[Ligusticum cycloprolactam inhibits IL-1ß-induced apoptosis and inflammation of rat chondrocytes via HMGB1/TLR4/NF-κB signaling pathway].

Chondrocytes are unique resident cells in the articular cartilage, and the pathological changes of them can lead to the occurrence of osteoarthritis(OA). Ligusticum cycloprolactam(LIGc) are derivatives of Z-ligustilide(LIG), a pharmacodynamic marker of Angelica sinensis, which has various biological functions such as anti-inflammation and inhibition of cell apoptosis. However, its protective effect on chondrocytes in the case of OA and the underlying mechanism remain unclear. This study conducted in vitro experiments to explore the molecular mechanism of LIGc in protecting chondrocytes from OA. The inflammation model of rat OA chondrocyte model was established by using interleukin-1ß(IL-1ß) to induce. LIGc alone and combined with glycyrrhizic acid(GA), a blocker of the high mobility group box-1 protein(HMGB1)/Toll-like receptor 4(TLR4)/nuclear factor-kappa B(NF-κB) signaling pathway, were used to intervene in the model, and the therapeutic effects were systematically evaluated. The viability of chondrocytes treated with different concentrations of LIGc was measured by the cell counting kit-8(CCK-8), and the optimal LIGc concentration was screened out. Annexin V-FITC/PI apoptosis detection kit was employed to examine the apoptosis of chondrocytes in each group. The enzyme-linked immunosorbent assay(ELISA) was employed to measure the expression of cyclooxygenase-2(COX-2), prostaglandin-2(PGE2), and tumor necrosis factor-alpha(TNF-α) in the supernatant of chondrocytes in each group. Western blot was employed to determine the protein levels of B-cell lymphoma-2(Bcl-2), Bcl-2-associated X protein(Bax), caspase-3, HMGB1, TLR4, and NF-κB p65. The mRNA levels of HMGB1, TLR4, NF-κB p65, and myeloid differentiation factor 88(MyD88) in chondrocytes were determined by real-time fluorescent quantitative PCR(RT-qPCR). The safe concentration range of LIGc on chondrocytes was determined by CCK-8, and then the optimal concentration of LIGc for exerting the effect was clarified. Under the intervention of IL-1ß, the rat chondrocyte model of OA was successfully established. The modeled chondrocytes showed increased apoptosis rate, promoted expression of COX-2, PGE2, and TNF-α, up-regulated protein levels of Bax, caspase-3, HMGB1, TLR4, and NF-κB p65 and mRNA levels of HMGB1, TLR4, NF-κB p65, and MyD88, and down-regulated protein level of Bcl-2. However, LIGc reversed the IL-1ß-induced changes of the above factors. Moreover, LIGc combined with GA showed more significant reversal effect than LIGc alone. These fin-dings indicate that LIGc extracted and derived from the traditional Chinese medicine A. sinensis can inhibit the inflammatory response of chondrocytes and reduce the apoptosis of chondrocytes, and this effect may be related to the HMGB1/TLR4/NF-κB signaling pathway. The pharmacological effect of LIGc on protecting chondrocytes has potential value in delaying the progression of OA and improving the clinical symptoms of patients, and deserves further study.

Differential cytokine expression in gastric tissues highlights helicobacter pylori's role in gastritis.

Helicobacter pylori (H. pylori), known for causing gastric inflammation, gastritis and gastric cancer, prompted our study to investigate the differential expression of cytokines in gastric tissues, which is crucial for understanding H. pylori infection and its potential progression to gastric cancer. Focusing on Il-1ß, IL-6, IL-8, IL-12, IL-18, and TNF-α, we analysed gene and protein levels to differentiate between H. pylori-infected and non-infected gastritis. We utilised real-time quantitative polymerase chain reaction (RT-qPCR) for gene quantification, immunohistochemical staining, and ELISA for protein measurement. Gastric samples from patients with gastritis were divided into three groups: (1) non-gastritis (N-group) group, (2) gastritis without H. pylori infection (G-group), and (3) gastritis with H. pylori infection (GH-group), each consisting of 8 samples. Our findings revealed a statistically significant variation in cytokine expression. Generally, cytokine levels were higher in gastritis, but in H. pylori-infected gastritis, IL-1ß, IL-6, and IL-8 levels were lower compared to H. pylori-independent gastritis, while IL-12, IL-18, and TNF-α levels were higher. This distinct cytokine expression pattern in H. pylori-infected gastritis underscores a unique inflammatory response, providing deeper insights into its pathogenesis.

Circ_0044235 regulates the development of osteoarthritis by the modulation of miR-375/PIK3R3 axis.

BACKGROUND: Circular RNAs (circRNAs) play an important role in osteoarthritis (OA). However, the role of circRNA in OA is still unclear. Here, we explored the role and mechanism of circ_0044235 in OA. METHODS: CHON-001 cells were treated with IL-1ß to establish OA model in vitro. The levels of circ_0044235, miR-375 and phosphoinositide 3-kinase (PI3K) regulatory subunit 3 (PIK3R3) were detected by quantitative real-time PCR. Cell count kit-8 assay and flow cytometry assay were used to detect cell viability and apoptosis. The concentrations of inflammation factors were determined by enzyme-linked immunosorbent assay. Western blot was used to detect protein levels. The interaction between miR-375 and circ_0044235 or PIK3R3 was analyzed by dual-luciferase reporter assay and RNA immunoprecipitation assay. RESULTS: Circ_0044235 was significantly decreased in OA cartilage tissue and IL-1ß-treated CHON-001 cells. Overexpression of circ_0044235 promoted IL-1ß-stimulated CHON-001 cell viability and inhibited apoptosis, inflammation, and extracellular matrix (ECM) degradation. In mechanism analysis, circ_0044235 could act as a sponge for miR-375 and positively regulate PIK3R3 expression. In addition, miR-375 ameliorated the effect of circ_0044235 overexpression on IL-1ß-mediated CHON-001 cells injury. In addition, miR-375 inhibition mitigated IL-1ß-induced CHON-001 cell injury, while PIK3R3 silencing restored the effect. CONCLUSION: Circ_0044235 knockdown alleviated IL-1ß-induced chondrocytes injury by regulating miR-375/PIK3R3 axis, confirming that circ_0044235 might be a potential target for OA treatment.

Optogenetically controlled inflammasome activation demonstrates two phases of cell swelling during pyroptosis.

Inflammasomes are multiprotein platforms that control caspase-1 activation, which process the inactive precursor forms of the inflammatory cytokines IL-1ß and IL-18, leading to an inflammatory type of programmed cell death called pyroptosis. Studying inflammasome-driven processes, such as pyroptosis-induced cell swelling, under controlled conditions remains challenging because the signals that activate pyroptosis also stimulate other signaling pathways. We designed an optogenetic approach using a photo-oligomerizable inflammasome core adapter protein, apoptosis-associated speck-like containing a caspase recruitment domain (ASC), to temporally and quantitatively manipulate inflammasome activation. We demonstrated that inducing the light-sensitive oligomerization of ASC was sufficient to recapitulate the classical features of inflammasomes within minutes. This system showed that there were two phases of cell swelling during pyroptosis. This approach offers avenues for biophysical investigations into the intricate nature of cellular volume control and plasma membrane rupture during cell death.

[Impact of chaperone-mediated autophagy on bilirubin-induced damage of mouse microglial cells]. / åˆ†å­ä¼´ä¾£ä»‹å¯¼çš„è‡ªå™¬å¯¹èƒ†çº¢ç´ è¯±å¯¼çš„å°é¼ å°èƒ¶è´¨ç»†èƒžæŸä¼¤çš„å½±å“.

OBJECTIVES: To investigate the effect of chaperone-mediated autophagy (CMA) on the damage of mouse microglial BV2 cells induce by unconjugated bilirubin (UCB). METHODS: The BV2 cell experiments were divided into two parts. (1) For the CMA activation experiment: control group (treated with an equal volume of dimethyl sulfoxide), QX77 group (treated with 20 µmol/L QX77 for 24 hours), UCB group (treated with 40 µmol/L UCB for 24 hours), and UCB+QX77 group (treated with both 20 µmol/L QX77 and 40 µmol/L UCB for 24 hours). (2) For the cell transfection experiment: LAMP2A silencing control group (treated with an equal volume of dimethyl sulfoxide), LAMP2A silencing control+UCB group (treated with 40 µmol/L UCB for 24 hours), LAMP2A silencing group (treated with an equal volume of dimethyl sulfoxide), and LAMP2A silencing+UCB group (treated with 40 µmol/L UCB for 24 hours). The cell viability was assessed using the modified MTT method. The expression levels of p65, nucleotide-binding oligomerization domain-like receptor protein 3 (NLRP3), and cysteinyl aspartate specific proteinase-1 (caspase-1) were detected by Western blot. The relative mRNA expression levels of the inflammatory cytokines interleukin (IL)-1ß, IL-6, and tumor necrosis factor-α (TNF-α) were determined by real-time quantitative polymerase chain reaction. Levels of IL-6 and TNF-α in the cell culture supernatant were measured using ELISA. The co-localization of heat shock cognate protein 70 with p65 and NLRP3 was detected by immunofluorescence. RESULTS: Compared to the UCB group, the cell viability in the UCB+QX77 group increased, and the expression levels of inflammation-related proteins p65, NLRP3, and caspase-1, as well as the mRNA relative expression levels of IL-1ß, IL-6, and TNF-α and levels of IL-6 and TNF-α decreased (P<0.05). Compared to the control group, there was co-localization of heat shock cognate protein 70 with p65 and NLRP3 in both the UCB and UCB+QX77 groups. After silencing the LAMP2A gene, compared to the LAMP2A silencing control+UCB group, the LAMP2A silencing+UCB group showed increased expression levels of inflammation-related proteins p65, NLRP3, and caspase-1, as well as increased mRNA relative expression levels of IL-1ß, IL-6, and TNF-α and levels of IL-6 and TNF-α (P<0.05). CONCLUSIONS: CMA is inhibited in UCB-induced BV2 cell damage, and activating CMA may reduce p65 and NLRP3 protein levels, suppress inflammatory responses, and counteract bilirubin neurotoxicity.

A novel fatty acid analogue triggers CD36-GPR120 interaction and exerts anti-inflammatory action in endotoxemia.

Inflammation is a mediator of a number of chronic pathologies. We synthesized the diethyl (9Z,12Z)-octadeca-9,12-dien-1-ylphosphonate, called NKS3, which decreased lipopolysaccharide (LPS)-induced mRNA upregulation of proinflammatory cytokines (IL-1ß, IL-6 and TNF-α) not only in primary intraperitoneal and lung alveolar macrophages, but also in freshly isolated mice lung slices. The in-silico studies suggested that NKS3, being CD36 agonist, will bind to GPR120. Co-immunoprecipitation and proximity ligation assays demonstrated that NKS3 induced protein-protein interaction of CD36 with GPR120in RAW 264.7 macrophage cell line. Furthermore, NKS3, via GPR120, decreased LPS-induced activation of TAB1/TAK1/JNK pathway and the LPS-induced mRNA expression of inflammatory markers in RAW 264.7 cells. In the acute lung injury model, NKS3 decreased lung fibrosis and inflammatory cytokines (IL-1ß, IL-6 and TNF-α) and nitric oxide (NO) production in broncho-alveolar lavage fluid. NKS3 exerted a protective effect on LPS-induced remodeling of kidney and liver, and reduced circulating IL-1ß, IL-6 and TNF-α concentrations. In a septic shock model, NKS3 gavage decreased significantly the LPS-induced mortality in mice. In the last, NKS3 decreased neuroinflammation in diet-induced obese mice. Altogether, these results suggest that NKS3 is a novel anti-inflammatory agent that could be used, in the future, for the treatment of inflammation-associated pathologies.

[3-Deazaadenosine (3-DAA) accelerates Japanese encephalitis virus replication and down-regulates levels of inflammatory factors in mouse and hamster cells].

Objective To investigate the effect of 3-deazaadenosine (3-DAA), an N6-methyladenosine (m6A) methylation modification inhibitor, on the replication of the Japanese encephalitis virus (JEV). Methods Neuro2a mouse neuroblastoma cells, N9 mouse microglial cells, and BHK baby hamster kidney cells were exposed to JEV and then treated with 3-DAA. JEV was also injected into the footpad of adult C57BL/6 mice, which were then administered 3-DAA intraperitoneally. Real-time quantitative PCR was utilized to measure mRNA expression levels of JEV, interleukin 1ß (IL-1ß), IL-6, tumor necrosis factor α (TNF-α), monocyte chemoattractant protein 1 (MCP-1), inducible nitric oxide synthase (iNOS), arginase 1 (Arg1), interferon (IFN)-α, IFN-ß, IFN-γ, and C-X-C motif chemokine ligand 10 (CXCL10) in the cells and mouse brain tissues. Western blot analysis was used to detect JEV protein expression in the cells and mouse brain tissues. Furthermore, the survival of the mice was monitored and pathological changes in mouse brains were observed via hematoxylin and eosin (HE) staining. Results 3-DAA had a dose-dependent effect on the replication of RNA and protein expression of JEV in both BHK, N9, Neuro 2α cells and mouse brain tissues, which resulted in rapid progression of JEV infection in mice and a decrease in their survival rate. Furthermore, 3-DAA suppressed the expression of inflammatory factors such as IL-6, TNF-α, CXCL10, IL-1ß and iNOS, thus weakening the immune response. Conclusion 3-DAA promotes JEV infection and hastens death of infected cells and mice, indicating that m6A modification may negatively regulate JEV replication.

Expression of TLR2, IL-1ß, and IL-10 Genes as a Possible Factor of Successful or Pathological Aging in Nonagenarians.

The expression of the gene of pattern recognition receptor TLR2, proinflammatory cytokine IL-1ß, and anti-inflammatory cytokine IL-10 was analyzed in the peripheral blood of nonagenarians (n=219; mean age 92.1 years, 77 men and 142 women) in comparison with healthy young donors (n=24; mean age 22.5 years, 16 women and 8 men). Nonagenarians were interviewed, medical records were analyzed, and a comprehensive geriatric assessment was performed according to the Clinical Guidelines on Frailty. The level of gene expression was determined by real-time PCR. The participation of inflammatory mechanisms in the immunosenescence was revealed. It was shown that increased expression of IL1B and TLR2 genes is associated with the development of frailty in nonagenarians and can be a factor of pathological aging. Increased expression of IL10 gene can be considered as a factor of successful aging in nonagenarians.

Interleukin-1ß promotes human metapneumovirus replication via activating the cGAS-STING pathway.

BACKGROUND: Human metapneumovirus(hMPV) is one of the most common viruses that cause acute lower respiratory tract infections. Interleukin-1ß (IL-1ß) has been reported to play an important role in multiple virus replication. Patients with hMPV infection have increased levels of IL-1ß which reminds IL-1ß is associated with hMPV infection. However, the mechanism by which IL-1ß affects hMPV replication remains unclear. In this study, we explore the effect of IL-1ß on hMPV replication and investigate its specific mechanism of action. METHODS: We established an hMPV infection model through Human bronchial epithelial cells (16HBE). qRT-PCR and Western Blot were used to detect the expression levels of IL-1ß, cyclic GMP-AMP synthase (cGAS), and interferon stimulating factor (STING). Regulating IL-1ß expression by small interfering RNA (siRNA) or exogenous supplementary to study the influence of hMPV replication. The selective cGAS inhibitor RU.521, G150, and STING inhibitor H-151 were utilized to detect hMPV replication in 16HBE cells. RESULTS: The level of IL-1ß protein increased in a time-dependent and dose-dependent manner after hMPV infection. The mRNA and protein levels of cGAS and STING were significantly up-regulated. Knockdown of IL-1ß could contribute to the decreased viral loads of hMPV. While the exogenous supplement of recombinant human IL-1ß in cells, replication of hMPV was significantly increased. Additionally, the level of cGAS-STING protein expression would be affected by regulating IL-1ß expression. Inhibitors of the cGAS-STING pathway led to a lower level of hMPV replication. CONCLUSION: This study found that IL-1ß could promote hMPV replication through the cGAS-STING pathway, which has the potential to serve as a candidate to fight against hMPV infection, targeting IL-1ß may be an effective new strategy to restrain virus replication.

Salmonella Enteritidis antitoxin DinJ inhibits NLRP3-dependent canonical inflammasome activation in macrophages.

The inflammasome is a pivotal component of the innate immune system, acting as a multiprotein complex that plays an essential role in detecting and responding to microbial infections. Salmonella Enteritidis have evolved multiple mechanisms to regulate inflammasome activation and evade host immune system clearance. Through screening S. Enteritidis C50336ΔfliC transposon mutant library, we found that the insertion mutant of dinJ increased inflammasome activation. In this study, we demonstrated the genetic connection between the antitoxin DinJ and the toxin YafQ in S. Enteritidis, confirming their co-transcription. The deletion mutant ΔfliCΔdinJ increased cell death and IL-1ß secretion in J774A.1 cells. Western blotting analysis further showed elevated cleaved Caspase-1 product (p10 subunits) and IL-1ß secretion in cells infected with ΔfliCΔdinJ compared to cells infected with ΔfliC. DinJ was found to inhibit canonical inflammasome activation using primary bone marrow-derived macrophages (BMDMs) from Casp-/- C57BL/6 mice. Furthermore, DinJ specifically inhibited NLRP3 inflammasome activation, as demonstrated in BMDMs from Nlrp3-/- and Nlrc4-/- mice. Fluorescence resonance energy transfer (FRET) experiments confirmed the translocation of DinJ into host cells during infection. Finally, we revealed that DinJ could inhibit the secretion of IL-1ß and IL-18 in vivo, contributing to S. Enteritidis evading host immune clearance. In summary, our findings provide insights into the role of DinJ in modulating the inflammasome response during S. Enteritidis infection, highlighting its impact on inhibiting inflammasome activation and immune evasion.

Different Gene Expression Patterns of IL-1 Family Members in Parkinson's Disease: Results from Bayesian Regression Model.

Parkinson's disease, the second most prevalent neurodegenerative disorder lacking a recognized etiology, is influenced by oxidative stress and alterations in inflammatory cytokine levels. This study aimed to investigate the expression levels of Interleukin(IL)1 receptor accessory protein (IL-1RAcP), IL1ß, IL1α, IL33, and IL36 genes in blood cells and serum IL-1ß levels in Parkinson's disease patients compared to healthy controls (HCs).I n this case-control study, 44 Parkinson's disease patients and 44 age- and sex-matched HCs were included. Gene expression levels were assessed using Quantitative Real-time PCR, and serum IL-1ß levels were measured via enzyme-linked immunosorbent assay. Advanced statistical analyses using the Bayesian regression model in R software were employed. Parkinson's disease patients exhibited elevated expression levels of IL-1RAcP and IL1ß genes  but decreased levels of IL1α, IL33, and IL36 compared to HCs. Age-based differences were not significant. Regarding gender, IL33 transcript levels were significantly higher in males, and serum IL-1ß levels were increased in patients. Subgroup analysis by gender indicated alterations in IL1ß and IL-1RAcP expression in both genders, while IL1α, IL33, and IL36 showed reduced expression only in males. Remarkably, only female patients displayed significantly higher serum IL-1ß levels than female HCs. These findings suggest that dysregulation of immune-related factors plays a crucial role in Parkinson's disease.

Investigating the association between the tissue expression of miRNA-101, JAK2/STAT3 with TNF-α, IL-6, IL-1ß, and IL-10 cytokines in the ulcerative colitis patients.

BACKGROUND: Ulcerative colitis (UC) is a chronic inflammatory bowel disease caused by numerous factors, such as immune system dysfunction and genetic factors. MicroRNAs (miRNAs) play a crucial role in UC pathogenesis, particularly via the JAK-STAT pathway. Our aim was to investigate the association between miRNA-101 and JAK2-STAT3 signaling pathway with inflammatory cytokines in UC patients. METHODS: We enrolled 35 UC patients and 35 healthy individuals as the control group, referred to Shariati Hospital, Tehran, Iran. Patients were diagnosed based on clinical, laboratory, histological, and colonoscopy criteria. RNA and protein extracted from tissue samples. Real-time PCR was used to assess the expression levels of miRNA-101, interleukin (IL)-1ß, IL-6, tumor necrosis factor (TNF)-α, and IL-10 genes, while western blot was employed to measure levels of P-STAT3, total STAT3, and JAK2 proteins. RESULTS: Expression of pro-inflammatory cytokines TNF-α, IL-1ß, and IL-6 significantly increased, while the expression of IL-10 significantly decreased in the case group versus controls. Additionally, miRNA-101 expression was significantly higher in UC patients. A significant correlation between miRNA-101 and IL-6 expression was observed, indicating their relationship and possible impact on cell signaling pathways, JAK2-STAT3. No significant changes were observed in phosphorylated and total STAT3 and JAK2 protein expression. CONCLUSION: This study provides evidence of increased miRNA-101 expression in UC tissue, suggesting a potential correlation between miRNA-101 and IL-6 expression and their involvement in the JAK2-STAT3 pathway. The study confirms alterations in UC patients' pro-inflammatory cytokines and anti-inflammatory IL-10. However, further investigations are needed to understand the exact role of miRNA-101 in UC pathogenesis fully.

Single-cell profiling identifies IL1Bhi macrophages associated with inflammation in PD-1 inhibitor-induced inflammatory arthritis.

Inflammatory arthritis (IA) is a common rheumatic adverse event following immune checkpoint inhibitors treatment. The clinical disparities between IA and rheumatoid arthritis (RA) imply disease heterogeneity and distinct mechanisms, which remain elusive. Here, we profile CD45+ cells from the peripheral blood or synovial fluid (SF) of patients with PD-1-induced IA (PD-1-IA) or RA using single-cell RNA sequencing. We report the predominant expansion of IL1Bhi myeloid cells with enhanced NLRP3 inflammasome activity, in both the SF and peripheral blood of PD-1-IA, but not RA. IL1Bhi macrophages in the SF of PD-1-IA shared similar inflammatory signatures and might originate from peripheral IL1Bhi monocytes. Exhausted CD8+ T cells (Texs) significantly accumulated in the SF of patients with PD-1-IA. IL1Bhi myeloid cells communicated with CD8+ Texs possibly via the CCR1-CCL5/CCL3 and CXCL10-CXCR3 axes. Collectively, these results demonstrate different cellular and molecular pathways in PD-1-IA and RA and highlight IL1Bhi macrophages as a possible therapeutic target in PD-1-IA.

High Mobility Group Box 1 Gene Polymorphism and Serum High Mobility Group Box 1, Interleukin 1 Beta, and Alpha-Klotho Crosstalk in Severe COVID-19 Patients.

AIM: To evaluate the serum levels of HMGB1, IL1ß, and α-klotho in COVID-19 patients with different disease severity, investigate their association with clinicopathological parameters, and to assess HMGB1 rs1045411 polymorphism and its relation with clinical severity. METHODS: 120 COVID-19 patients (89 critically ill, 15 severe, and 16 moderately severe) along with 80 healthy control were enrolled.The serum levels of HMGB1,IL1ß, and α-klotho were determined by ELISA. The HMGB1 rs1045411 polymorphism was detected by RT- PCR. RESULTS: The serum levels of HMGB1, IL1ß, and α-klotho were significantly higher in critically ill COVID-19 patients compared to other groups. The HMGB1rs1045411 polymorphism revealed a significant decrease in the percentage of T/T genotypes in COVID-19 patients compared to controls. The (ROC) analysis showed moderate diagnostic potential for serum HMGB1, IL1ß, and α-klotho. CONCLUSION: The serum HMGB1, IL1ß, and α-klotho may be severity markers and promising therapeutic targets for COVID-19 patients.

Effect of electroacupuncture on neuronal programmed necrosis by regulating RIP1/RIP3/MLKL pathway in rats with cerebral ischemia reperfusion injury. / 电针调节RIP1/RIP3/MLKLé€šè·¯å¯¹è„‘ç¼ºè¡€å†çŒæ³¨æŸä¼¤å¤§é¼ ç¥žç»å ƒç¨‹åºæ€§åæ­»çš„å½±å“.

OBJECTIVES: To investigate the neuroprotective effect of electroacupuncture (EA) at "Quchi"(LI11) and "Zusanli"(ST36) in the rats with cerebral ischemia reperfusion injury and its influence on programmed necrosis of cerebral cortical neurons. METHODS: Sixty male SD rats were randomly divided into sham-operation group, model group, EA group and inhibitor group, with 15 rats in each group. Left middle cerebral artery occlusion model was established using the modified thread embolism method. In the sham-operation group, the carotid artery was exposed and dissociated in each rat. EA was applied to "Quchi"(LI11) and "Zusanli"(ST36) on the right side for 30 min each time, once daily for 7 days in the rats of the EA group. The rats in the inhibitor group were intraperitoneally injected with norstatin-1 (0.6 mg/kg) for consecutive 7 days. The neurological deficit score of rats in each group was observed. HE staining was adopted to detect the degree of pathological damage of the cerebral cortex in the infarction area. Using TUNEL staining, the apoptosis of cortical neurons in the infarction area was determined；the contents of tumor necrosis factor α (TNF-α), interleukin (IL)-1ß and IL-6 were detected by ELISA；the mRNA and protein expression of the receptor interacting protein-1 (RIP1), the receptor interacting protein-3 (RIP3) and the substrate mixed lineage kinase like protein (MLKL) were detected by fluorescence quantitative PCR and Western blot, respectively. RESULTS: In comparison with the sham-operation group, the neurological deficit score in the model group was higher(P<0.01)；HE staining showed that there was the pathological damage in the infarction area；the neuron apoptosis rate, the contents of TNF-α, IL-1ß and IL-6, and the mRNA and protein expressions of RIP1, RIP3 and MLKL increased(P<0.01) in the model group. In the EA group, the neurological deficit score was reduced(P<0.01)；HE staining showed that the pathological damage was ameliorated in the infarction area；the neuron apoptosis rate, the contents of TNF-α, IL-1ß and IL-6, and the mRNA and protein expressions of RIP1, RIP3, MLKL decreased(P<0.05, P<0.01) when compared with those in the model group. CONCLUSIONS: EA can attenuate cerebral ischemia reperfusion injury and display its neuroprotective effect probably through inhibiting programmed necrosis of cerebral cortical neurons in the rats.

HDAC11 Regulates Palmitate-induced NLRP3 Inflammasome Activation by Inducing YAP Expression in THP-1 Cells and PBMCs.

Histone deacetylase 11 (HDAC11) has been implicated in the pathogenesis of metabolic diseases characterized by chronic low-grade inflammation, such as obesity. However, the influence of HDAC11 on inflammation and the specific effect of HDAC11 on the palmitic acid (PA)-induced NLR family pyrin domain containing 3 (NLRP3) inflammasome activation are poorly understood. The effect of PA treatment on HDAC11 activity and the NLRP3 inflammasome was investigated in human peripheral blood mononuclear cells and THP-1 cells. The PA-induced responses of key markers of NLRP3 inflammasome activation, including NLRP3 gene expression, caspase-1 p10 activation, cleaved IL-1ß production, and extracellular IL-1ß release, were assessed as well. The role of HDAC11 was explored using a specific inhibitor of HDAC11 and by knockdown using small interfering (si)HDAC11 RNA. The relationship between HDAC11 and yes-associated protein (YAP) in the PA-induced NLRP3 inflammasome was investigated in THP-1 cells with HDAC11 or YAP knockdown. Following PA treatment, HDAC11 activity and protein levels increased significantly, concomitant with activation of the NLRP3 inflammasome. Notably, PA-induced the upregulation of NLRP3, caspase-1 p10 activation, the production of cleaved IL-1ß, and the release of IL-1ß into the extracellular space, all of which were attenuated by FT895 treatment and by HDAC11 knockdown. In THP-1 cells, PA induced the expression of YAP and its interaction with NLRP3, resulting in NLRP3 inflammasome activation, whereas both were inhibited by FT895 and siHDAC11 RNA. These findings demonstrate a pivotal role for HDAC11 in the PA-induced activation of the NLRP3 inflammasome. HDAC11 inhibition thus represents a promising therapeutic strategy for mitigating NLRP3 inflammasome-related inflammation in the context of obesity.

NLRP3 Contributes to Sarcopenia Associated to Dependency Recapitulating Inflammatory-Associated Muscle Degeneration.

Sarcopenia, a complex and debilitating condition characterized by progressive deterioration of skeletal muscle, is the primary cause of age-associated disability and significantly impacts healthspan in elderly patients. Despite its prevalence among the aging population, the underlying molecular mechanisms are still under investigation. The NLRP3 inflammasome is crucial in the innate immune response and has a significant impact on diseases related to inflammation and aging. Here, we investigated the expression of the NLRP3 inflammasome pathway and pro-inflammatory cytokines in skeletal muscle and peripheral blood of dependent and independent patients who underwent hip surgery. Patients were categorized into independent and dependent individuals based on their Barthel Index. The expression of NLRP3 inflammasome components was significantly upregulated in sarcopenic muscle from dependent patients, accompanied by higher levels of Caspase-1, IL-1ß and IL-6. Among older dependent individuals with sarcopenia, there was a significant increase in the MYH3/MYH2 ratio, indicating a transcriptional shift in expression from mature to developmental myosin isoforms. Creatine kinase levels and senescence markers were also higher in dependent patients, altogether resembling dystrophic diseases and indicating muscle degeneration. In summary, we present evidence for the involvement of the NLRP3/ASC/NEK7/Caspase-1 inflammasome pathway with activation of pro-inflammatory SASP in the outcome of sarcopenia in the elderly.

The involvement of cytokine gene polymorphism in determining the vulnerability to Blastocystis and Helicobacter pylori co-infection in the Egyptian population.

AIMS: The present study aimed to identify any potential association between IL-1ß and TNF-α gene polymorphism and the risk of Blastocystis infection as well as co-infection of Blastocystis with Helicobacter pylori (H.pylori). METHODOLOGY: A total of 314 stool samples were collected and examined microscopically for the detection of parasitic infection. DNA was extracted from all samples and utilized to identify Blastocystis molecularly. Positive samples were used for H. pylori detection by rapid tests and PCR. Moreover, we investigate polymorphism in the TNF-α gene at position -1031T/C, -308 G/A, and IL-1ß at position +3954C/T using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assay. RESULTS: Out of the 314 stool samples, Blastocystis was detected in 93 (29.6 %); among them, 54 (58.1 %) had a mixed infection of Blastocystis with H. pylori. The TT genotype of the IL-1ß gene at position +3954 was significantly higher in Blasocystis-infected patients than in uninfected patients (17.2% vs. 6.3 %, P = 0.02), which might be considered a risk factor (OR = 3.2; CI =1.21-8.52). The TNF-α at position -1031 TT genotype was significantly higher in Blastocystis-infected patients than uninfected patients (44.1% vs. 10.8 %, P< 0.0001). The T allele (OR= 2.67; CI=1.51-4.72, P = 0.0008) might be considered a risk factor. The TNF- α at position -308 AA genotype is higher in Blasocystis infected than uninfected (17.2% vs 7.2 %, P = 0.03). TNF-α -308 AA (OR = 2.72; CI = 1.08-6.89) and A allele (OR= 1.46; CI= 0.797-2.66) might be considered risk factors. The TNF- α at position -308 G/A showed that the GG is the most frequent genotype in Blastocystis with H. pylori-positive patients with a significant association (P = 0.004), as well as the G allele (P = 0.02). The G allele (OR=1.924; CI= 1.071-3.454) might be considered a risk factor for co-infection of Blastocystis and H. pylori. CONCLUSION: SNPs (-1031 T/C and -308 G/A) of the TNF-α and (+3954 C/T) of the IL-1ß may be a useful marker in the assessment of the risk of Blastocystis infection, and TNF-α at position -308 G/A) may be a predictor for co-infection of Blastocystis with H. pylori.

A pivotal role for the IL-1ß and the inflammasome in preterm labor.

During labor, monocytes infiltrate massively the myometrium and differentiate into macrophages secreting high levels of reactive oxygen species and of pro-inflammatory cytokines (i.e. IL-1ß), leading to myometrial contraction. Although IL-1ß is clearly implicated in labor, its function and that of the inflammasome complex that cleaves the cytokine in its active form, has never been studied on steps preceding contraction. In this work, we used our model of lipopolysaccharide-induced preterm labor to highlight their role. We demonstrated that IL-1ß was secreted by the human myometrium during labor or in presence of infection and was essential for myometrial efficient contractions as its blockage with an IL-1 receptor antagonist (Anakinra) or a neutralizing antibody completely inhibited the induced contractions. We evaluated the implication of the inflammasome on myometrial contractions and differentiation stages of labor onset. We showed that the effects of macrophage-released IL-1ß in myometrial cell transactivation were blocked by inhibition of the inflammasome, suggesting that the inflammasome by producing IL-1ß was essential in macrophage/myocyte crosstalk during labor. These findings provide novel innovative approaches in the management of preterm labor, specifically the use of an inflammasome inhibitor to block the precursor stages of labor before the acquisition of the contractile phenotype.

M1 polarization of macrophages promotes stress-induced hair loss via interleukin-18 and interleukin-1ß.

Stress-induced hair loss is a prevalent health concern, with mechanisms that remain unclear, and effective treatment options are not yet available. In this study, we investigated whether stress-induced hair loss was related to an imbalanced immune microenvironment. Screening the skin-infiltrated immune cells in a stressed mouse model, we discovered a significant increase in macrophages upon stress induction. Clearance of macrophages rescues mice from stress-induced hair shedding and depletion of hair follicle stem cells (HFSCs) in the skin, demonstrating the role of macrophages in triggering hair loss in response to stress. Further flow cytometry analysis revealed a significant increase in M1 phenotype macrophages in mice under stressed conditions. In searching for humoral factors mediating stress-induced macrophage polarization, we found that the hormone Norepinephrine (NE) was elevated in the blood of stressed mice. In addition, in-vivo and in-vitro studies confirm that NE can induce macrophage polarization toward M1 through the ß-adrenergic receptor, Adrb2. Transcriptome, enzyme-linked immunosorbent assay (ELISA), and western blot analyses reveal that the NLRP3/caspase-1 inflammasome signaling and its downstream effector interleukin 18 (IL-18) and interleukin 1 beta (IL-1ß) were significantly upregulated in the NE-treated macrophages. However, inhibition of the NE receptor Adrb2 with ICI118551 reversed the upregulation of NLRP3/caspase-1, IL-18, and IL-1ß. Indeed, IL-18 and IL-1ß treatments lead to apoptosis of HFSCs. More importantly, blocking IL-18 and IL-1ß signals reversed HFSCs depletion in skin organoid models and attenuated stress-induced hair shedding in mice. Taken together, this study demonstrates the role of the neural (stress)-endocrine (NE)-immune (M1 macrophages) axis in stress-induced hair shedding and suggestes that IL-18 or IL-1ß may be promising therapeutic targets.