Release Of IL-1beta Research Articles

Abstract 372: MicroRNA-9 Inhibits Hyperglycemia-induced Cardiac Pyroptosis by Targeting ELAVL1

Diabetic cardiomyopathy is a common complication in patients with diabetes and is associated with underlying chronic inflammation and cardiac cell death, subsequently leading to left ventricular dysfunction and heart failure. ELAV-like protein 1 (ELAVL1, mRNA stabilizing protein) and NLRP3 activation (inflammasome complex protein)-mediated IL-1beta synthesis play a critical role in the progression of heart failure. However, ELAVL1 regulation of pyroptosis (caspase-1-mediated programmed apoptosis) and associated IL-1beta release in cardiomyocytes, especially under diabetic condition, remains elusive. Human diabetic, non-diabetic heart tissues, human ventricular cardiomyocytes and rat cardiomyoblasts exposed to high glucose (HG) were used for our studies. Our data demonstrates that human ventricular cardiomyocytes exposed to high glucose condition showed significant increase in ELAVL1 and NLRP3 expression with a concomitant increase in caspase-1 and IL-1 beta expression. Furthermore, human cardiac tissue from diabetic patients showed increased ELAVL1, caspase-1 and NLRP3 expression as compared to non-diabetic hearts. Intriguingly, ELAVL1 knockdown abrogates TNF-α induced canonical pyroptosis via NLRP3, caspase-1 and IL-1beta suppression. Interestingly, miRNA-9 expression significantly reduces in high glucose treated cardiomyocytes and in human diabetic hearts. Bioinformatics analysis and target validation assays showed that miR-9 directly targets ELAVL1. Inhibition of miR-9 up regulates ELAVL1 expression and activates caspase-1. Alternatively, miR-9 mimics attenuate hyperglycemia-induced ELAVL1 and inhibit cardiomyocyte pyroptosis. To our knowledge, this is the first report to demonstrate the role of miR-9/ELAVL1 in hyperglycemia-induced cardiac pyroptosis. Taken together our study highlights the potential therapeutic implications in preventing cardiomyocyte cell loss in human diabetic failing heart.

Evaluations of thyme extract effects in human normal bronchial and tracheal epithelial cell lines and in human lung cancer cell line

Thyme (Thymus vulgaris) is used traditionally to prepare herbal remedies possessing expectorant, mucolytic, antitussive and antispasmodic properties.The aim of the present study was to investigate the effects of a standardized hydroalcoholic extract of thyme on primary human airway (bronchial/tracheal) epithelial cell lines in a model of lung inflammation induced by LPS. In addition, the effects of thyme extract on human lung cancer cell line (H460) were analysed. Thyme extract showed significant anti-inflammatory properties by reducing the NF-κB p65 and NF-κB p52 transcription factors protein levels followed by the decrease of pro-inflammatory cytokines (IL-1 beta and IL-8), and Muc5ac secretion in human normal bronchial and tracheal epithelial cells. Moreover, the extract showed cytotoxic effects on H460 cancer cells, modulated the release of IL-1 beta, IL-8 and down-regulated NF-κB p65 and NF-κB p52 proteins.Taken together, these results substantiated the traditional uses of thyme in the treatment of respiratory diseases. Thyme extract might be an effective treatment of chronic diseases based on inflammatory processes when hypersecretion of mucus overwhelms the ciliary clearance and obstructs airways, causing morbidity and mortality. Moreover thyme extract, evaluated in H460 lung cancer cell line, demonstrated to induce cell cytotoxicity in addition to reduce inflammatory cell signals.

The major contribution of IL-1 alpha and caspase-11 in oviduct pathology during Chlamydia infection in the mouse model

Abstract The mouse model of genital Chlamydia trachomatis infection simulates the human chlamydial disease resulting in oviduct damage leading to hydrosalpinx. Using this model, we have shown that signaling by IL-1R is a major contributor of oviduct disease during infection. In the current study, we have investigated the specific contribution of IL-1 alpha and IL-1 beta that signal thorough IL-1R. Further, the contribution of caspase11-mediated cell-death that results in the release of IL-1 alpha and IL-1 beta were investigated. IL-1 alpha KO mice developed significantly less incidence (5%) of hydrosalpinx compared to WT mice (75%) (p=<0.001) upon infection. IL-1 beta KO mice had a reduced incidence in hydrosalpinx than WT mice, but significantly higher than IL-1 alpha KO mice. To determine the mechanism of IL-1 alpha release, infection course and pathology in caspase-1-11 KO mice and caspase-11 KO mice were determined. Both caspase-11 and caspase-1-11 KO mice displayed a reduced incidence of hydrosalpinx. Since the caspase-1 adaptor, ASC KO mice do not show a reduced incidence in pathology, our data suggest that caspase-11 activation is sufficient to drive the pathology. Oviduct pathology is a result of excessive PMN infiltration. Flow cytometric analyses demonstrate significantly reduced PMN infiltration to the genital tracts in IL-1 alpha and caspase-11 KO mice. However, T cell infiltration to the genital tracts is unchanged in both the groups, which reflected in normal infection clearance in IL-1 alpha and caspase-11 KO mice. These data suggest that activation of cell death pathways and local release of IL-1 alpha causes long-term damage to the oviducts during Chlamydia infection. [Supported by PHS funding NIAID AI067678 (UN)]

Abstract A20: Mammary tumor-derived CCL2 enhances pro-metastatic systemic inflammation through upregulation of macrophage-derived IL1beta

Abstract Metastatic breast cancer remains a major cause of cancer-related death among women. Metastasis is regulated by extensive crosstalk between cancer cells and immune cells. Besides the onset of a local inflammatory microenvironment, tumors frequently induce a systemic inflammatory state characterized by the release of various cytokines, chemokines and growth factors to mobilize myeloid cells that support metastasis, emphasizing the notion that cancer should be regarded as a systemic disease. Utilizing the K14cre;Cdh1F/F;Trp53F/F (KEP) conditional mouse model of metastatic breast cancer, we have recently unraveled a novel mechanistic link between mammary tumor-induced IL17-producing gamma delta T cells, systemic expansion of immunosuppressive neutrophils and metastasis formation. We identified tumor-derived IL1beta as one of the drivers of this cascade by inducing IL17 release from gamma delta T cells. However, it remains unclear how other inflammatory mediators are involved in this systemic inflammatory cascade. In the current study, we identified the pro-inflammatory cytokine CCL2 as a key regulator of the gamma delta T cell IL17 neutrophil axis. CCL2 blockade resulted in reduced IL17-production by gamma delta T cells, decreased systemic neutrophil accumulation and enhanced CD8+ T cell activity. We show that activation of this cascade relies on CCL2-mediated release of IL1beta from CCR2+ tumor-associated macrophages. These results indicate that CCL2, via IL1beta, acts as a key player in regulating the pro-metastatic gamma delta T cell IL17 neutrophil systemic inflammatory cascade and might be an interesting candidate for therapeutic targeting. Citation Format: Kelly Kersten, Seth B. Coffelt, Niels J. Verstegen, Kim Vrijland, Metamia Ciampricotti, Chris W. Doornebal, Cheei-Sing Hau, Parul Doshi, Karin E. de Visser. Mammary tumor-derived CCL2 enhances pro-metastatic systemic inflammation through upregulation of macrophage-derived IL1beta. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Metastasis; 2015 Nov 30-Dec 3; Austin, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(7 Suppl):Abstract nr A20.

Actin dynamics shape microglia effector functions.

Impaired actin filament dynamics have been associated with cellular senescence. Microglia, the resident immune cells of the brain, are emerging as a central pathophysiological player in neurodegeneration. Microglia activation, which ranges on a continuum between classical and alternative, may be of critical importance to brain disease. Using genetic and pharmacological manipulations, we studied the effects of alterations in actin dynamics on microglia effector functions. Disruption of actin dynamics did not affect transcription of genes involved in the LPS-triggered classical inflammatory response. By contrast, in consequence of impaired nuclear translocation of phospho-STAT6, genes involved in IL-4 induced alternative activation were strongly downregulated. Functionally, impaired actin dynamics resulted in reduced NO secretion and reduced release of TNFalpha and IL-6 from LPS-stimulated microglia and of IGF-1 from IL-4 stimulated microglia. However, pathological stabilization of the actin cytoskeleton increased LPS-induced release of IL-1beta and IL-18, which belong to an unconventional secretory pathway. Reduced NO release was associated with decreased cytoplasmic iNOS protein expression and decreased intracellular arginine uptake. Furthermore, disruption of actin dynamics resulted in reduced microglia migration, proliferation and phagocytosis. Finally, baseline and ATP-induced [Ca(2+)]int levels were significantly increased in microglia lacking gelsolin, a key actin-severing protein. Together, the dynamic state of the actin cytoskeleton profoundly and distinctly affects microglia behaviours. Disruption of actin dynamics attenuates M2 polarization by inhibiting transcription of alternative activation genes. In classical activation, the role of actin remodelling is complex, does not relate to gene transcription and shows a major divergence between cytokines following conventional and unconventional secretion.

Abstract O.21: Inositol 1,4,5 triphosphate 3- kinase C regulates NLRP3 Inflammasome activation in Kawasaki disease

Background: ITPKC controls calcium homeostasis and was identified in genome-wide studies to be associated with susceptibility and severity of KD. NLRP3 is activated by danger signals, leading to inflammasome activation and release of IL-1beta and IL-18. Ca 2+ mobilization mediates NLRP3 activation. We studied the contribution of ITPKC and NLRP3 inflammasome activation in KD. Methods: Cytokine levels were measured using ELISA. Gene expression was assayed using Illumina HumanHT-12v4. EBV-transformed B-cell lines from KD patients with different ITPKC genotypes were used to study the gene and protein expression and Ca 2+ flux using qRT-PCR, Western blot analysis, and FACS respectively. In vivo and in vitro analyses from ITPKC-deficient and wildtype mice were performed using ELISA, spinning disc confocal and Westerns, to determine cytokine release, Ca 2+ flux and protein expression responses respectively. Results: Children with KD show increased circulating IL-1beta, IL-18, IL-1RA and IL18BP protein during acute phase of KD (KD n=48, febrile controls n=41 p<0.001). NLRP3 and its associated inflammasome complex, and the IL-1[[Unsupported Character - Symbol Font ]] and IL18 receptor genes were up-regulated nearly two fold during the acute KD compared to matched convalescent controls (n=171, p<0.001). Pathway analysis showed specific up-regulation of NLRP3 related genes. The KD-associated genetic polymorphism in ITPKC (rs28493229) is functional, directly modulating [Ca 2+ ] i mobilization resulting in NLRP3 activation and increased production of IL-1beta and IL-18, as demonstrated by gene expression, circulating protein levels and functional assays of PBMCs and immortalized cell lines from affected children. These biologic data are partnered with evidence showing resistance to IVIG therapy in those with the ITPKC polymorphism associated with the highest basal and stimulated [Ca 2+ ] i levels. Studies using ITPKC-deficient mice in an animal model of KD support the molecular, cellular and clinical findings in affected children. Significance: ITPKC regulates NLRP3 activation via control of [Ca 2+ ] i mobilization, directing production of IL-1beta and IL-18, pointing to the key role of calcium mobilization in the immunopathogenesis of KD.

Flavonoids casticin and chrysosplenol D from Artemisia annua L. inhibit inflammation in vitro and in vivo

BackgroundThe aim of our experiments was to investigate the anti-inflammatory properties of casticin and chrysosplenol D, two flavonoids present in Artemisia annua L. MethodsTopical inflammation was induced in ICR mice using croton oil. Mice were then treated with casticin or chrysosplenol D. Cutaneous histological changes and edema were assessed. ICR mice were intragastrically administrated with casticin or chrysosplenol D followed by intraperitoneal injection of lipopolysaccharide (LPS). Mouse Raw264.7 macrophage cells were incubated with casticin or chrysosplenol D. Intracellular phosphorylation was detected, and migration was assessed by trans-well assay. HT-29/NFκB-luc cells were incubated with casticin or chrysosplenol D in the presence or absence of LPS, and NF-κB activation was quantified. ResultsIn mice, administration of casticin (0.5, 1 and 1.5μmol/cm2) and chrysosplenol D (1 and 1.5μmol/cm2) inhibited croton oil-induced ear edema (casticin: 29.39–64.95%; chrysosplenol D: 37.76–65.89%, all P<0.05) in a manner similar to indomethacin (0.5, 1 and 1.5μmol/cm2; 55.63–84.58%). Casticin (0.07, 0.13 and 0.27mmol/kg) and chrysosplenol D (0.07, 0.14 and 0.28mmol/kg) protected against LPS-induced systemic inflammatory response syndrome (SIRS) in mice (all P<0.05), in a manner similar to dexamethasone (0.03mmol/kg). Casticin and chrysosplenol D suppressed LPS-induced release of IL-1 beta, IL-6 and MCP-1, inhibited cell migration, and reduced LPS-induced IκB and c-JUN phosphorylation in Raw264.7 cells. JNK inhibitor SP600125 blocked the inhibitory effect of chrysosplenol D on cytokine release. ConclusionsThe flavonoids casticin and chrysosplenol D from A. annua L. inhibited inflammation in vitro and in vivo.

Anti-inflammatory mechanism research of flavonoid compounds in Dalbergiae Odoriferae Lignum by module-based network analysis

Dalbergiae Odoriferae Lignum as a traditional Chinese medicine (TCM) has been widely used for promoting blood circulation and removing blood stasis. Flavonoid compounds are main chemical constituents of Dalbergiae Odoriferae Lignum, which exert anti-inflammatory property. However, the underlying anti-inflammatory mechanisms of flavonoid compounds are incompletely understood. It has been reported that isoliquiritigenin, liquiritigenin, naringenin and butein possess anti-inflammatory property. The purpose of this study is to illuminate the anti-inflammatory mechanism of flavonoid compounds based on the protein interaction network (PIN) analysis on molecular network level. 130 targets of the main medicinal ingredients of flavonoid compounds were gained though database retrieval. A protein interaction network of flavonoid compounds was constructed with 589 nodes and 216 interactions. By a graph theoretic clustering algorithm Molecular Complex Detection (MCODE), 26 modules were identified and analyzed by Gene ontology (GO) enrichment. Two modules were associated with anti-inflammatory actions. The most interesting finding of this study was that the anti-inflammatory effect of flavonoid compounds may be partly attributable to inhibite FOS, PTGS2 expression, inhibite of IL-1beta release, and block the MAPK pathway and toll-like receptor pathway.

IL-1beta release in human lung tissue and mononuclear cells is dependent on pneumococcal pneumolysin

IL-1beta release in human lung tissue and mononuclear cells is dependent on pneumococcal pneumolysin

Effect of the addition of pentoxifylline on the therapeutic and inflammatory response in patients with cutaneous leishmaniasis: A Randomized Placebo Controlled Trial

Effect of the addition of pentoxifylline on the therapeutic and inflammatory response in patients with cutaneous leishmaniasis: A Randomized Placebo Controlled Trial

Biodegradable chitosan microparticles induce delayed STAT-1 activation and lead to distinct cytokine responses in differentially polarized human macrophages in vitro

Current data suggest that chitosan activates wound macrophages to release endogenous factors that guide mesenchymal stem cell (MSC) to bone fractures. We tested the hypothesis that chitosan, a polymer containing glucosamine and N-acetyl glucosamine, stimulates macrophages in different polarization states to release functional MSC chemokines and mainly anabolic factors. Low-serum conditioned medium was collected from M0, M1 and M2a U937 macrophages previously differentiated with phorbol myristate acetate (PMA) and exposed or not for 24h to chitosan microparticles (80% degree of deacetylation, DDA, 130kDa). Chitosan particles were highly phagocytosed. Chitosan enhanced anabolic factor release from M0 and M2a macrophages (MCP-1, IP-10, MIP-1beta, IL-1ra, IL-10, PDGF), and IL-1beta release, with 25- to 400-fold excess IL-1ra over IL-1beta. In M1 macrophages, chitosan enhanced IL-1beta without enhancing or suppressing inflammatory factor release (IL-6, IP-10, IL-8). M0 and M2a macrophages, with or without chitosan stimulation, produced conditioned medium that promoted 2-fold more MSC chemotaxis than low-serum control medium, while M1-conditioned medium failed to induce MSC chemotaxis. Acetylated chitosan induced U937 macrophages to release IL-1ra without STAT-6 activation, and also induced a delayed STAT-1 activation/IP-10 release response that was not observed using non-biodegradable chitosan (98% DDA, 130kDa). In primary human macrophages, acetylated chitosan enhanced IL-1ra release without inducing IL-1beta, and required PMA priming to elicit STAT-1 activation and IP-10 release. We conclude that biodegradable chitosan particles enhance M0 and M2a macrophage anabolic responses independent of the IL4/STAT-6 axis, by inducing excess IL-1ra over IL-1beta and more chemokine release, without altering their inherent capacity to attract MSCs.

HIV-1 Vpr activates the NLRP3 inflammasome in primary human microglia

Human immunodeficiency virus type 1 (HIV-1) infects microglia and macrophages in the brain during seroconversion and eventually results in neuroinflammation and neurodegeneration in susceptible individuals. Inflammasomes are cytosolic protein complexes that serve as platforms for caspase-1 activation and ensuing cleavage and release of IL-1beta and IL-18. Our group recently showed that HIV-1 infection of human microglia induces the NLRP3 inflammasome. The viral protein R (Vpr) is an accessory protein encoded by HIV-1, which is essential for macrophage infection and is detected in serum and cerebrospinal fluid of HIV-infected patients.

Differential polarization between resident microglia and brain infiltrating macrophages in mouse models of Alzheimer's disease

Human immunodeficiency virus type 1 (HIV-1) infects microglia and macrophages in the brain during seroconversion and eventually results in neuroinflammation and neurodegeneration in susceptible individuals. Inflammasomes are cytosolic protein complexes that serve as platforms for caspase-1 activation and ensuing cleavage and release of IL-1beta and IL-18. Our group recently showed that HIV-1 infection of human microglia induces the NLRP3 inflammasome. The viral protein R (Vpr) is an accessory protein encoded by HIV-1, which is essential for macrophage infection and is detected in serum and cerebrospinal fluid of HIV-infected patients. We investigated the actions of extracellular and intracellular Vpr expression on inflammasome activation in cells of monocytic lineage. Extracellular Vpr exposure to differentiated human monocytic cells (THP-1) and primary human microglia caused caspase-1 activation and reduced cell viability (p b 0.01). Vpr exposure also significantly increased transcription and secretion of IL-1beta and IL-18 (p b 0.001) although this effect was eliminated in THP-1 cells deficient in NLRP3. Infection of THP-1s and microglia with HIV-1 HxBruR−E+ (Vprdeficient) showed significantly reduced caspase-1 activation and IL1beta production (p b 0.01) compared to cells infected with HIV-1 HxBruRE (Vpr-encoding). To examine the effects of intracellular Vpr expression on inflammasome activation, THP-1 cells were transfected with Vpr, revealing enhanced caspase-1 activation and reduced cellular viability (p b 0.001) but without significant IL-1beta release. These observations indicated that extracellular Vpr activates the NLRP3 inflammasome by acting as both Signals 1 and 2. Thus, Vpr-induced NLRP3 inflammasome activation likely contributes to HIV-1 associated neuroinflammation.

PS-336 Influence Of N-acetylcysteine Amide (naca) On The Inflammasome Pathway. A Study On Neonatal Pigs

Background and aims Severe perinatal hypoxia contributes to approximately 6% of spastic cerebral palsy (CP). Studies have indicated an association between elevation of IL-1beta after perinatal asphyxia and the development of CP. The NLRP3 Inflammasome complex may lead to release of the cytokines IL-1beta and IL-18 and cell death. Reactive oxygen species (ROS) have been proposed to be an upstream inducer of this complex and the anti-oxidant N-Acetylcysteine amide (NACA) may provide organ protection after hypoxia. Objectives To map inflammasome activation in specific brain regions of the pig after neonatal hypoxia-reoxygenation and to investigate if the expression of different proteins in this pathway are modulated by NACA. Study design ([Table 1][1]). ELISA was used to measure IL-1b protein in cerebral cortex and Realtime PCR for mRNA expression of NLRP3, ASC, IL-1b and Il18 in cortex, cerebellum, hippocampus and striatum. ![Abstract PS-336 Table 1][2] Abstract PS-336 Table 1 Study design: Fifty-four newborn piglets, age 1236h, were included. Invasive blood pressure, EEG and ECG were measured continuously. One control group (n = 6) and 4 experimental groups (n = 12), exposed to global hypoxia, until BE was either -15 or -20 mmol/l (moderate/severe asphyxia with or without NACA) The pigs were observed for 9.5 h Results After severe hypoxia the protein expression of IL-1b in cerebral cortex was reduced for the NACA treated pigs vs. saline, p < 0.05. When comparing all the pigs treated with NACA vs. saline after hypoxia Fold Change of ASC in cortex was significantly reduced, p ([Table 2][3] ). ![Abstract PS-336 Table 2][2] Abstract PS-336 Table 2 Fold change for NLRP3, ASC, IL-18 and IL-1b measured in cerebral cortex. Significant difference in Fold Change of ASC (*), p < 0.05. All values are Median values In hippocampus, cortex and Striatum Fold Change of IL-1b was elevated in all the hypoxia groups compared with the control group, p Conclusion NACA reduces the protein expression of Il-1beta and mRNA-expression of ASC in cortex after hypoxia. This may indicate that NACA has some neuroprotective abilities after perinatal asphyxia. Upcoming analyses of histopathology and injury markers will elucidate possible neuroprotective effects of NACA treatment following birth asphyxia. [1]: #F1 [2]: pending:yes [3]: #F2

177: The NLRP3 inflammasome is regulated by phosphorylation and ubiquitinylation

NLRP3 is an intracellular pattern-recognition receptor that senses many different activators including bacterial toxins and endogenous danger signals. Upon activation, the NLRP3 inflammasome is formed, leading to the secretion of pro-inflammatory cytokines and cell death. Because of its involvement in many diseases such as atherosclerosis and Alzheimer’s Disease, a better understanding of NLRP3 regulation may help to shape therapeutic approaches. The aim of this study was to identify post-translational modifications on NLRP3 and their impact on inflammasome activation. We applied mass spectrometry of NLRP3 to identify post-translational modifications. We identified ubiquitinated and phosphorylated residues. To analyze the function of the modifications, we mutated the residues in NLRP3 expression plasmids and used them for reconstitution of NLRP3-deficient macrophages or overexpression studies in HEK293T cells. Ubiquitinylated residues were found in the NACHT and LRR domains of NLRP3 and mutation of these residues resulted in hyperactive NLRP3. In addition, mutation of one of the phosphorylated residues to a phosphomimetic residue showed reduced IL-1beta release and caspase-1 cleavage upon NLRP3 inflammasome activation, whereas mutation to alanine had no effect. Further studies in HEK293T cells found that the recruitment of the downstream adaptor ASC as well as the self-interaction of NLRP3 were reduced with the phosphomimetic residue. Thus, NLRP3 is likely regulated by post-translational modification and needs to be deubiquitinylated and possibly dephosphorylated before activation. Knowledge about the modifications and the pathways involved may help shape novel strategies for treating diseases that involve NLRP3.

Effects of N-acetyl-glucosamine-coated glycodendrimers as biological modulators in the B16F10 melanoma model in vivo

Glyco-coat changes on cancer cells due to aberrant glycosylation are potential targets for immune recognition through lectin-like receptors on immune cells. These cells include natural killer (NK), CD8+ and CD4+ lymphocytes, all reported to have, together with cytokines, important functions in antitumor immunity. The aim of this study was to evaluate a possible role of synthetic monodisperse multivalent neo-glycoconjugates, namely glycodendrimers, as a new approach to anticancer immune modulation through carbohydrate-mediated immune recognition. Octavalent polyamidoamine dendrimers functionalized with N-acetyl-glucosamine residues (PAMAM-GlcNAc8), with in vitro high affinity for the recombinant lymphocyte receptor NKR-P1A, were employed. To follow the fate of the compound, a fluorescent marker was conjugated to the tetra-branched semi-component of the dendrimer. Tumor development and immunity were evaluated in C57BL/6 mice. Animals were inoculated with B16F10 melanoma cells and underwent different protocols of PAMAM-GlcNAc8 administration. Advantages on survival and reduction of tumor growth were obtained in dose-dependent manner, by IP route. Increase of CD69+ cells in the spleen and their appearance inside the tumors, early progressive release of IL-1beta, a later production of INFgamma and IL-2 concomitant to an increment of CD4+ cells were observed. Cytotoxicity assays, performed ex vivo, showed an enhanced NK cell activity proportioned to the percentage of activated NK cells. Our data suggest that well-defined multivalent neo-glycoconjugates can stimulate an antitumor immune response engaging both innate and acquired immunity.

Association ofTLR2Gene Polymorphisms With Ocular Behçet's Disease in a Chinese Han Population

TLR2, TLR4, TLR8, and TLR9 have been reported to be associated with several autoimmune diseases. The current study aimed to explore whether singe nucleotide polymorphisms (SNPs) of these four genes were associated with ocular Behçet's disease (BD), Vogt-Koyanagi-Harada (VKH) syndrome, acute anterior uveitis (AAU) with or without ankylosing spondylitis (AS), or pediatric uveitis in Han Chinese. Genotyping was performed by PCR-restriction fragment length polymorphism. The first stage study comprised 400 ocular BD patients, 400 VKH syndrome patients, 400 AAU ± AS patients, 400 pediatric uveitis patients and 600 healthy subjects. The second stage included 438 ocular BD patients and 1000 healthy subjects. Allele and genotype frequencies were compared between patients and controls using the χ(2) test. Real-time PCR was used to detect mRNA expression from PBMCs obtained from healthy controls. Levels of TNF-α, IL-6, IL-10, and IL-1beta in culture supernatants were measured by ELISA. In the first stage study, only the frequencies of the rs2289318/TLR2 genotype A and C allele and rs3804099/TLR2 genotype CT were significantly higher in ocular BD patients (Pc = 0.048; Pc = 0.008; Pc = 0.005, respectively) compared with controls. The second stage and combined studies confirmed the association (Pc = 0.001; Pc = 6.89E-06, Pc = 2.426E-06, respectively). TLR2 mRNA expression in PBMCs was increased in healthy carriers of the CC genotype of rs2289318/TLR2 and TT genotype of rs3804099/TLR2 following stimulation with peptidoglycan (PGN; P = 0.028; P = 0.004, respectively). No effect of the various TLR2 rs2289318 and rs3804099 genotypes on the release of TNF-α, IL-6, IL-10, and IL-1beta could be detected. This study provides evidence that the TLR2 gene is involved in the susceptibility to ocular BD.

Lipid-Core Nanocapsules Improve the Effects of Resveratrol Against A&lt;I&gt;β&lt;/I&gt;-Induced Neuroinflammation

Resveratrol, a natural polyphenolic compound, has attracted considerable interest for its anti-inflammatory and neuroprotective properties. However, the biological effects of resveratrol appear strongly limited because it is photosensitive, easily oxidized, and has unfavorable pharmacokinetics. The present study aimed to elucidate the effect of resveratrol on Abeta-triggered neuroinflammation by comparing the effects of free resveratrol (RSV) treatment with those of treatment with resveratrol-loaded lipid-core nanocapsules (RSV-LNC). Organotypic hippocampal cultures were stimulated by Abeta1-42 with or without different concentrations of RSV or RSV-LNC. We found that Abeta triggered a harmful neuroinflammation process in organotypic hippocampal cultures. Pre- and co-treatments with RSV-LNC were able to protect cultures against ROS formation and cell death induced by Abeta, possibly through sustained blocking of TNF-alpha, IL-1beta, and IL-6 release. Furthermore, RSV-LNC was able to increase IL-10 release even in the presence of Abeta and prevent or decrease both glial and JNK activation. On the other hand, both pre- and co-treatment with RSV exhibited a lower ability to prevent or decrease neuroinflammation, ROS formation, and cell death, and failed to increase IL-10 release. Our findings suggest that modulation of neuroinflammation through a combination of resveratrol and a lipid-core nanocapsule-based delivery system might represent a promising approach for preventing or delaying the neurodegenerative process triggered by Abeta. The results open new vistas to the interplay between inflammation and amyloid pathology.

LPS induces mediators of neuroinflammation, cell proliferation, and GFAP expression in human astrocytoma cells U373MG: the anti-inflammatory and anti-proliferative effect of guggulipid

Neuroinflammation has been considered to be an integrated part of human neurodegenerative diseases. In this study, we examined the effect of guggulipid on cell proliferation, nitrite release, interleukin IL-6 and IL-1 beta release, and expression of COX-2 and glial fibrillary acidic protein (GFAP) in LPS-stimulated U373MG cells. LPS significantly stimulated human astrocytoma cells U373MG by up-regulating these neuroinflammatory mediators. Guggulipid alone had no effect on the cell proliferation of U373MG cells. The up regulation in nitrite release, cell proliferation, and release of IL-6 and IL-1 beta in LPS stimulated human astrocytoma cells were dose-dependently inhibited by co-treatment with guggulipid. The expression level of COX-2 and GFAP proteins was up regulated by LPS but the increased level of COX-2 and GFAP was significantly down regulated by treatment with guggulipid. These data indicate that guggulipid has a modulatory effect on all these parameters, which might explain its beneficial effect in the treatment of neuroinflammation-associated disorders directly relating to human aspects.

ATP-mediated inflammasome activation is efficiently inhibited by acetylcholine

Event Abstract Back to Event ATP-mediated inflammasome activation is efficiently inhibited by acetylcholine Mira Kuellmar1*, Andreas Hecker1, Sigrid Wilker1, Srebrena Atanasova1, Anna Zakrzewicz1, Marion Meixner1, Andreas Kaufmann2, Wolfgang Kummer1, Winfried Padberg1 and Veronika Grau1 1 Justus-Liebig-University Giessen, Germany 2 Philipps-University Marburg, Germany The release of IL-1beta by mononuclear phagocytes is tightly controlled. Lipopolysaccharide induces synthesis of pro-IL-1beta. A second danger signal such as ATP triggers inflammasome activation resulting in cleavage and secretion of mature IL-1beta. Acetylcholine is known for its anti-inflammatory effects on the innate immune system. In this study, we test the hypothesis, that acetylcholine dampens ATP-mediated activation of the inflammasome. Human monocytic U937 cells were primed with lipopolysaccharide for 3 hours. In addition, mononuclear blood leukocytes from healthy human donors and from blood vessels of renal rat allografts undergoing acute rejection were investigated. Cells were stimulated with BzATP or nigericin in the presence or absence of cholinergics and inhibitors of NO-synthases. The release of IL-1beta was measured by ELISA. Acetylcholine and nicotine efficiently inhibited BzATP-induced release of IL-1beta from U937 cells and primary blood leukocytes. Mecamylamine, strychnine, alpha-bungarotoxin and inhibitors of NO-synthases reversed this effect. Nicotinic agonists did not inhibit IL-1beta release induced by nigericin. Activated leukocytes from allografts, which are known to synthesize acetylcholine, did not release IL-1beta unless acetylcholine esterase was added together with BzATP. As revealed by RT-PCR, alpha-9 subunits of acetylcholine receptors and NO-synthases 1-3 were expressed. We describe a novel anti-inflammatory mechanism of acetylcholine. Acetylcholine inhibits ATP-mediated inflammasome activation. Our pharmacological data suggest that P2X7 receptors, alpha-9 nicotinic receptor subunits and NO-synthases are involved in the signaling cascade. We suggest that this novel anti-inflammatory pathway is active in renal allografts during acute rejection, where endogenous leukocytic acetylcholine prevents IL-1beta secretion into the blood stream. Keywords: IL-1beta, Inflammasome, P2X7 ATP Receptor., lipopolysaccharide, Cholinergics Conference: 15th International Congress of Immunology (ICI), Milan, Italy, 22 Aug - 27 Aug, 2013. Presentation Type: Abstract Topic: Innate immunity Citation: Kuellmar M, Hecker A, Wilker S, Atanasova S, Zakrzewicz A, Meixner M, Kaufmann A, Kummer W, Padberg W and Grau V (2013). ATP-mediated inflammasome activation is efficiently inhibited by acetylcholine . Front. Immunol. Conference Abstract: 15th International Congress of Immunology (ICI). doi: 10.3389/conf.fimmu.2013.02.00429 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 27 Mar 2013; Published Online: 22 Aug 2013. * Correspondence: Mrs. Mira Kuellmar, Justus-Liebig-University Giessen, Giessen, Germany, mira.kuellmar@chiru.med.uni-giessen.de Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Mira Kuellmar Andreas Hecker Sigrid Wilker Srebrena Atanasova Anna Zakrzewicz Marion Meixner Andreas Kaufmann Wolfgang Kummer Winfried Padberg Veronika Grau Google Mira Kuellmar Andreas Hecker Sigrid Wilker Srebrena Atanasova Anna Zakrzewicz Marion Meixner Andreas Kaufmann Wolfgang Kummer Winfried Padberg Veronika Grau Google Scholar Mira Kuellmar Andreas Hecker Sigrid Wilker Srebrena Atanasova Anna Zakrzewicz Marion Meixner Andreas Kaufmann Wolfgang Kummer Winfried Padberg Veronika Grau PubMed Mira Kuellmar Andreas Hecker Sigrid Wilker Srebrena Atanasova Anna Zakrzewicz Marion Meixner Andreas Kaufmann Wolfgang Kummer Winfried Padberg Veronika Grau Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.