and caspase-1 was detected by Western blot. We characterized the NLRP3 inflammasome in trophoblast cells according to whether L. monocytogenes infection increased the activation of caspase-1 and the release of IL-1β. For human or mouse IL-1β in the culture supernatants and mouse tissue lysates were analyzed using ELISA Kits. Trophoblast cells constitutively expressed the components of the NLRP3 inflammasome. In vitro, L. monocytogenes triggers NLRP3 inflammasome activation in trophoblast cells by inducing caspase-1 activation, increasing the NLRP3 protein levels, IL-1β maturation and secretion in HTR-8/SVneo cells. In vivo, L. monocytogenes induces fetal resorption and IL-1β processing in pregnant mice. In addition, NLRP3-deficient mice were more prone to fetal loss than their wild-type counterparts following infection with L. monocytogenes at a lower infective dose. We conclude that trophoblast cells respond to L. monocytogenes infection through the NLRP3 receptor, resulting in inflammasome activation and IL-1β production, which prevents listeriosis during pregnancy.

Acacetin improves cognitive function of APP/PS1 Alzheimer's disease model mice via the NLRP3 inflammasome signaling pathway.

Acacetin (5,7-dihydroxy-4'-methoxyflavone), one of the main extractions from Saussurea involucrata, has anti-inflammatory effects. Our previous study found that acacetin inhibited the Nod-like receptor pyrin domain containing 3 (NLRP3) signaling pathway after cerebral ischemia-reperfusion injury. NLRP3 inflammasome plays a role in Alzheimer's disease (AD) process. However, few studies have examined the effects of acacetin in AD. We randomly divided APP swe/PS1dE9 double transgenic mice into acacetin group (intraperitoneal injection of 25 mg/kg acacetin) and AD model group (intraperitoneal injection of same volume of saline). C57BL/6 mice were selected as control group (same treatment with AD model group). After treating for 30 days, a Morris water maze test was conducted to evaluate spatial learning and memory of the mice. Senile plaque (SP) formation was evaluated by immunohistochemistry. NLRP3 inflammasome-related inflammatory factors and amyloid-β-42 were detected by Western blot or enzyme-linked immunosorbent assay. Acacetin improved spatial learning and memory of AD mice and reduced APP/β expression, thereby decreasing SP formation in the brain. Acacetin also reduced the expression of NLRP3, cysteinyl aspartate-specific proteinase 1 (caspase-1), and interleukin-1β (IL-1β) and the release of inflammatory factors, tumor necrosis factor-α (TNF-α) and IL-1β. Acacetin improved the learning and memory abilities of AD mice and exerted a protective effect on AD by inhibiting the NLRP3 signaling pathway and reducing SP formation.

IL-18BP Alleviates Anxiety-Like Behavior Induced by Traumatic Stress via Inhibition of the IL-18R-NLRP3 Signaling Pathway in a Mouse Model of Hemorrhagic Shock and Resuscitation.

Psychological distress and posttraumatic stress, including anxiety, severely influence life quality. Previously, we reported that interleukin-18 (IL-18) was involved in pyroptosis-induced emotional changes in a rodent model of hemorrhagic shock and resuscitation (HSR). Here, we aimed to continue our investigation on the role of IL-18 binding protein (IL-18BP), which exhibits excellent anti-inflammatory effects as an IL-18 negative regulator. Mice were administered with an intraperitoneal injection of IL-18BP after HSR exposure and anxiety-like behavior was examined using the open-field test and elevated plus maze test. Moreover, the following variables post-HSR were measured: (1) the activation of astrocytes; (2) pyroptosis-associated factors including cleaved caspase-1, GSDMD, IL-18; (3) the roles of IL-18 receptor (IL-18R)-NOD-like receptor pyrin domain-containing-3 (NLRP3) signal with the application of the NLRP3 specific agonist or astrocyte-specific NLRP3 knockout mice. IL-18BP administration remarkably alleviated HSR-induced anxiety-like behavior, astrocytic activation, and increases in pyroptosis-associated factors, while NLRP3 agonist nigericin partially reversed IL-18BP-induced neuroprotective effects. Astrocyte-specific NLRP3 knockout mice exhibited relatively less anxiety-like behavior. Similarly, IL-18BP exhibited an anti-pyroptosis effect in astrocytes in an in vitro model of low oxygen-glucose deprivation. These findings offer unique perspectives on HSR-induced posttraumatic stress and indicate that inhibition of IL-18R-NLRP3 signal via IL-18BP can attenuate astrocytic activation and pyroptosis, broadening the therapeutic landscape for patients with psychological distress and posttraumatic stress.

Ferulic acid relieved ulcerative colitis by inhibiting the TXNIP/NLRP3 pathway in rats.

Ulcerative colitis (UC) is a disorder of the bowel that is characterized by a chronic inflammatory response. The traditional Chinese herbal medicine ferulic acid (FA) is known for its antioxidant, antiapoptotic, and antiinflammatory properties. However, its role in UC is still unclear. Thus, the current study was conducted to investigate the role of FA in UC. Rats were treated with 2,4,6-triabrobenzene sulfonic acid to induce UC and subjected to FA. Human intestinal microvascular endothelial cells (HIMECs) were treated with tumor necrosis factor-α (TNF-α) and pretreated with FA. Pathological changes in colonic tissues were visualized via hematoxylin-eosin staining. Enzyme linked immunosorbent assay was conducted to detect interleukin (IL)-6, IL-12, and IL-1β levels. Cell morphology was visualized by using a microscope, and viability was detected by using MTT. The percentage of apoptosis was detected via flow cytometry. Western blot analysis was performed to detect the expression of the apoptosis-related proteins thioredoxin-interacting protein (TXNIP) and NOD-like receptor pyrin domain-containing 3 (NLRP3). In vivo FA administration alleviated intestinal injury in UC rats and inhibited inflammatory factor levels (IL-6, IL-12, and IL-1β), apoptosis-related protein expression (caspase-1 and caspase-3) and the TXNIP/NLRP3 signaling pathway. In vitro, TNF-α treatment reduced HIMEC viability, increased cell apoptosis and inflammatory factor levels and activated the TXNIP/NLRP3 signaling pathway. However, FA treatment restored the viability of HIMECs, reduced TNF-α-induced cell apoptosis and inflammation and inhibited the TXNIP/NLRP3 signaling pathway. Furthermore, with increasing FA concentration, the effects were stronger. In summary, FA inhibits the inflammatory injury of endothelial cells in ulcerative colitis or alleviates TNF-α-induced HIMEC injury by inhibiting the TXNIP/NLRP3 signaling pathway.

Long noncoding RNA maternally expressed gene 3 improves trophoblast dysfunction and inflammation in preeclampsia through the Wnt/β-Catenin/nod-like receptor pyrin domain-containing 3 axis.

Inadequate trophoblastic infiltration and resulting placental hypoxia and inflammation comprise the core pathological basis of preeclampsia (PE). Maternally expressed gene 3 (MEG3) is known to be involved in the pathogenesis of preeclampsia by inhibiting the migration and invasion of trophoblasts and promoting their apoptosis. Nevertheless, the specific underlying downstream molecular mechanism of MEG3 is less well characterized. In this study, we detected lower expression levels of MEG3 and β-Catenin and higher expression of nod-like receptor pyrin domain-containing 3 (NLRP3) in placental tissues of pregnant women with severe preeclampsia (sPE) than in normal pregnancies. Elevated serum levels of IL-1β and TNF-α were also observed in the sPE group. Then, we established a hypoxia/reoxygenation (H/R) model to mimic preeclampsia. Similar results with sPE group were found in the H/R group compared with the control group. In addition, suppressive trophoblast proliferation, migration and invasion and increases in the apoptotic rate and inflammation were also detected in the H/R group. Notably, overexpressing MEG3 markedly improved trophoblast dysfunction and inflammation caused by H/R. However, the effects of MEG3 on trophoblasts, whether upregulated or downregulated, can be reversed by DKK-1 (Wnt/β-Catenin inhibitor) and MCC950 (NLRP3 inhibitor). The current study revealed that MEG3 regulates trophoblast function and inflammation through the Wnt/β-Catenin/NLRP3 axis and provided new insights into the pathogenesis of preeclampsia.

Sodium Danshensu protects against oxygen glucose deprivation/reoxygenation-induced astrocytes injury through regulating NOD-like receptor pyrin domain containing 3 (NLRP3) inflammasome and tuberous sclerosis complex-2 (TSC2)/mammalian target of rapamycin (mTOR) pathways.

Cerebral ischemic stroke is a serious condition with high incidence, mortality, and associated disability. Currently, effective therapeutic options are available for ischemic stroke are limited. Accumulating evidence indicates that sodium Danshensu, mono sodium compound derived from Salvia miltiorrhiza, plays protective roles in ischemic stroke. However, the underlying protective mechanism of sodium Danshensu in cerebral ischemic stroke remains unknown. In the current study, we explored the role and mechanism of sodium Danshensu on astrocytes exposed to oxygen-glucose deprivation/reoxygenation (OGD/R), which mimics the process of ischemia-reperfusion. The impact of sodium Danshensu on cell viability and apoptosis after OGD/R were evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-dophenyl tetrazolium bromide (MTT) assay and flow cytometry. Quantitative reverse transcription polymerase chain reaction (qRT-PCR) and western blot were used to detect the expression of target messenger RNA (mRNA) and proteins associated with apoptosis and autophagy. The release of lactate dehydrogenase (LDH) was determined, and the production of proinflammatory cytokines were detected using enzyme-linked immunosorbent assay (ELISA) kits. It was found that sodium Danshensu could significantly increase cell viability and decrease LDH release and apoptosis. Besides, it inhibited the production of proinflammatory cytokines, including tumor necrosis factor-α (TNF-α), interleukin (IL)-1β, and IL-6. Sodium Danshensu also dose-dependently decreased protein and mRNA levels of nucleotide binding oligomerization NOD-like receptor pyrin domain containing 3 (NLRP3) and high mobility group box 1 (HMGB1), which play a crucial role in promoting ischemic stroke-induced cell injury. Moreover, sodium Danshensu dose-dependently upregulated Beclin 1 expression, downregulated P62 protein expression, and further increased LC3B-II/LC3B-I ratio through inducing autophagy in astrocytes. Additionally, we noticed that sodium Danshensu dose-dependently increased tuberous sclerosis complex-2 (TSC2) protein expression, while significantly reduced the levels of mammalian target of rapamycin (mTOR) in the presence of OGD/R insult. These findings suggest that sodium Danshensu protects against OGD/R-induced injury by modulating the NLRP3 inflammasome and TSC2/mTOR pathways.

Policosanol ameliorates renal inflammation and pyroptosis in hypercholesterolemic rabbits via modulation of HMGB1/PI3K/mTOR/NLRP3/Caspase-1 pathway

Hyperlipidemia represents a major risk factor for cardiovascular and renal diseases. This study aimed to demonstrate the possible ameliorating effects of policosanol (PC) on renal inflammation and pyroptosis. Eighteen male New Zealand rabbits were randomly divided into 3 groups (n = 6/group), one received normal chow diet for 12 weeks while the others received either 0.5 % w/w high cholesterol diet (HCD) or HCD and concurrently treated with PC (5 mg/kg body weight/day) for 12 weeks. Rabbits fed with HCD showed significant increases in body and kidney weights; renal dysfunction, dyslipidemia, oxidative stress status, elevated levels of oxidized low density lipoprotein (Ox-LDL) and renal High-mobility group box 1 (HMGB1). These increases were accompanied with up-regulation of phosphatidylinositol-3-kinase (PI3K)/mammalian target of rapamycin (mTOR) signaling in renal tissue and activation of NOD-like receptor, pyrin domain-containing-3 (NLRP3) inflammasome. Administration of PC concurrently with HCD significantly modulated the HCD-induced activation of Ox-LDL/HMGB1/PI3K/mTOR/NLRP3/Caspase-1 signaling pathway and subsequently attenuated the resulted renal injury.

MiR-30a-5p and miR-153-3p regulate LPS-induced neuroinflammatory response and neuronal apoptosis by targeting NeuroD1

Neurogenic differentiation 1 (NeuroD1) is an essential transcription factor for neuronal differentiation, maturation, and survival, and is associated with inflammation in lipopolysaccharide (LPS)-induced glial cells; however, the concrete mechanisms are still ambiguous. Therefore, we investigated whether NeuroD1-targeting miRNAs affect inflammation and neuronal apoptosis, as well as the underlying mechanism. First, we confirmed that miR-30a-5p and miR-153-3p, which target NeuroD1, reduced NeuroD1 expression in microglia and astrocytes. In LPS-induced microglia, miR-30a-5p and miR-153-3p suppressed pro-inflammatory cytokines, reactive oxygen species, the phosphorylation of c-Jun N-terminal kinase, extracellular-signal-regulated kinase (ERK), and p38, and the expression of cyclooxygenase and inducible nitric oxide synthase (iNOS) via the NF-κB pathway. Moreover, miR-30a-5p and miR-153-3p inhibited the expression of NOD-like receptor pyrin domain containing 3 (NLRP3) inflammasomes, NLRP3, cleaved caspase-1, and IL-1β, which are involved in the innate immune response. In LPS-induced astrocytes, miR-30a-5p and miR-153-3p reduced ERK phosphorylation and iNOS expression via the STAT-3 pathway. Notably, miR-30a-5p exerted greater anti-inflammatory effects than miR-153-3p. Together, these results indicate that miR-30a-5p and miR-153-3p inhibit MAPK/NF-κB pathway in microglia as well as ERK/STAT-3 pathway in astrocytes to reduce LPS-induced neuronal apoptosis. This study highlights the importance of NeuroD1 in microglia and astrocytes neuroinflammation and suggests that it can be regulated by miR-30a-5p and miR-153-3p.

Pioglitazone Protects Against Hypoxia-Induced Cardiomyocyte Apoptosis Through Inhibiting NLRP3/Caspase-1 Pathway in vivo and in vitro.

This study aims to explore the underlying mechanisms of how Pioglitazone (Pio) affects myocardial ischemia-reperfusion (I/R) injury. In this study, after pretreatment of Pio, the pathologic change of myocardial tissues was measured via hematoxylin and eosin staining. The release of lactate dehydrogenase (LDH), superoxide dismutase (SOD), and nitric oxide (NO) were measured. The cardiomyocyte apoptosis was detected via TUNEL assay and flow cytometry assay. The mitochondrial membrane potential (ΔΨm) was estimated using the JC-1 probe. The release of cytochrome c in mitochondria and the translocation of cytochrome c in the cytosol were measured using western blot. Additionally, apoptosis-associated molecules and NOD-like receptor pyrin domain containing-3 (NLRP3)/caspase-1 pathway-related molecules were measured using western blot, quantitative real-time-polymerase chain reaction, and immunofluorescence staining. Results showed that the pretreatment of Pio significantly decreased myocardial tissue damage. Pio pretreatment inhibited the release of creatine kinase and LDH but promoted NO release in serum and H9c2 cell supernatants. Moreover, the pretreatment of Pio notably alleviated cardiomyocyte apoptosis. Pio pretreatment also maintained the mitochondrial membrane potential and prevented cytochrome c release in H/R-induced cardiomyocytes. Additionally, we confirmed that Pio pretreatment inhibited cardiomyocyte apoptosis via repressing the NLRP3/caspase-1 pathway. In conclusion, our study demonstrated that Pio could inhibit myocardial I/R injury and cardiomyocyte apoptosis by inhibiting the activation of the NLRP3/caspase-1 signaling pathway.

Palmitic acid impairs INS-1 cells and alters the global gene expression profile

Chronic elevated free fatty acids (FFAs) impair pancreatic β cells, but the mechanisms remain elusive. In this study, palmitic acid (PA) impaired viability and glucose-stimulated insulin secretion of INS-1 cells. Microarray analysis showed that PA markedly altered the expression of 277 probe sets of genes with 232 upregulated and 45 downregulated (fold change ≥ 2.0 or ≤ -2.0; P < 0.05). Gene Ontology analysis displayed a series of the biological process of the differentially expressed genes, such as intrinsic apoptotic signaling pathway in response to endoplasmic reticulum (ER) stress and oxidative stress, inflammatory response, positive regulation of macroautophagy, regulation of insulin secretion, cell proliferation and cycle, fatty acid metabolic process, glucose metabolic process and so on. Kyoto Encyclopedia of Genes and Genomes analysis demonstrated molecular pathways with which the differentially expressed genes associated, including NOD-like receptor, NF-κB and PI3K-Akt signaling pathways, apoptosis, adipocytokine signaling pathway, ferroptosis, protein processing in ER, fatty acid biosynthesis and cell cycle. Moreover, PA promoted protein expression of CHOP, cleaved caspase-3, microtubule-associated proteins light chain 3 (LC3)-II, NOD-like receptor pyrin domain containing 3 (NLRP3), cleaved IL-1β and Lcn2, increased reactive oxygen species, apoptosis and the ratio of LC3-II/I, and reduced p62 protein expression, intracellular glutathione peroxidase and catalase levels, suggesting activation of ER stress, oxidative stress, autophagy and NLRP3 inflammasome. The results indicate the impaired role of PA and the global gene expression profile of INS-1 cells following PA intervention, providing new insights into the mechanisms involving the damage of pancreatic β cells by FFAs.

Targeting the PANoptosome with 3,4-Methylenedioxy-β-Nitrostyrene, Reduces PANoptosis and Protects the Kidney against Renal İschemia-Reperfusion Injury

Objectıves The objectives of this study were a) to investigate the effect of targeting the PANoptosome with 3,4-methylenedioxy-β-nitrostyrene (MNS) on PANoptosis in the Renal ischemia-reperfussion (RIR) model b) to investigate the kidney protective effect of MNS toward RIR injury. Methods Thirty-two rats were divided into four groups randomly. The groups were assigned as Control, Sham, DMSO (dimethyl sulfoxide) and MNS groups. The rats in the MNS group were intraperitoneally given 20 mg/kg of MNS 30 minutes before reperfusion. 2% DMSO solvent that dissolves MNS were given to the rats in DMSO group. Left nephrectomy was performed on the rats under anesthesia at the 6th hour after reperfusion. Glutathione peroxidase (GPx), malondialdehyde (MDA), catalase (CAT), superoxide dismutase (SOD) and 8-Okso-2′-deoksiguanozin (8-OHdG) levels were measured. Immunohistochemical analysis, electron microscopic and histological examinations were carried out in the tissues. Results Total tubular injury score was lower in the MNS group (p < 0.001). Caspase-3, Gasdermin D and MLK (Mixed Lineage Kinase Domain Like Pseudokinase) expressions were considerably decreased in the MNS group (p < 0.001). Apoptotic index (AI) was found to be low in the MNS group (p < 0.001). CAT and SOD levels were higher in the MNS Group (p = 0.006, p = 0.0004, respectively). GPx, MDA, and 8-OH-dG levels were similar (p > 0.05) in all groups. MNS considerably improved the tissue structure, based on the electron microscopic analysis. Conclusıons Our results suggested that MNS administrated before the reperfusion reduces pyroptosis, apoptosis and necroptosis. These findings suggest that MNS significantly protects the kidney against RIR injury by reducing PANoptosis as a result of specific inhibition of Nod-like receptor pyrin domain-containing 3 (NLRP 3), one of the PANoptosome proteins.

Microglial Responses and Pain Behaviors Are Exacerbated by Chronic Sleep Deprivation in Rats with Chronic Pain Via Neuroinflammatory Pathways

The inflammatory response of central nervous system (CNS) and microglial activation is important in the development of pain behaviors induced by sleep deprivation. We found that chronic sleep deprivation (CSD) aggravated pain behaviors in rats with chronic pain by upregulating expression of Toll-like receptor 4 (TLR4), NOD-like receptor pyrin domain containing 3 (NLRP3), and interleukin 1β (IL-1β), which promoted microglial activation in the brain. We also found that CSD increased numbers of Iba1+ and TLR4+ cells, as well as neuronal apoptosis. Inhibitors of TLR4 and NLRP3 (TAK-242 and MCC950, respectively) reduced expression levels of inflammatory factor proteins and M1-related factor mRNA, decreased microglial activation, and relieved the hyperalgesia caused by CSD. These results suggest that CSD aggravated pain behavior in rats with chronic pain through the TLR4/NLRP3/IL-1β signaling pathway, which mediates microglial activation and promotes CNS inflammation and neuronal apoptosis.

Does Pyroptosis Play a Role in Inflammasome-Related Disorders?

Inflammasomes are multiprotein complexes orchestrating intracellular recognition of endogenous and exogenous stimuli, cellular homeostasis, and cell death. Upon sensing of certain stimuli, inflammasomes typically activate inflammatory caspases that promote the production and release of the proinflammatory cytokines IL-1β, IL-1α, and IL-18 and induce a type of inflammatory cell death known as “pyroptosis”. Pyroptosis is an important form of regulated cell death executed by gasdermin proteins, which is largely different from apoptosis and necrosis. Recently, several signaling pathways driving pyroptotic cell death, including canonical and noncanonical inflammasome activation, as well as caspase-3-dependent pathways, have been reported. While much evidence exists that pyroptosis is involved in the development of several inflammatory diseases, its contribution to inflammasome-related disorders (IRDs) has not been fully clarified. This article reviews molecular mechanisms leading to pyroptosis, and attempts to provide evidence for its possible role in inflammasome-related disorders, including NLR pyrin domain containing 3 (NLRP3) inflammasome disease, NLR containing a caspase recruitment domain 4 (NLRC4) inflammasome disease, and pyrin inflammasome disease. Although the specific mechanism needs further investigations, these studies have uncovered the role of pyroptosis in inflammasome-related disorders and may open new avenues for future therapeutic interventions.

Glucocorticoid induces GSDMD-dependent pyrolysis in PC12 cells via endoplasmic reticulum stress.

The present study explored whether pyroptosis is involved in the injury process of PC12 cells induced by glucocorticoid (GC) and the regulatory relationship between endoplasmic reticulum stress (ERS) and pyrolysis. LDH leakage of PC12 cells was detected by LDH assay. The number of dead cells was detected by SYTOX green nucleic acid staining. The levels of IL-1β and IL-18 in the supernatants was detected by ELSIA assay. The expression levels of glucose regulated protein 78 (GRP78), cleaved gasdermin D-NT (cleaved-GSDMD-NT), NLR-pyrin domain-containing 3 (NLRP3) and cleaved-caspase-1 were observed by immunofluorescence staining and western blot. The LDH assay revealed that GC exposure significantly increased the release of LDH. The results of SYTOX green acid staining showed that GC exposure significantly increased the number of SYTOX green acid-positive cells. The ELSIA assay revealed that GC exposure significantly increased the levels of IL-1β and IL-18 in the supernatants. The results of immunofluorescence staining and western blot showed that GC exposure significantly increased the expression of GRP78, cleaved-GSDMD-NT, NLRP3 and cleaved caspase-1. Treatment with the ERS inhibitor tauroursodeoxycholate (TUDCA) and siRNA GSDMD attenuated related damage and downregulated the expression of the abovementioned proteins. The present study clearly demonstrated that GC exposure can induce GSDMD-dependent pyrolysis, and ERS is involved in the above damage process.

CREBH alleviates mitochondrial oxidative stress through SIRT3 mediating deacetylation of MnSOD and suppression of Nlrp3 inflammasome in NASH.

Lipotoxicity and unresolved oxidative stress are key drivers of metabolic inflammation in nonalcoholic steatohepatitis (NASH). cAMP-response element binding protein H(CREBH) is a liver-specific transcription factor and regulates the glucose and lipid metabolism of NASH. However, its role in mitochondrial oxidative stress and its association with sirtuin 3 (SIRT3), a master regulator of deacetylation for mitochondrial proteins, remains elusive. In this study, AML-12cells were treated with palmitic acid to imitate the pathological changes of NASH in vitro and 8-week-old male C57BL/6J mice were fed with a high-fat (HF) diet or a methionine-choline-deficient (MCD) diet to build the widely accepted in vivo model of NASH. We found that lipid overload induced mitochondrial oxidative stress and stimulated the expression of CREBH and SIRT3. CREBH overexpression alleviated the mitochondrial oxidative stress. Moreover, CREBH promoted SIRT3 expression, which regulated the deacetylation of manganese superoxide dismutase (MnSOD) and inhibited NOD-Like Receptor Pyrin Domain Containing 3 (Nlrp3) inflammasome activation whereas suppression of SIRT3 damaged the protecting ability of CREBH in mitochondrial oxidative stress. CREBH knockout mice were highly susceptible to HF and MCD diet-induced NASH with more severe oxidative stress. Collectively, our results firstly provided the support that CREBH could serve as a protective factor in the progression of NASH by regulating the acetylation of MnSOD and the activation of Nlrp3 inflammasome through SIRT3. These results suggest that CREBH might be a valuable therapeutic candidate for NASH.