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Sample records for activation cytokine production

  1. Whole Blood Activation Results in Altered T Cell and Monocyte Cytokine Production Profiles by Flow Cytometry

    NASA Technical Reports Server (NTRS)

    Crucian, Brian E.; Sams, Clarence F.

    2001-01-01

    An excellent monitor of the immune balance of peripheral circulating cells is to determine their cytokine production patterns in response to stimuli. Using flow cytometry, a positive identification of cytokine producing cells in a mixed culture may be achieved. Recently, the ability to assess cytokine production following a whole-blood activation culture has been described. In this study, whole blood activation was compared to traditional PBMC activation and the individual cytokine secretion patterns for both T cells, T cell subsets and monocytes was determined by flow cytometry. RESULTS: For T cell cytokine assessment (IFNg/IL-10 and IL-21/L-4) following PMA +ionomycin activation: (1) a Significantly greater percentages of T cells producing IFNgamma and IL-2 were observed following whole-blood culture and (2) altered T cell cytokine production kinetics were observed by varying whole blood culture times. Four-color analysiS was used to allow assessment of cytokine production by specific T cell subsets. It was found that IFNgamma production was significantly elevated in the CD3+/CD8+ T cell population as compared to the CD3+/CD8- population following five hours of whole blood activation. Conversely, IL-2 and IL-10 production were Significantly elevated in the CD3+/CD8- T cell population as compared to the CD3+/CD8+ population. Monocyte cytokine production was assessed in both culture systems following LPS activation for 24 hours. A three-color flow cytometric was used to assess two cytokines (IL-1a/IL-12 and TNFa/IL-10) in conjunction with CD14. Nearly all monocytes were stimulated to produce IL-1a, IL-12 and TNFa. equally well in both culture systems, however monocyte production of IL-10 was significantly elevated in whole blood culture as compared to PBMC culture. IL-12 producing monocytes appeared to be a distinct subpopulation of the IL-1a producing set, whereas IL-10 and TNFa producing monocytes were largely mutually exclusive. IL-10 and TNFa producing

  2. Carvedilol differentially regulates cytokine production from activated human peripheral blood mononuclear cells.

    PubMed

    Yang, Shih-Ping; Ho, Ling-Jun; Cheng, Shu-Meng; Hsu, Yu-Lin; Tsao, Tien-Ping; Chang, Deh-Ming; Lai, Jenn-Haung

    2004-05-01

    Chronic inflammation is one of the important mechanisms involved in atherosclerosis formation. The activated monocytes and their secreted cytokines contribute significantly to this inflammatory process. Here we examined the effects of carvedilol, a recently introduced cardio-protective alpha-1- and beta-receptor blocker, on cytokine production from various stimuli-activated human immune effector cells. By ELISA analysis, we showed that carvedilol inhibited interferon-gamma (IFN-gamma), but enhanced interleukin (IL)-12 production in phytohemagglutinin (PHA)- and concanavalin A (ConA)-stimulated human peripheral blood mononuclear cells (PBMCs). The production of tumor necrosis factor-alpha (TNF-alpha) was marginally affected. When purified monocytes were examined, we observed the consistent up-regulation of IL-12 production while both IL-10 and TNF-alpha were unaffected or marginally down-regulated, respectively, by carvedilol. In agreement with the observation in monocytes, the production of IL-12 from activated macrophages was also up-regulated by carvedilol. We concluded that carvedilol might mediate its therapeutic effects through differentially regulating cytokine production from activated mononuclear cells, including at least monocytes and macrophages.

  3. Monocyte activation and cytokine production in Malawian children presenting with P. falciparum malaria.

    PubMed

    Mandala, W L; Msefula, C L; Gondwe, E N; Drayson, M T; Molyneux, M E; MacLennan, C A

    2016-05-01

    Malaria in malaria-naïve adults is associated with an inflammatory response characterized by expression of specific activation markers on innate immune cells. Here, we investigate activation and adhesion marker expression, and cytokine production in monocytes from children presenting with cerebral malaria (CM, n = 36), severe malarial anaemia (SMA, n = 42) or uncomplicated malaria (UM, n = 66), and healthy aparasitemic children (n = 52) in Blantyre, Malawi. In all malaria groups, but particularly in the two severe malaria groups, monocyte expression of CD11b, CD11c, CD18, HLA-DR and CD86, and percentages of TNF-α- and IL-6-producing monocytes were lower than in healthy controls, while expression of CD11a, TLR2 and TLR4 was lower in children with severe malaria compared with controls. These levels mostly normalized during convalescence, but percentages of cytokine-producing monocytes remained suppressed in children with SMA. In all malaria groups, especially the SMA group, a greater proportion of monocytes were loaded with haemozoin than among controls. In a P. falciparum hyperendemic area, monocytes in children with acute symptomatic malaria have reduced expression of adhesion molecules and activation markers and reduced inflammatory cytokine production. This immune suppression could be due to accumulation of haemozoin and/or previous exposure to P. falciparum.

  4. Moxibustion activates host defense against herpes simplex virus type I through augmentation of cytokine production.

    PubMed

    Takayama, Yuko; Itoi, Manami; Hamahashi, Takashi; Tsukamoto, Noriyuki; Mori, Kazuya; Morishita, Daisuke; Wada, Kumiko; Amagai, Takashi

    2010-09-01

    Moxibustion is a technique used in traditional oriental medicine, the aim of which is to cure and/or prevent illness by activating a person's ability for self-healing. In this study, we assessed how moxibustion would affect the immune system and whether it would augment protective immunity. Mice were treated with moxibustion at Zusanli (ST36) acupoints; we analyzed mortality and cytokine activity in sera after infection with herpes simplex virus type 1 (HSV-1), and cytokine gene expression in the skin and the spleen without a virus challenge. Our study demonstrates that pretreatment of BALB/c mice with moxibustion resulted in a marked increase in the survival rate after infection with lethal doses of HSV-1, and elevated serum levels of IL-1β and IFN-γ on days 1 and 6 post-infection with HSV-1. Semi-quantitative RT-PCR assay showed that moxibustion treatment augmented the expression of IL-1α, IL-1β, IL-6, universal-IFN-α, MIP-1α, and TNF-α mRNA in the skin, and IL-1α, IL-1β, IL-12p40, IL-15, u-IFN-α, MIP-1α, and TNF-α mRNA in the spleen. Moreover, moxibustion induces augmentation of natural killer cell activity. Collectively, our study demonstrates that moxibustion activates protective responses against HSV-1 infection through the activation of cytokine production including IFN, and of NK cells.

  5. Effect of polyclonal activators on cytokine production by blood cells and by malignant breast cancer cells.

    PubMed

    Kunts, T A; Karpukhina, K V; Mikhaylova, E S; Marinkin, I O; Varaksin, N A; Autenshlyus, A I; Lyakhovich, V V

    2016-01-01

    The production of cytokines by peripheral blood cells and biopsy specimens of tumors stimulated by polyclonal activators (PAs) was evaluated in 34 patients with invasive ductal breast carcinoma using enzyme-linked immunosorbent assay (ELISA). Positive correlation between the stimulation index of polyclonal activators (SIPA) for IL-18 production by the tumor and the relative content of poorly differentiated cells was revealed. The latter, in turn, was positively correlated with the numbers of normal and pathologic mitoses and the degree of malignancy. Cancer cells can produce IL-18, which is involved in the process of angiogenesis, stimulates invasion and metastasis. Decrease in SIPA for the production of IL-6 and GCSF by peripheral blood cells could serve as an indicator of malignant progression in invasive ductal breast carcinoma.

  6. Potassium humate inhibits complement activation and the production of inflammatory cytokines in vitro

    SciTech Connect

    van Rensburg, C.E.J.; Naude, P.J.

    2009-08-15

    The effects of brown coal derived potassium humate on lymphocyte proliferation, cytokine production and complement activation were investigated in vitro. Potassium humate increased lymphocyte proliferation of phytohaemaglutinin A (PHA) and pokeweed mitogen (PWM) stimulated mononuclear lymphocytes (MNL) in vitro from concentrations of 20 to 80 {mu} g/ml, in a dose dependant manner. On the other hand potassium humate, at 40 {mu} g/ml, significantly inhibited the release of TNF-alpha, IL-1 beta, IL-6 and IL-10 by PHA stimulated MNL. Regarding complement activation it was found that potassium humate inhibits the activation of both the alternative and classical pathways without affecting the stability of the red blood cell membranes. These results indicate that the anti-inflammatory potential of potassium humate could be partially due to the inhibition of pro-inflammatory cytokines responsible for the initiation of these reactions as well as inhibition of complement activation. The increased lymphocyte proliferation observed, might be due to increased IL-2 production as previously been documented.

  7. Whole Blood Activation Results in Enhanced Detection of T Cell and Monocyte Cytokine Production by Flow Cytometry

    NASA Technical Reports Server (NTRS)

    Sams, Clarence F.; Crucian, Brian E.

    2001-01-01

    An excellent monitor of the immune balance of peripheral circulating cells is to determine their cytokine production patterns in response to stimuli. Using flow cytometry a positive identification of cytokine producing cells in a mixed culture may be achieved. Recently, the ability to assess cytokine production following a wholeblood activation culture has been described. We compared whole blood culture to standard PBMC culture and determined the individual cytokine secretion patterns for both T cells and monocytes via flow cytometry. For T cells cytokine assessment following PMA +ionomycin activation: (1) a significantly greater percentages of T cells producing IFNgamma and IL-2 were observed following whole-blood culture; (2) altered T cell cytokine production kinetics were observed by varying whole blood culture times. In addition, a four-color cytometric analysis was used to allow accurate phenotyping and quantitation of cytokine producing lymphocyte populations. Using this technique we found IFNgamma production to be significantly elevated in the CD3+/CD8+ T cell population as compared to the CD3+/CD8- population following five hours of whole blood activation. Conversely, IL-2 and IL-10 production were significantly elevated in the CD3+/CD8- T cell population as compared to the CD3+/CD8+ population. Monocyte cytokine production was assessed in both culture systems following LPS activation for 24 hours. A three-color flow cytometric was used to assess two cytokines in conjunction with CD 14. The cytokine pairs used for analysis were IL-1a/IL-12, and IL-10ITNFa. Nearly all monocytes were stimulated to produce IL-1a, IL-12 and TNFalpha equally well in both culture systems. Monocyte production of IL-10 was significantly elevated following whole blood culture as compared to PBMC culture. IL-12 producing monocytes appeared to be a distinct subpopulation of the IL-1a producing set, whereas IL-10 and TNFa producing monocytes were largely mutually exclusive. IL-10 and

  8. Analgesic activity of piracetam: effect on cytokine production and oxidative stress.

    PubMed

    Navarro, Suelen A; Serafim, Karla G G; Mizokami, Sandra S; Hohmann, Miriam S N; Casagrande, Rubia; Verri, Waldiceu A

    2013-04-01

    Piracetam is a prototype of nootropic drugs used to improve cognitive impairment. However, recent studies suggest that piracetam can have analgesic and anti-inflammatory effects. Inflammatory pain is the result of a process that depends on neutrophil migration, cytokines and prostanoids release and oxidative stress. We analyze whether piracetam has anti-nociceptive effects and its mechanisms. Per oral pretreatment with piracetam reduced in a dose-dependent manner the overt pain-like behavior induced by acetic acid, phenyl-p-benzoquinone, formalin and complete Freund's adjuvant. Piracetam also diminished carrageenin-induced mechanical and thermal hyperalgesia, myeloperoxidase activity, and TNF-α-induced mechanical hyperalgesia. Piracetam presented analgesic effects as post-treatment and local paw treatment. The analgesic mechanisms of piracetam were related to inhibition of carrageenin- and TNF-α-induced production of IL-1β as well as prevention of carrageenin-induced decrease of reduced glutathione, ferric reducing ability and free radical scavenging ability in the paw. These results demonstrate that piracetam presents analgesic activity upon a variety of inflammatory stimuli by a mechanism dependent on inhibition of cytokine production and oxidative stress. Considering its safety and clinical use for cognitive function, it is possible that piracetam represents a novel perspective of analgesic.

  9. Premalignant Oral Lesion Cells Elicit Increased Cytokine Production and Activation of T-cells

    PubMed Central

    JOHNSON, SARA D.; LEVINGSTON, CORINNE; YOUNG, M. RITA I.

    2016-01-01

    Background Head and neck squamous cell carcinomas (HNSCC) are known to evade the host immune response. How premalignant oral lesions modulate the immune response, however, has yet to be elucidated. Materials and Methods A mouse model of oral carcinogenesis was used to determine how mediators from premalignant oral lesion cells vs. HNSCC cells impact on immune cytokine production and activation. Results Media conditioned by premalignant lesion cells elicited an increased production of T cell-associated cytokines and proinflammatory mediators from cervical lymph node cells compared to media conditioned by HNSCC cells or media alone. In the presence of premalignant lesion cell-conditioned media, CD4+ T cell expression of the IL-2 receptor CD25 and CD8+ T cell expression of the activation marker CD69 was greater, compared to what was induced in HNSCC cell-conditioned media or media alone. Conclusion Premalignant lesion cells promote a proinflammatory environment and induce immune changes before HNSCC tumors are established. PMID:27354582

  10. Bestatin, an inhibitor for aminopeptidases, modulates the production of cytokines and chemokines by activated monocytes and macrophages.

    PubMed

    Lkhagvaa, Battur; Tani, Kenji; Sato, Keiko; Toyoda, Yuko; Suzuka, Chiyuki; Sone, Saburo

    2008-12-01

    The aim of this study was to clarify the effect of bestatin, an aminopeptidase inhibitor, on the production of cytokines from peripheral blood monocytes and alveolar macrophages (AM). Human monocytes isolated from peripheral blood of healthy volunteers were incubated with or without lipopolysaccharide (LPS) in the presence or absence of bestatin. AM obtained from patients with sarcoidosis were incubated in the presence or absence of bestatin. The concentration of cytokines in the culture supernatant was determined by enzyme-linked immunosorbent assay. The expression of mRNA was determined by reverse transcription polymerase chain reaction. Bestatin suppressed the production and expression of proinflammatory cytokines and chemokines, interleukin (IL)-6, CXCL8/IL-8, CCL3/macrophage inflammatory protein (MIP)-1alpha by LPS-stimulated monocytes. The mean percentage of the inhibition of IL-6, CXCL8/IL-8, CCL3/MIP-1alpha by bestatin at a concentration of 50 microg/mL was 71.2%, 29.7% and 61.0%, respectively. On the other hand, bestatin increased the production and mRNA expression of IL-10 by LPS-stimulated monocytes. The treatment with bestatin significantly inhibited the production of IL-6 and CXCL8/IL-8 by AM from patients with sarcoidosis. The data presented here indicate that bestatin suppresses the production of the pro-inflammatory cytokines and stimulates the anti-inflammatory cytokine by activated human monocytes. This study suggests that bestatin may be useful as an anti-inflammatory agent in various inflammatory diseases.

  11. Propolis Ethanol Extract Stimulates Cytokine and Chemokine Production through NF-κB Activation in C2C12 Myoblasts

    PubMed Central

    Washio, Kohei; Kobayashi, Mao; Saito, Natsuko; Amagasa, Misato; Kitamura, Hiroshi

    2015-01-01

    Myoblast activation is a triggering event for muscle remodeling. We assessed the stimulatory effects of propolis, a beehive product, on myoblasts. After an 8 h treatment with 100 μg/mL of Brazilian propolis ethanol extract, expression of various chemokines, including CCL-2 and CCL-5, and cytokines, such as IL-6, increased. This propolis-induced cytokine production appears to depend on NF-κB activation, because the IKK inhibitor BMS-345541 repressed mRNA levels of CCL-2 by ~66%, CCL-5 by ~81%, and IL-6 by ~69% after propolis treatment. Supernatant from propolis-conditioned C2C12 cells upregulated RAW264 macrophage migration. The supernatant also stimulated RAW264 cells to produce angiogenic factors, including VEGF-A and MMP-12. Brazilian green propolis therefore causes myoblasts to secrete cytokines and chemokines, which might contribute to tissue remodeling of skeletal muscle. PMID:26604971

  12. Activation of cytokine production by secreted phospholipase A2 in human lung macrophages expressing the M-type receptor.

    PubMed

    Granata, Francescopaolo; Petraroli, Angelica; Boilard, Eric; Bezzine, Sofiane; Bollinger, James; Del Vecchio, Luigi; Gelb, Michael H; Lambeau, Gerard; Marone, Gianni; Triggiani, Massimo

    2005-01-01

    Secreted phospholipases A(2) (sPLA(2)) are enzymes released in plasma and extracellular fluids during inflammatory diseases. Because human group IB and X sPLA(2)s are expressed in the lung, we examined their effects on primary human lung macrophages (HLM). Both sPLA(2)s induced TNF-alpha and IL-6 release in a concentration-dependent manner by increasing their mRNA expression. This effect was independent of their enzymatic activity because 1) the capacity of sPLA(2)s to mobilize arachidonic acid from HLM was unrelated to their ability to induce cytokine production; and 2) two catalytically inactive isoforms of group IB sPLA(2) (bromophenacyl bromide-inactivated human sPLA(2) and the H48Q mutant of the porcine sPLA(2)) were as effective as the catalytically active sPLA(2)s in inducing cytokine production. HLM expressed the M-type receptor for sPLA(2)s at both mRNA and protein levels, as determined by RT-PCR, immunoblotting, immunoprecipitation, and flow cytometry. Me-indoxam, which decreases sPLA(2) activity as well as binding to the M-type receptor, suppressed sPLA(2)-induced cytokine production. Incubation of HLM with the sPLA(2)s was associated with phosphorylation of ERK1/2, and a specific inhibitor of this pathway, PD98059, significantly reduced the production of IL-6 elicited by sPLA(2)s. In conclusion, two distinct sPLA(2)s produced in the human lung stimulate cytokine production by HLM via a mechanism that is independent of their enzymatic activity and involves activation of the ERK1/2 pathway. HLM express the M-type receptor, but its involvement in eliciting cytokine production deserves further investigation.

  13. Activation of platelet-activating factor receptor in SZ95 sebocytes results in inflammatory cytokine and prostaglandin E2 production.

    PubMed

    Zhang, Qiwei; Seltmann, Holger; Zouboulis, Christos C; Travers, Jeffrey B

    2006-10-01

    Platelet-activating factor (PAF) is a group of phosphocholines with various biological effects mediated by the PAF receptor (PAF-R). Activation of the epidermal PAF-R induces the expression of inflammatory mediators, including cyclooxygenase-2 (COX-2) and prostaglandin E(2) (PGE(2)). The upregulation of COX-2 expression has been shown to be involved in sebocyte proliferation, sebaceous gland inflammation and carcinogenesis. The present study was designed to investigate whether PAF-R activation could induce the expression of COX-2 and production of PGE(2), as well as secretion of the inflammatory cytokine, interleukin-8 (IL-8), in the immortalized sebaceous gland cell line SZ95. Using calcium mobilization studies, we first confirmed that PAF can signal through PAF-R in SZ95 sebocytes. We then found that the production of IL-8 was induced following treatment with PAF-R agonist, however blocked by a specific PAF-R antagonist. Induction of COX-2 expression and increased PGE(2) production were observed in SZ95 sebocytes after PAF-R activation. Finally, it was demonstrated that the production of PGE(2), induced by PAF-R activation and mediated by COX-2 expression, was blocked following PAF-R antagonism in SZ95 sebocytes. These studies suggest that SZ95 sebocytes express functional PAF-Rs and PAF-Rs are involved in regulating the expression of inflammatory mediators, including COX-2, PGE(2) and IL-8.

  14. The Cytotoxic Enterotoxin of Aeromonas hydrophila Induces Proinflammatory Cytokine Production and Activates Arachidonic Acid Metabolism in Macrophages

    PubMed Central

    Chopra, A. K.; Xu, X.-J.; Ribardo, D.; Gonzalez, M.; Kuhl, K.; Peterson, J. W.; Houston, C. W.

    2000-01-01

    An aerolysin-related cytotoxic enterotoxin (Act) of Aeromonas hydrophila possesses multiple biological activities, which include its ability to lyse red blood cells, destroy tissue culture cell lines, evoke a fluid secretory response in ligated intestinal loop models, and induce lethality in mice. The role of Act in the virulence of the organism has been demonstrated. In this study, we evaluated the potential of Act to induce production of proinflammatory cytokines associated with Act-induced tissue injury and Act's capacity to activate in macrophages arachidonic acid (AA) metabolism that leads to production of eicosanoids (e.g., prostaglandin E2 [PGE2]). Our data indicated that Act stimulated the production of tumor necrosis factor alpha and upregulated the expression of genes encoding interleukin-1β (IL-1β) and IL-6 in the murine macrophage cell line RAW264.7. Act also activated transcription of the gene encoding inducible nitric oxide synthase. Act evoked the production of PGE2 coupled to the cyclooxygenase-2 (COX-2) pathway. AA is a substrate for PGE2, and Act produced AA from phospholipids by inducing group V secretory phospholipase A2. We also demonstrated that Act increased cyclic AMP (cAMP) production in macrophages. cAMP, along with PGE2, could potentiate fluid secretion in animal models because of infiltration and activation of macrophages resulting from Act-induced tissue injury. After Act treatment of RAW cells, we detected an increased translocation of NF-κB and cAMP-responsive element binding protein (CREB) to the nucleus using gel shift assays. Act also upregulated production of antiapoptotic protein Bcl-2 in macrophages, suggesting a protective role for Bcl-2 against cell death induced by proinflammatory cytokines. The increased expression of genes encoding the proinflammatory cytokines, COX-2, and Bcl-2 appeared correlated with the activation of NF-κB and CREB. This is the first report of the detailed mechanisms of action of Act from A

  15. Inhibition of NF-kB activation and cytokines production in THP-1 monocytes by 2-styrylchromones.

    PubMed

    Gomes, Ana; Capela, João P; Ribeiro, Daniela; Freitas, Marisa; Silva, Artur M S; Pinto, Diana C G A; Santos, Clementina M M; Cavaleiro, José A S; Lima, José L F C; Fernandes, Eduarda

    2015-01-01

    Nuclear factor kappa B (NF-kB) is one of the most important transcription factors whose modulation triggers a cascade of signaling events, namely the expression of many cytokines, enzymes, chemokines, and adhesion molecules, some of which being potential key targets for intervention in the treatment of inflammatory conditions. The 2-styrylchromones (2-SC) designation represents a well-recognized group of natural and synthetic chromones, vinylogues of flavones (2-phenylchromones). Several 2-SC were recently tested for their anti-inflammatory potential, regarding the arachidonic acid metabolic cascade, showing some motivating results. In addition, several flavones with structural similarities to 2-SC have shown NF-kB inhibitory properties. Hence, the aim of the present work was to continue the investigation on the interference of 2-SC in inflammatory pathways. Herein we report their effects on lipopolysaccharide (LPS)-induced NF-kB activation and consequent production of proinflammatory cytokines/chemokine, using a human monocytic cell line (THP-1). From the twelve 2-SC tested, three of them were able to significantly inhibit the NF-kB activation and to reduce the production of the proinflammatory cytokines/chemokine. The compound 3',4',5-trihydroxy-2- styrylchromone stood up as the most active in both assays, being a promising candidate for an anti-inflammatory drug.

  16. Regulatory T cell levels and cytokine production in active non-infectious uveitis: in-vitro effects of pharmacological treatment

    PubMed Central

    Molins, B; Mesquida, M; Lee, R W J; Llorenç, V; Pelegrín, L; Adán, A

    2015-01-01

    The aim of this study was to quantify the proportion of regulatory T cells (Treg) and cytokine expression by peripheral blood mononuclear cells (PBMCs) in patients with active non-infectious uveitis, and to evaluate the effect of in-vitro treatment with infliximab, dexamethasone and cyclosporin A on Treg levels and cytokine production in PBMCs from uveitis patients and healthy subjects. We included a group of 21 patients with active non-infectious uveitis and 18 age-matched healthy subjects. The proportion of forkhead box protein 3 (FoxP3)+ Treg cells and intracellular tumour necrosis factor (TNF)-α expression in CD4+ T cells was determined by flow cytometry. PBMCs were also either rested or activated with anti-CD3/anti-CD28 and cultured in the presence or absence of dexamethasone, cyclosporin A and infliximab. Supernatants of cultured PBMCs were collected and TNF-α, interleukin (IL)-10, IL-17 and interferon (IFN)-γ levels were measured by enzyme-linked immunosorbent assay (ELISA). No significant differences were observed in nTreg levels between uveitis patients and healthy subjects. However, PBMCs from uveitis patients produced significantly higher amounts of TNF-α and lower amounts of IL-10. Dexamethasone treatment in vitro significantly reduced FoxP3+ Treg levels in PBMCs from both healthy subjects and uveitis patients, and all tested drugs significantly reduced TNF-α production in PBMCs. Dexamethasone and cyclosporin A significantly reduced IL-17 and IFN-γ production in PBMCs and dexamethasone up-regulated IL-10 production in activated PBMCs from healthy subjects. Our results suggest that PBMCs from patients with uveitis express more TNF-α and less IL-10 than healthy subjects, and this is independent of FoxP3+ Treg levels. Treatment with infliximab, dexamethasone and cyclosporin A in vitro modulates cytokine production, but does not increase the proportion of FoxP3+ Treg cells. PMID:25354724

  17. Regulatory T cell levels and cytokine production in active non-infectious uveitis: in-vitro effects of pharmacological treatment.

    PubMed

    Molins, B; Mesquida, M; Lee, R W J; Llorenç, V; Pelegrín, L; Adán, A

    2015-03-01

    The aim of this study was to quantify the proportion of regulatory T cells (Treg ) and cytokine expression by peripheral blood mononuclear cells (PBMCs) in patients with active non-infectious uveitis, and to evaluate the effect of in-vitro treatment with infliximab, dexamethasone and cyclosporin A on Treg levels and cytokine production in PBMCs from uveitis patients and healthy subjects. We included a group of 21 patients with active non-infectious uveitis and 18 age-matched healthy subjects. The proportion of forkhead box protein 3 (FoxP3)(+) Treg cells and intracellular tumour necrosis factor (TNF)-α expression in CD4(+) T cells was determined by flow cytometry. PBMCs were also either rested or activated with anti-CD3/anti-CD28 and cultured in the presence or absence of dexamethasone, cyclosporin A and infliximab. Supernatants of cultured PBMCs were collected and TNF-α, interleukin (IL)-10, IL-17 and interferon (IFN)-γ levels were measured by enzyme-linked immunosorbent assay (ELISA). No significant differences were observed in nTreg levels between uveitis patients and healthy subjects. However, PBMCs from uveitis patients produced significantly higher amounts of TNF-α and lower amounts of IL-10. Dexamethasone treatment in vitro significantly reduced FoxP3(+) Treg levels in PBMCs from both healthy subjects and uveitis patients, and all tested drugs significantly reduced TNF-α production in PBMCs. Dexamethasone and cyclosporin A significantly reduced IL-17 and IFN-γ production in PBMCs and dexamethasone up-regulated IL-10 production in activated PBMCs from healthy subjects. Our results suggest that PBMCs from patients with uveitis express more TNF-α and less IL-10 than healthy subjects, and this is independent of FoxP3(+) Treg levels. Treatment with infliximab, dexamethasone and cyclosporin A in vitro modulates cytokine production, but does not increase the proportion of FoxP3(+) Treg cells.

  18. SoSoSo or its active ingredient chrysophanol regulates production of inflammatory cytokines & adipokine in both macrophages & adipocytes

    PubMed Central

    Hong-Kun, Rim; Phil-Dong, Moon; In-Hwa, Choi; Eun-Hee, Lee; Hyung-Min, Kim; Hyun-Ja, Jeong

    2013-01-01

    Background & objectives: Obesity is now considered as a major risk factor for the development of fatty liver diseases, cardiovascular diseases, and atherosclerosis. SoSoSo is a newly developed dietary supplement made of seven medicinal herbs. This study was aimed at examining the anti-obesity effect of SoSoSo or its active ingredient chrysophanol on the production of inflammatory cytokines and adipokine in macrophyage cell line RAW264 and 3T3-L1 adipocytes. Methods: No release was measured as a form of nitrite by Griess method. The production of inflammatory cytokines and adipokine were measured with the ELISA method. The m-RNA expression of each cytokine and adipokine were measured using RT-PCR. The nuclear proteins for NF-κB were analyzed with western blotting. Results: SoSoSo or chrysophanol significantly inhibited the nitric oxide production in lipopolysaccharide-stimulated RAW264 cells as well as in RAW264 cells-conditioned medium (CM)-treated 3T3-L1 cells. The production of interleukin (IL)-6 and tumour necrosis factor (TNF)-α were inhibited by SoSoSo or chrysophanol. In addition, SoSoSo or chrysophanol inhibited the activation of nuclear factor-κB in RAW264 cells. SoSoSo or chrysophanol inhibited the productions of IL-6, TNF-α, and monocyte chemoattractant protein-1 as well as the reduction of adiponectin production in CM-treated 3T3-L1 cells. Interpretation & conclusions: These results suggest a potential of SoSoSo or chrysophanol as a source of anti-inflammatory agent for obesity. Further in vivo studies would be required to confirm these findings. PMID:23481064

  19. Paeonia japonica, Houttuynia cordata, and Aster scaber water extracts induce nitric oxide and cytokine production by lipopolysaccharide-activated macrophages.

    PubMed

    Kim, Jin; Park, Chang-Shin; Lim, Yunsook; Kim, Hyun-Sook

    2009-04-01

    Natural products are increasingly recognized as potential targets for drug discovery and development. We previously reported that Paeonia japonica, Houttuynia cordata, and Aster scaber enhanced macrophage activation both in vitro and in vivo. In the present study we investigated the immunomodulating effects of these plants on lipopolysacharide (LPS)-stimulated macrophages. An aqueous extract of each plant was administered to female BALB/c mice every other day for 4 weeks. Peritoneal macrophages were then collected and incubated to examine the immunoreactivity of macrophages against LPS at different time points. The expression levels of inducible nitric oxide (NO) synthetase (iNOS), cyclooxygenase (COX)-2, and inhibitory factor kappaB alpha (IkappaBalpha) proteins and the production of NO metabolite (nitrite), prostaglandin (PG) E(2), and the pro-inflammatory cytokines interleukin-1beta, interleukin-6, and tumor necrosis factor-alpha were determined in the activated macrophages treated with extracts from each plant individually or combined. High levels of pro-inflammatory cytokines were produced by A. scaber-, P. japonica-, and H. cordata-treated macrophages following 24 hours of LPS stimulation. P. japonica, H. cordata, and A. scaber treatment also induced the production of nitrate by LPS-treated macrophages. Induction of iNOS mRNA and protein was also different in each group. PGE(2) secretion was up-regulated by all extract-treated macrophages at early time points; however, no significant differences were observed between the groups by 8 hours post-LPS stimulation. Treatment with A. scaber extract resulted in the highest levels of IkappaBalpha degradation. Our findings illustrate that the natural plant products P. japonica, H. cordata, and A. scaber may enhance immune function by modulating ex vivo pro-inflammatory cytokine and NO production as well as the expression of iNOS and COX-2.

  20. Human MAIT-cell responses to Escherichia coli: activation, cytokine production, proliferation, and cytotoxicity

    PubMed Central

    Dias, Joana; Sobkowiak, Michał J.; Sandberg, Johan K.; Leeansyah, Edwin

    2016-01-01

    Mucosa-associated invariant T cells are a large and relatively recently described innate-like antimicrobial T-cell subset in humans. These cells recognize riboflavin metabolites from a range of microbes presented by evolutionarily conserved major histocompatibility complex, class I-related molecules. Given the innate-like characteristics of mucosa-associated invariant T cells and the novel type of antigens they recognize, new methodology must be developed and existing methods refined to allow comprehensive studies of their role in human immune defense against microbial infection. In this study, we established protocols to examine a range of mucosa-associated invariant T-cell functions as they respond to antigen produced by Escherichia coli. These improved and dose- and time-optimized experimental protocols allow detailed studies of MR1-dependent mucosa-associated invariant T-cell responses to Escherichia coli pulsed antigen-presenting cells, as assessed by expression of activation markers and cytokines, by proliferation, and by induction of apoptosis and death in major histocompatibility complex, class I-related–expressing target cells. The novel and optimized protocols establish a framework of methods and open new possibilities to study mucosa-associated invariant T-cell immunobiology, using Escherichia coli as a model antigen. Furthermore, we propose that these robust experimental systems can also be adapted to study mucosa-associated invariant T-cell responses to other microbes and types of antigen-presenting cells. PMID:27034405

  1. Effect of silica and gold nanoparticles on macrophage proliferation, activation markers, cytokine production, and phagocytosis in vitro.

    PubMed

    Bancos, Simona; Stevens, David L; Tyner, Katherine M

    2015-01-01

    The accumulation of durable nanoparticles (NPs) in macrophages following systemic administration is well described. The ultimate biological impact of this accumulation on macrophage function, however, is not fully understood. In this study, nontoxic doses of two durable NPs, SiO2 and Au, at particle sizes of ~10 nm and 300 nm were used to evaluate the effect of bioaccumulation on macrophage function in vitro using RAW 264.7 mouse macrophage-like cells as a model system. Cell proliferation, cell cycle, cytokine production, surface marker activation, and phagocytosis responses were evaluated through a panel of assays using flow cytometry and confocal microscopy. The most dramatic change in RAW 264.7 cell function was a reduction in phagocytosis as monitored by the uptake of Escherichia coli. Cells exposed to both 10 nm Au NPs and 10 nm SiO2 NPs showed ~50% decrease in phagocytosis, while the larger NPs caused a less dramatic reduction. In addition to modifying phagocytosis profiles, 10 nm SiO2 NPs caused changes in proliferation, cell cycle, and cell morphology. Au NPs had no effect on cell cycle, cytokine production, or surface markers and caused interference in phagocytosis in the form of quenching when the assay was performed via flow cytometry. Confocal microscopy analysis was used to minimize this interference and demonstrated that both sizes of Au NPs decreased the phagocytosis of E. coli. Overall, our results demonstrate that Au and SiO2 NP uptake by macrophages can influence macrophage phagocytosis in vitro without altering surface markers and cytokine production in vitro. While the biological impact of these findings remains unclear, our results indicate that bioaccumulation of durable NPs within the macrophages may lead to a suppression of bacterial uptake and possibly impair bactericidal activity.

  2. Effect of silica and gold nanoparticles on macrophage proliferation, activation markers, cytokine production, and phagocytosis in vitro

    PubMed Central

    Bancos, Simona; Stevens, David L; Tyner, Katherine M

    2015-01-01

    The accumulation of durable nanoparticles (NPs) in macrophages following systemic administration is well described. The ultimate biological impact of this accumulation on macrophage function, however, is not fully understood. In this study, nontoxic doses of two durable NPs, SiO2 and Au, at particle sizes of ~10 nm and 300 nm were used to evaluate the effect of bioaccumulation on macrophage function in vitro using RAW 264.7 mouse macrophage-like cells as a model system. Cell proliferation, cell cycle, cytokine production, surface marker activation, and phagocytosis responses were evaluated through a panel of assays using flow cytometry and confocal microscopy. The most dramatic change in RAW 264.7 cell function was a reduction in phagocytosis as monitored by the uptake of Escherichia coli. Cells exposed to both 10 nm Au NPs and 10 nm SiO2 NPs showed ~50% decrease in phagocytosis, while the larger NPs caused a less dramatic reduction. In addition to modifying phagocytosis profiles, 10 nm SiO2 NPs caused changes in proliferation, cell cycle, and cell morphology. Au NPs had no effect on cell cycle, cytokine production, or surface markers and caused interference in phagocytosis in the form of quenching when the assay was performed via flow cytometry. Confocal microscopy analysis was used to minimize this interference and demonstrated that both sizes of Au NPs decreased the phagocytosis of E. coli. Overall, our results demonstrate that Au and SiO2 NP uptake by macrophages can influence macrophage phagocytosis in vitro without altering surface markers and cytokine production in vitro. While the biological impact of these findings remains unclear, our results indicate that bioaccumulation of durable NPs within the macrophages may lead to a suppression of bacterial uptake and possibly impair bactericidal activity. PMID:25565813

  3. Upregulated LINE-1 Activity in the Fanconi Anemia Cancer Susceptibility Syndrome Leads to Spontaneous Pro-inflammatory Cytokine Production.

    PubMed

    Brégnard, Christelle; Guerra, Jessica; Déjardin, Stéphanie; Passalacqua, Frank; Benkirane, Monsef; Laguette, Nadine

    2016-06-01

    Fanconi Anemia (FA) is a genetic disorder characterized by elevated cancer susceptibility and pro-inflammatory cytokine production. Using SLX4(FANCP) deficiency as a working model, we questioned the trigger for chronic inflammation in FA. We found that absence of SLX4 caused cytoplasmic DNA accumulation, including sequences deriving from active Long INterspersed Element-1 (LINE-1), triggering the cGAS-STING pathway to elicit interferon (IFN) expression. In agreement, absence of SLX4 leads to upregulated LINE-1 retrotransposition. Importantly, similar results were obtained with the FANCD2 upstream activator of SLX4. Furthermore, treatment of FA cells with the Tenofovir reverse transcriptase inhibitor (RTi), that prevents endogenous retrotransposition, decreased both accumulation of cytoplasmic DNA and pro-inflammatory signaling. Collectively, our data suggest a contribution of endogenous RT activities to the generation of immunogenic cytoplasmic nucleic acids responsible for inflammation in FA. The additional observation that RTi decreased pro-inflammatory cytokine production induced by DNA replication stress-inducing drugs further demonstrates the contribution of endogenous RTs to sustaining chronic inflammation. Altogether, our data open perspectives in the prevention of adverse effects of chronic inflammation in tumorigenesis.

  4. Inhibitory effects of diallyl disulfide on the production of inflammatory mediators and cytokines in lipopolysaccharide-activated BV2 microglia

    SciTech Connect

    Park, Hye Young; Kim, Nam Deuk; Kim, Gi-Young; Hwang, Hye Jin; Kim, Byung-Woo; Kim, Wun Jae; Choi, Yung Hyun

    2012-07-15

    Diallyl disulfide (DADS), a main organosulfur component responsible for the diverse biological effects of garlic, displays a wide variety of internal biological activities. However, the cellular and molecular mechanisms underlying DADS' anti-inflammatory activity remain poorly understood. In this study, therefore, the anti-inflammatory effects of DADS were studied to investigate its potential therapeutic effects in lipopolysaccharide (LPS)-stimulated BV2 microglia. We found that pretreatment with DADS prior to treatment with LPS significantly inhibited excessive production of nitric oxide (NO) and prostaglandin E{sub 2} (PGE{sub 2}) in a dose-dependent manner. The inhibition was associated with down-regulation of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) expression. DADS also attenuated the production of pro-inflammatory cytokines and chemokines, including interleukin-1β (IL-1β), tumor necrosis factor (TNF)-α, and monocyte chemoattractant protein-1 (MCP-1) by suppressing the expression of mRNAs for these proteins. The mechanism underlying this protective effect might be related to the inhibition of nuclear factor-kappaB, Akt and mitogen-activated protein kinase signaling pathway activation in LPS-stimulated microglial cells. These findings indicated that DADS is potentially a novel therapeutic candidate for the treatment of various neurodegenerative diseases. -- Highlights: ► DADS attenuates production of NO and PGE2 in LPS-activated BV2 microglia. ► DADS downregulates levels of iNOS and COX-2. ► DADS inhibits production and expression of inflammatory cytokines and chemokine. ► DADS exhibits these effects by suppression of NF-κB, PI3K/Akt and MAPKs pathways.

  5. Enhanced natural killer activity and production of pro-inflammatory cytokines in mice selected for high acute inflammatory response (AIRmax).

    PubMed

    Castoldi, Lindsey; Golim, Marjorie Assis; Filho, Orlando Garcia Ribeiro; Romagnoli, Graziela Gorete; Ibañez, Olga Célia Martinez; Kaneno, Ramon

    2007-03-01

    Strains of mice with maximal and minimal acute inflammatory responsiveness (AIRmax and AIRmin, respectively) were developed through selective breeding based on their high- or low-acute inflammatory responsiveness. Previous reports have shown that AIRmax mice are more resistant to the development of a variety of tumours than AIRmin mice, including spontaneous metastasis of murine melanoma. Natural killer activity is involved in immunosurveillance against tumour development, so we analysed the number and activity of natural killer cells (CD49b(+)), T-lymphocyte subsets and in vitro cytokine production by spleen cells of normal AIRmax and AIRmin mice. Analysis of lymphocyte subsets by flow cytometry showed that AIRmax mice had a higher relative number of CD49b(+) cells than AIRmin mice, as well as cytolytic activity against Yac.1 target cells. The number of CD3(+) CD8(+) cells was also higher in AIRmax mice. These findings were associated with the ability of spleen cells from AIRmax mice in vitro to produce higher levels of the pro-inflammatory cytokines tumour necrosis factor-alpha, interleukin-12p40 and interferon-gamma but not the anti-inflammatory interleukin-10. Taken together, our data suggest that the selective breeding to achieve the AIRmax and AIRmin strains was able to polarize the genes associated with cytotoxic activity, which can be responsible for the antitumour resistance observed in AIRmax mice.

  6. Store-operated CRAC channels regulate PAR2-activated Ca2+ signaling and cytokine production in airway epithelial cells

    PubMed Central

    Jairaman, Amit; Yamashita, Megumi; Schleimer, Robert P.; Prakriya, Murali

    2015-01-01

    The G-protein coupled protease-activated receptor 2 (PAR2) plays an important role in the pathogenesis of various inflammatory and auto-immune disorders. In airway epithelial cells (AECs), stimulation of PAR2 by allergens and proteases triggers the release of a host of inflammatory mediators to regulate bronchomotor tone and immune cell recruitment. Activation of PAR2 turns on several cell signaling pathways of which the mobilization of cytosolic Ca2+ is likely a critical but poorly understood event. Here, we show that Ca2+ release-activated Ca2+ (CRAC) channels encoded by STIM1 and Orai1 are a major route of Ca2+ entry in primary human AECs and drive the Ca2+ elevations seen in response to PAR2 activation. Activation of CRAC channels induces the production of several key inflammatory mediators from AECs including TSLP, IL-6 and PGE2, in part through stimulation of gene expression via NFAT (nuclear factor of activated T-cells). Furthermore, PAR2 stimulation induces the production of many key inflammatory mediators including PGE2, IL-6, IL-8 and GM-CSF in a CRAC-channel dependent manner. These findings indicate that CRAC channels are the primary mechanism for Ca2+ influx in AECs and a vital checkpoint for the induction of PAR2-induced proinflammatory cytokines. PMID:26238490

  7. Velutin reduces lipopolysaccharide-induced proinflammatory cytokine TNFa and IL-6 production by inhibiting NF-Kappa B activation

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Recent studies have shown that some flavonoids are modulators of proinflammatory cytokine expression. Velutin, an uncommon flavone isolated from acai (Euterpe oleraceas) berry, was tested for the effects in reducing LPS-induced TNFa and IL-6 production in RAW 264.7 peripheral macrophages and periton...

  8. Pretreatment of activated human CD8 T cells with IL-12 leads to enhanced TCR-induced signaling and cytokine production.

    PubMed

    Vacaflores, Aldo; Freedman, Samantha N; Chapman, Nicole M; Houtman, Jon C D

    2017-01-01

    During the immune response to pathogens and autoantigens, CD8T cells are exposed to numerous inflammatory agents including the cytokine IL-12. Previous studies have focused on how IL-12 regulates T cell functions when present during or after the activation of the T cell receptor (TCR). However, recent studies suggest that prior exposure to IL-12 also alters the TCR responsiveness of murine T cells. Whether similar phenomena occur in human activated CD8T cells and the mechanisms mediating these effects remain unexplored. In this study, we observed that pretreatment of human activated CD8T cells with IL-12 results in increased cytokine mRNA and protein production following subsequent TCR challenge. The potentiation of TCR-mediated cytokine release was transient and required low doses of IL-12 for at least 24h. Mechanistically, prior exposure to IL-12 increased the TCR induced activation of select MAPKs and AKT without altering the activation of more proximal TCR signaling molecules, suggesting that the IL-12 mediated changes in TCR signaling are responsible for the increased production of cytokines. Our data suggest that prior treatment with IL-12 potentiates human CD8T cell responses at sites of infection and inflammation, expanding our understanding of the function of this clinically important cytokine.

  9. GRB2 Nucleates T Cell Receptor-Mediated LAT Clusters That Control PLC-γ1 Activation and Cytokine Production

    PubMed Central

    Bilal, Mahmood Yousif; Houtman, Jon C. D.

    2015-01-01

    GRB2 is a ubiquitously expressed adaptor protein required for signaling downstream of multiple receptors. To address the role of GRB2 in receptor-mediated signaling, the expression of GRB2 was suppressed in human CD4+ T cells and its role downstream of the T cell receptor (TCR) was examined. Interestingly, GRB2 deficient T cells had enhanced signaling from complexes containing the TCR. However, GRB2 deficient T cells had substantially reduced production of IL-2 and IFN-γ. This defect was attributed to diminished formation of linker for activation of T cells (LAT) signaling clusters, which resulted in reduced MAP kinase activation, calcium flux, and PLC-γ1 recruitment to LAT signaling clusters. Add back of wild-type GRB2, but not a novel N-terminal SH3 domain mutant, rescued LAT microcluster formation, calcium mobilization, and cytokine release, providing the first direct evidence that GRB2, and its ability to bind to SH3 domain ligands, is required for establishing LAT microclusters. Our data demonstrate that the ability of GRB2 to facilitate protein clusters is equally important in regulating TCR-mediated functions as its capacity to recruit effector proteins. This highlights that GRB2 regulates signaling downstream of adaptors and receptors by both recruiting effector proteins and regulating the formation of signaling complexes. PMID:25870599

  10. Impact of short-term systemic hypoxia on phagocytosis, cytokine production, and transcription factor activation in peripheral blood cells.

    PubMed

    Fritzenwanger, Michael; Jung, Christian; Goebel, Bjoern; Lauten, Alexander; Figulla, Hans R

    2011-01-01

    Hypoxia frequently associated with certain physiologic and pathologic conditions influences numerous cellular functions. Because the effects of short-term hypoxia are incompletely understood, we examined phagocytosis and cytokine production as well as the activation of the transcription factors HIF-1 and NFκB in peripheral blood cells of healthy volunteers exposed to an oxygen concentration equivalent to that found at a height of 5500 m. Furthermore, we analysed plasma HIF-1 and serum concentrations of various HIF-1-dependent genes. Results showed that short-term hypoxia increased phagocytosis in neutrophils without affecting monocyte phagocytosis. Hypoxia decreased basal TNFα concentration in monocytes and basal interferon γ concentration in CD4(+) T lymphocytes. In contrast, plasma HIF and serum VEGF concentrations were not affected by hypoxia, although serum EPO concentration was raised. In PBMC, hypoxia increased cytosolic HIF-1 concentration without affecting nuclear HIF-1 concentration and led to a rise in the nuclear NFκB in PBMC. Our results show that short-term hypoxia affects immune functions in healthy individuals. Furthermore, we speculate that the effects of hypoxia are not due to HIF-1, but are caused by the activation of NFκB .

  11. GRB2 Nucleates T Cell Receptor-Mediated LAT Clusters That Control PLC-γ1 Activation and Cytokine Production.

    PubMed

    Bilal, Mahmood Yousif; Houtman, Jon C D

    2015-01-01

    GRB2 is a ubiquitously expressed adaptor protein required for signaling downstream of multiple receptors. To address the role of GRB2 in receptor-mediated signaling, the expression of GRB2 was suppressed in human CD4+ T cells and its role downstream of the T cell receptor (TCR) was examined. Interestingly, GRB2 deficient T cells had enhanced signaling from complexes containing the TCR. However, GRB2 deficient T cells had substantially reduced production of IL-2 and IFN-γ. This defect was attributed to diminished formation of linker for activation of T cells (LAT) signaling clusters, which resulted in reduced MAP kinase activation, calcium flux, and PLC-γ1 recruitment to LAT signaling clusters. Add back of wild-type GRB2, but not a novel N-terminal SH3 domain mutant, rescued LAT microcluster formation, calcium mobilization, and cytokine release, providing the first direct evidence that GRB2, and its ability to bind to SH3 domain ligands, is required for establishing LAT microclusters. Our data demonstrate that the ability of GRB2 to facilitate protein clusters is equally important in regulating TCR-mediated functions as its capacity to recruit effector proteins. This highlights that GRB2 regulates signaling downstream of adaptors and receptors by both recruiting effector proteins and regulating the formation of signaling complexes.

  12. Activity of Uncleaved Caspase-8 Controls Anti-bacterial Immune Defense and TLR-Induced Cytokine Production Independent of Cell Death

    PubMed Central

    DeLaney, Alexandra; Santos-Marrero, Melanie; Grier, Jennifer T.; Sun, Yan; Zwack, Erin E.; Hu, Baofeng; Olsen, Tayla M.; Rongvaux, Anthony; López, Carolina B.; Oberst, Andrew; Beiting, Daniel P.; Brodsky, Igor E.

    2016-01-01

    Caspases regulate cell death programs in response to environmental stresses, including infection and inflammation, and are therefore critical for the proper operation of the mammalian immune system. Caspase-8 is necessary for optimal production of inflammatory cytokines and host defense against infection by multiple pathogens including Yersinia, but whether this is due to death of infected cells or an intrinsic role of caspase-8 in TLR-induced gene expression is unknown. Caspase-8 activation at death signaling complexes results in its autoprocessing and subsequent cleavage and activation of its downstream apoptotic targets. Whether caspase-8 activity is also important for inflammatory gene expression during bacterial infection has not been investigated. Here, we report that caspase-8 plays an essential cell-intrinsic role in innate inflammatory cytokine production in vivo during Yersinia infection. Unexpectedly, we found that caspase-8 enzymatic activity regulates gene expression in response to bacterial infection as well as TLR signaling independently of apoptosis. Using newly-generated mice in which caspase-8 autoprocessing is ablated (Casp8DA/DA), we now demonstrate that caspase-8 enzymatic activity, but not autoprocessing, mediates induction of inflammatory cytokines by bacterial infection and a wide variety of TLR stimuli. Because unprocessed caspase-8 functions in an enzymatic complex with its homolog cFLIP, our findings implicate the caspase-8/cFLIP heterodimer in control of inflammatory cytokines during microbial infection, and provide new insight into regulation of antibacterial immune defense. PMID:27737018

  13. Mitochondrial proteins NIP-SNAP-1 and -2 are a target for the immunomodulatory activity of clarithromycin, which involves NF-κB-mediated cytokine production.

    PubMed

    Yamamoto, Soh; Ogasawara, Noriko; Yamamoto, Keisuke; Uemura, Chika; Takaya, Yoshiaki; Shiraishi, Tsukasa; Sato, Toyotaka; Hashimoto, Shin; Tsutsumi, Hiroyuki; Takano, Kenichi; Himi, Tetsuo; Yokota, Shin-Ichi

    2017-02-12

    Macrolide antibiotics have immunomodulatory activities, including suppression of cytokine production, cell adhesion molecule expression, and mucin production. These immunomodulatory activities improve the symptoms of respiratory diseases associated with chronic inflammation. However, the underlying molecular mechanism(s) is not well understood yet. To address this, we prepared clarithromycin (CAM)-conjugated Sepharose and examined bound cellular proteins by proteome analysis. We identified mitochondrial proteins 4-nitrophenylphosphatase domain and non-neuronal synaptosomal associated protein 25-like protein homolog (NIP-SNAP)-1 and -2 and very long-chain acyl-CoA dehydrogenase (VLCAD) as CAM-binding proteins. Production of proinflammatory cytokines (IL-8 and IL-6) induced by lipopolysaccharides (LPSs) and Pam3-CSK4 in human epithelial cell lines BEAS-2B and T24 were suppressed by knockdown of NIP-SNAP-1 or -2, and partly by knockdown of VLCAD. Also, knockdown of NIP-SNAP-1 or -2 in various cell lines suppressed LPS-induced expression of IL-8 and IL-6 mRNA and NF-κB activity. Thus, CAM suppresses NF-κB-mediated proinflammatory cytokine production by interacting with mitochondrial proteins, NIP-SNAP-1 and -2.

  14. Activation of p38 MAPK by feline infectious peritonitis virus regulates pro-inflammatory cytokine production in primary blood-derived feline mononuclear cells.

    PubMed

    Regan, Andrew D; Cohen, Rebecca D; Whittaker, Gary R

    2009-02-05

    Feline infectious peritonitis (FIP) is an invariably fatal disease of cats caused by systemic infection with a feline coronavirus (FCoV) termed feline infectious peritonitis virus (FIPV). The lethal pathology associated with FIP (granulomatous inflammation and T-cell lymphopenia) is thought to be mediated by aberrant modulation of the immune system due to infection of cells such as monocytes and macrophages. Overproduction of pro-inflammatory cytokines occurs in cats with FIP, and has been suggested to play a significant role in the disease process. However, the mechanism underlying this process remains unknown. Here we show that infection of primary blood-derived feline mononuclear cells by FIPV WSU 79-1146 and FIPV-DF2 leads to rapid activation of the p38 MAPK pathway and that this activation regulates production of the pro-inflammatory cytokine tumor necrosis factor alpha (TNF-alpha) and interleukin-1 beta (IL-1 beta). FIPV-induced p38 MAPK activation and pro-inflammatory cytokine production was inhibited by the pyridinyl imidazole inhibitors SB 203580 and SC 409 in a dose-dependent manner. FIPV-induced p38 MAPK activation was observed in primary feline blood-derived mononuclear cells individually purified from multiple SPF cats, as was the inhibition of TNF-alpha production by pyridinyl imidazole inhibitors.

  15. Depressant effects of ambroxol and erdosteine on cytokine synthesis, granule enzyme release, and free radical production in rat alveolar macrophages activated by lipopolysaccharide.

    PubMed

    Jang, Yoon Young; Song, Jin Ho; Shin, Yong Kyoo; Han, Eun Sook; Lee, Chung Soo

    2003-04-01

    The present study examined the effects of ambroxol and erdosteine, bronchial expectorants, on the cytokine synthesis, granule enzyme release, and free radical production in rat alveolar macrophages activated by lipopolysaccharide. Ambroxol and erdosteine significantly decreased the production of tumour necrosis factors-alpha, interleukin-1beta, and interleukin-6 in alveolar macrophages activated by lipopolysaccharide. These drugs significantly reduced the production of superoxide anion, hydrogen peroxide, and nitric oxide and the release of acid phosphatase and lysozyme in lipopolysaccharide-activated macrophages. Ambroxol and erdosteine showed no scavenging effect on superoxide anion and hydrogen peroxide, whereas both drugs effectively decomposed nitric oxide. The results show that ambroxol and erdosteine may inhibit the responses, including cytokine synthesis and free radical production, in rat alveolar macrophages activated by lipopolysaccharide. Unlike the production of reactive oxygen species, the inhibitory effect of ambroxol and erdosteine on the production of nitric oxide in lipopolysaccharide-activated alveolar macrophages may be accomplished by a scavenging action on the species and inhibition of the respiratory burst.

  16. Immune Response to Snake Envenoming and Treatment with Antivenom; Complement Activation, Cytokine Production and Mast Cell Degranulation

    PubMed Central

    Stone, Shelley F.; Isbister, Geoffrey K.; Shahmy, Seyed; Mohamed, Fahim; Abeysinghe, Chandana; Karunathilake, Harendra; Ariaratnam, Ariaranee; Jacoby-Alner, Tamara E.; Cotterell, Claire L.; Brown, Simon G. A.

    2013-01-01

    Background Snake bite is one of the most neglected public health issues in poor rural communities worldwide. In addition to the clinical effects of envenoming, treatment with antivenom frequently causes serious adverse reactions, including hypersensitivity reactions (including anaphylaxis) and pyrogenic reactions. We aimed to investigate the immune responses to Sri Lankan snake envenoming (predominantly by Russell's viper) and antivenom treatment. Methodology/Principal Findings Plasma concentrations of Interleukin (IL)-6, IL-10, tumor necrosis factor α (TNFα), soluble TNF receptor I (sTNFRI), anaphylatoxins (C3a, C4a, C5a; markers of complement activation), mast cell tryptase (MCT), and histamine were measured in 120 Sri Lankan snakebite victims, both before and after treatment with antivenom. Immune mediator concentrations were correlated with envenoming features and the severity of antivenom-induced reactions including anaphylaxis. Envenoming was associated with complement activation and increased cytokine concentrations prior to antivenom administration, which correlated with non-specific systemic symptoms of envenoming but not with coagulopathy or neurotoxicity. Typical hypersensitivity reactions to antivenom occurred in 77/120 patients (64%), satisfying criteria for a diagnosis of anaphylaxis in 57/120 (48%). Pyrogenic reactions were observed in 32/120 patients (27%). All patients had further elevations in cytokine concentrations, but not complement activation, after the administration of antivenom, whether a reaction was noted to occur or not. Patients with anaphylaxis had significantly elevated concentrations of MCT and histamine. Conclusions/Significance We have demonstrated that Sri Lankan snake envenoming is characterized by significant complement activation and release of inflammatory mediators. Antivenom treatment further enhances the release of inflammatory mediators in all patients, with anaphylactic reactions characterised by high levels of mast

  17. Cytokine profile of murine malaria: stage-related production of inflammatory and anti-inflammatory cytokines.

    PubMed

    Bakir, Hanaa Y; Tomiyama, Chikako; Abo, Toru

    2011-06-01

    Balance between inflammatory and anti-inflammatory cytokines may be important in malaria presentation and outcome. To clarify cytokine interactions that produce pathology of malaria and control infection, C57BL/6 mice were infected with 10(4) parasitized RBCs from a non-lethal strain of Plasmodium yoelii. Kinetics was monitored showing the course of parasitemia, and cytokines were determined by RT-PCR from liver and spleen tissues. Inflammatory cytokines such as interferon-γ (IFNγ), interleukin (IL)-12, IL-6, tumor necrosis factor-α (TNFα) and anti-inflammatory cytokines, including IL-4 and IL-10, were investigated as key molecules that interact with immune cells in the activation of the immune responses. The production of IFNγ mRNA was found to be higher on day 7 than on day 21 after infection, and IL-12 and IL-6 showed higher expression in the liver than in the spleen. Though TNFα was highly expressed on day 14 after infection and on day 21 in the liver, such expression was decreased on day 21 in the spleen. Anti-inflammatory cytokines showed high expression in both the liver and spleen. The results suggest that a relative balance between inflammatory and anti-inflammatory cytokines is crucial and that the increase of inflammatory cytokine levels during the acute phase of malaria may reflect an early and effective immune response.The counteraction effect of anti-inflammatory cytokines is thought to play a role in limiting progression from uncomplicated malaria to severe life-threatening complications.

  18. Dysregulation of in vitro cytokine production by monocytes during sepsis.

    PubMed Central

    Munoz, C; Carlet, J; Fitting, C; Misset, B; Blériot, J P; Cavaillon, J M

    1991-01-01

    The production by monocytes of interleukin-1 alpha (IL-1 alpha), interleukin-1 beta (IL-1 beta), interleukin-6 (IL-6), and tumor necrosis factor alpha (TNF alpha) in intensive care unit (ICU) patients with sepsis syndrome (n = 23) or noninfectious shock (n = 6) is reported. Plasma cytokines, cell-associated cytokines within freshly isolated monocytes and LPS-induced in vitro cytokine production were assessed at admission and at regular intervals during ICU stay. TNF alpha and IL-6 were the most frequently detected circulating cytokines. Despite the fact that IL-1 alpha is the main cytokine found within monocytes upon in vitro activation of cells from healthy individuals, it was very rarely detected within freshly isolated monocytes from septic patients, and levels of cell-associated IL-1 beta were lower than those of TNF alpha. Cell-associated IL-1 beta and TNF alpha were not correlated with corresponding levels in plasma. Upon LPS stimulation, we observed a profound decrease of in vitro IL-1 alpha production by monocytes in all patients, and of IL-1 beta, IL-6, and TNF alpha in septic patients. This reduced LPS-induced production of cytokines was most pronounced in patients with gram-negative infections. Finally, monocytes from survival patients, but not from nonsurvival ones recovered their capacity to produce normal amounts of cytokines upon LPS stimulation. In conclusion, our data indicate an in vivo activation of circulating monocytes during sepsis as well as in noninfectious shock and suggest that complex regulatory mechanisms can downregulate the production of cytokines by monocytes during severe infections. Images PMID:1939659

  19. Cytokine and nitric oxide production following severe envenomation.

    PubMed

    Petricevich, Vera L

    2004-09-01

    Venom is a complex mixture of many substances such as toxins, enzymes, growth factor activators, and inhibitors are particularly responsible for the deleterious effects of cells. These constituents interact in the body with a large number of proteins and receptors, and this interaction determines the eventual inflammatory effect of the compounds. Envenomation by bees, scorpions, snakes, spiders and wasps involves the activation of the inflammatory response with the release and activation of pro-inflammatory cytokines and other mediators, such as nitric oxide. Recently, a battery of cytokines produced by activated T cells or macrophages have been added to in envenomations. Cytokines are important for the interactions between cells in the immune and inflammatory responses. Although the pathophysiology of envenomation is not fully understood, venom and immune responses are known to trigger the release of cytokines and nitric oxide. The cytokines initiate a cascade of events that lead to illness behaviors such as fever, anorexia, and, as well as a host of physiologic events such as activation of vasodilation, hypotension and increased nitric oxide production. Accumulating evidence indicates that these cytokines play important roles in mediating cell recruitment and activation necessary for inflammation and the repair of tissue damage. A better understanding of the involvement of the inflammatory system in different envenoming syndromes may have future therapeutic benefits.

  20. Cytokine Regulation Immunoglobulin Isotype Production

    DTIC Science & Technology

    1994-11-08

    FerRIll - Fc receptor for IgG , type III GAF interferon-gamma activated factor G AS interferon-gamma activation site G(lM~ - goat anti-mouse IgD GM...present. The prevalence of IgA at mucous membranes effects a first line of immunologic defense against invading pathogens and reinforces the physical...demonstrated following several distinct types of immunization. Small increases In serum IgG2a are seen in mice injected with goat anti-mouse IgD antibody

  1. Effect of Blood Component Coatings of Enosseal Implants on Proliferation and Synthetic Activity of Human Osteoblasts and Cytokine Production of Peripheral Blood Mononuclear Cells

    PubMed Central

    Hulejova, Hana; Bartova, Jirina; Riedel, Tomas; Pesakova, Vlasta

    2016-01-01

    The study monitored in vitro early response of connective tissue cells and immunocompetent cells to enosseal implant materials coated by different blood components (serum, activated plasma, and plasma/platelets) to evaluate human osteoblast proliferation and synthetic activity and inflammatory response presented as a cytokine profile of peripheral blood mononuclear cells (PBMCs) under conditions imitating the situation upon implantation. The cells were cultivated on coated Ti-plasma-sprayed (Ti-PS), Ti-etched (Ti-Etch), Ti-hydroxyapatite (Ti-HA), and ZrO2 surfaces. The plasma/platelets coating supported osteoblast proliferation only on osteoconductive Ti-HA and Ti-Etch whereas activated plasma enhanced proliferation on all surfaces. Differentiation (BAP) and IL-8 production remained unchanged or decreased irrespective of the coating and surface; only the serum and plasma/platelets-coated ZrO2 exhibited higher BAP and IL-8 expression. RANKL production increased on serum and activated plasma coatings. PBMCs produced especially cytokines playing role in inflammatory phase of wound healing, that is, IL-6, GRO-α, GRO, ENA-78, IL-8, GM-CSF, EGF, and MCP-1. Cytokine profiles were comparable for all tested surfaces; only ENA-78, IL-8, GM-CSF, and MCP-1 expression depended on materials and coatings. The activated plasma coating led to uniformed surfaces and represented a favorable treatment especially for bioinert Ti-PS and ZrO2 whereas all coatings had no distinctive effect on bioactive Ti-HA and Ti-Etch. PMID:27651560

  2. Decreased Gaq expression in T cells correlates with enhanced cytokine production and disease activity in systemic lupus erythematosus

    PubMed Central

    Luo, Jiao; Yu, Bing; Qian, Hongyan; Duan, Lihua; Shi, Guixiu

    2016-01-01

    Aberrant T cell immune responses appear central to the development of systemic lupus erythematosus (SLE). We previously reported that Gαq, the alpha subunit of Gq, regulates T and B cell immune responses, promoting autoimmunity. To address whether Gαq contributes to the pathogenesis of SLE, Gαq mRNA expression was studied using real time-PCR in PBMCs and T cells from SLE patients as well as age- and sex-matched healthy controls. Our results showed that Gαq mRNA expression was decreased in PBMCs and T cells from SLE patients compared to healthy individuals. Correlation analyses showed that Gαq expression in T cells from SLE patients was associated with disease severity (as per SLE Disease Activity Index), the presence of lupus nephritis, and expression of Th1, Th2 and Th17 cytokines. In keeping with clinical results, T-helper cell subsets (Th1, Th2 and Th17) were over-represented in Gαq knockout mice. In addition, Gαq expression in SLE T cells was negatively correlated with the expression of Bcl-2, an anti-apoptotic gene, and positively correlated with the expression of Bax, a pro-apoptotic gene. These data suggest that reduced Gαq levels in T cells may promote enhanced and prolonged T cell activation, contributing to the clinical manifestations of SLE. PMID:27965465

  3. Dendritic Cell Activation and Cytokine Production Induced by Group B Neisseria meningitidis: Interleukin-12 Production Depends on Lipopolysaccharide Expression in Intact Bacteria

    PubMed Central

    Dixon, Garth L. J.; Newton, Phillippa J.; Chain, Benjamin M.; Katz, David; Andersen, Svein Rune; Wong, Simon; van der Ley, Peter; Klein, Nigel; Callard, Robin E.

    2001-01-01

    Interactions between dendritic cells (DCs) and microbial pathogens are fundamental to the generation of innate and adaptive immune responses. Upon stimulation with bacteria or bacterial components such as lipopolysaccharide (LPS), immature DCs undergo a maturation process that involves expression of costimulatory molecules, HLA molecules, and cytokines and chemokines, thus providing critical signals for lymphocyte development and differentiation. In this study, we investigated the response of in vitro-generated human DCs to a serogroup B strain of Neisseria meningitidis compared to an isogenic mutant lpxA strain totally deficient in LPS and purified LPS from the same strain. We show that the parent strain, lpxA mutant, and meningococcal LPS all induce DC maturation as measured by increased surface expression of costimulatory molecules and HLA class I and II molecules. Both the parent and lpxA strains induced production of tumor necrosis factor alpha (TNF-α), interleukin-1α (IL-1α), and IL-6 in DCs, although the parent was the more potent stimulus. In contrast, high-level IL-12 production was only seen with the parent strain. Compared to intact bacteria, purified LPS was a very poor inducer of IL-1α, IL-6, and TNF-α production and induced no detectable IL-12. Addition of exogenous LPS to the lpxA strain only partially restored cytokine production and did not restore IL-12 production. These data show that non-LPS components of N. meningitidis induce DC maturation, but that LPS in the context of the intact bacterium is required for high-level cytokine production, especially that of IL-12. These findings may be useful in assessing components of N. meningitidis as potential vaccine candidates. PMID:11401973

  4. Dendritic cell activation and cytokine production induced by group B Neisseria meningitidis: interleukin-12 production depends on lipopolysaccharide expression in intact bacteria.

    PubMed

    Dixon, G L; Newton, P J; Chain, B M; Katz, D; Andersen, S R; Wong, S; van der Ley, P; Klein, N; Callard, R E

    2001-07-01

    Interactions between dendritic cells (DCs) and microbial pathogens are fundamental to the generation of innate and adaptive immune responses. Upon stimulation with bacteria or bacterial components such as lipopolysaccharide (LPS), immature DCs undergo a maturation process that involves expression of costimulatory molecules, HLA molecules, and cytokines and chemokines, thus providing critical signals for lymphocyte development and differentiation. In this study, we investigated the response of in vitro-generated human DCs to a serogroup B strain of Neisseria meningitidis compared to an isogenic mutant lpxA strain totally deficient in LPS and purified LPS from the same strain. We show that the parent strain, lpxA mutant, and meningococcal LPS all induce DC maturation as measured by increased surface expression of costimulatory molecules and HLA class I and II molecules. Both the parent and lpxA strains induced production of tumor necrosis factor alpha (TNF-alpha), interleukin-1alpha (IL-1alpha), and IL-6 in DCs, although the parent was the more potent stimulus. In contrast, high-level IL-12 production was only seen with the parent strain. Compared to intact bacteria, purified LPS was a very poor inducer of IL-1alpha, IL-6, and TNF-alpha production and induced no detectable IL-12. Addition of exogenous LPS to the lpxA strain only partially restored cytokine production and did not restore IL-12 production. These data show that non-LPS components of N. meningitidis induce DC maturation, but that LPS in the context of the intact bacterium is required for high-level cytokine production, especially that of IL-12. These findings may be useful in assessing components of N. meningitidis as potential vaccine candidates.

  5. Fenoterol inhibits LPS-induced AMPK activation and inflammatory cytokine production through β-arrestin-2 in THP-1 cell line

    SciTech Connect

    Wang, Wei; Zhang, Yuan; Xu, Ming; Zhang, You-Yi; He, Bei

    2015-06-26

    The AMP-activated protein kinase (AMPK) pathway is involved in regulating inflammation in several cell lines. We reported that fenoterol, a β{sub 2}-adrenergic receptor (β{sub 2}-AR) agonist, had anti-inflammatory effects in THP-1 cells, a monocytic cell line. Whether the fenoterol anti-inflammatory effect involves the AMPK pathway is unknown. In this study, we explored the mechanism of β{sub 2}-AR stimulation with fenoterol in a lipopolysaccharide (LPS)-induced inflammatory cytokine secretion in THP-1 cells. We studied whether fenoterol and β-arrestin-2 or AMPKα1 subunit knockdown could affect LPS-induced AMPK activation, nuclear factor-kappa B (NF-κB) activation and inflammatory cytokine secretion. LPS-induced AMPK activation and interleukin 1β (IL-1β) release were reduced with fenoterol pretreatment of THP-1 cells. SiRNA knockdown of β-arrestin-2 abolished the fenoterol inhibition of LPS-induced AMPK activation and interleukin 1β (IL-1β) release, thus β-arrestin-2 mediated the anti-inflammatory effects of fenoterol on LPS-treated THP-1 cells. In addition, siRNA knockdown of AMPKα1 significantly attenuated the LPS-induced NF-κB activation and IL-1β release, so AMPKα1 was a key signaling molecule involved in LPS-induced inflammatory cytokine production. These results suggested the β{sub 2}-AR agonist fenoterol inhibited LPS-induced AMPK activation and IL-1β release via β-arrestin-2 in THP-1 cells. The exploration of these mechanisms may help optimize therapeutic agents targeting these pathways in inflammatory diseases. - Highlights: • β{sub 2}-AR agonist fenoterol exerts its protective effect on LPS-treated THP-1 cells. • Fenoterol inhibits LPS-induced AMPK activation and IL-1β production. • β-arrestin2 mediates fenoterol-inhibited AMPK activation and IL-1β release. • AMPKα1 is involved in LPS-induced NF-κB activation and IL-1β production.

  6. Vinpocetine inhibits amyloid-beta induced activation of NF-κB, NLRP3 inflammasome and cytokine production in retinal pigment epithelial cells

    PubMed Central

    Liu, Ruozhou Tom; Wang, Aikun; To, Eleanor; Gao, Jiangyuan; Cao, Sijia; Cui, Jing Z.; Matsubara, Joanne A.

    2015-01-01

    Chronic inflammation is a key pathogenic process in age-related macular degeneration (AMD). Amyloid-beta (Aβ) is a constituent of AMD drusen and promotes the activation of NLRP3 inflammasome which facilitates the production of cytokines. We investigated the role of transcription factor NF-κB in the activation of inflammasome in the RPE and the effect of vinpocetine, a dietary supplement with inhibitory effect on NF-κB. ARPE19/NF-κB-luciferase reporter cells treated with Aβ demonstrated enhanced NF-κB activation that was significantly suppressed by vinpocetine. Intraperitoneal injection of vinpocetine (15 mg/kg) inhibited NF-κB nuclear translocation and reduced the expression and activation of NLRP3, caspase-1, IL-1β, IL-18, and TNF-α in the RPE of adult rats that received intraocular Aβ, as measured by retinal immunohistochemistry and Western blot. Cytokine level in the vitreous was assayed using multiplex suspension arrays and revealed significantly lower concentration of MIP-3α, IL-6, IL-1α, IL-1β, IL-18, and TNF-α in vinpocetine treated animals. These results suggest that the NF-κB pathway is activated by Aβ in the RPE and signals the priming of NLRP3 inflammasome and the expression of pro-inflammatory cytokines including the inflammasome substrates IL-1β and IL-18. NF-κB inhibition may be an effective approach to stem the chronic inflammatory milieu that underlies the development of AMD. Vinpocetine is a potentially useful anti-inflammatory agent that is well-tolerated in long term use. PMID:25041941

  7. Activation of KCNN3/SK3/K(Ca)2.3 channels attenuates enhanced calcium influx and inflammatory cytokine production in activated microglia.

    PubMed

    Dolga, Amalia M; Letsche, Till; Gold, Maike; Doti, Nunzianna; Bacher, Michael; Chiamvimonvat, Nipavan; Dodel, Richard; Culmsee, Carsten

    2012-12-01

    In neurons, small-conductance calcium-activated potassium (KCNN/SK/K(Ca)2) channels maintain calcium homeostasis after N-methyl-D-aspartate (NMDA) receptor activation, thereby preventing excitotoxic neuronal death. So far, little is known about the function of KCNN/SK/K(Ca)2 channels in non-neuronal cells, such as microglial cells. In this study, we addressed the question whether KCNN/SK/K(Ca)2 channels activation affected inflammatory responses of primary mouse microglial cells upon lipopolysaccharide (LPS) stimulation. We found that N-cyclohexyl-N-[2-(3,5-dimethyl-pyrazol-1-yl)-6-methyl-4-pyrimidinamine (CyPPA), a positive pharmacological activator of KCNN/SK/K(Ca)2 channels, significantly reduced LPS-stimulated activation of microglia in a concentration-dependent manner. The general KCNN/SK/K(Ca)2 channel blocker apamin reverted these effects of CyPPA on microglial proliferation. Since calcium plays a central role in microglial activation, we further addressed whether KCNN/SK/K(Ca)2 channel activation affected the changes of intracellular calcium levels, [Ca(2+)](i), in microglial cells. Our data show that LPS-induced elevation of [Ca(2+)](i) was attenuated following activation of KCNN2/3/K(Ca)2.2/K(Ca)2.3 channels by CyPPA. Furthermore, CyPPA reduced downstream events including tumor necrosis factor alpha and interleukin 6 cytokine production and nitric oxide release in activated microglia. Further, we applied specific peptide inhibitors of the KCNN/SK/K(Ca)2 channel subtypes to identify which particular channel subtype mediated the observed anti-inflammatory effects. Only inhibitory peptides targeting KCNN3/SK3/K(Ca)2.3 channels, but not KCNN2/SK2/K(Ca)2.2 channel inhibition, reversed the CyPPA-effects on LPS-induced microglial proliferation. These findings revealed that KCNN3/SK3/K(Ca)2.3 channels can modulate the LPS-induced inflammatory responses in microglial cells. Thus, KCNN3/SK3/K(Ca)2.3 channels may serve as a therapeutic target for reducing microglial

  8. Flexible cytokine production by macrophages and T cells in response to probiotic bacteria: a possible mechanism by which probiotics exert multifunctional immune regulatory activities.

    PubMed

    Shida, Kan; Nanno, Masanobu; Nagata, Satoru

    2011-01-01

    Probiotics have been reported to be efficacious against cancers, infections, allergies, inflammatory bowel diseases and autoimmune diseases, and it is important to explain how such multifunctional activities are realized. Lactobacillus casei Shirota (LcS) is one of these multifunctional probiotics, and its ability to augment the host immune system has been extensively examined. We have shown that the cell wall structure of this probiotic strain is responsible for potently inducing IL-12 production. In addition, we have recently found that LcS differentially controls the inflammatory cytokine responses of macrophages and T cells in either Peyer's patches or the spleen. Other studies revealed that LcS-induced IL-12 production by macrophages is modified when other bacteria or their cell components are simultaneously present. These findings can provide a theoretical basis for understanding the multifunctional activities of specific probiotics.

  9. Transcription Factors Regulating Inflammatory Cytokine Production Are Differentially Expressed in Peripheral Blood Mononuclear Cells of Behçet Disease Depending on Disease Activity

    PubMed Central

    Woo, Min-Yeong; Yun, Su Jin; Lee, Mi Jin; Kim, Kyongmin

    2017-01-01

    Background Behçet disease (BD) is a relapsing inflammatory disease with increased production of inflammatory cytokines in peripheral blood mononuclear cells (PBMCs); however, the underlying molecular mechanisms are not well known. Objective To analyze whether the differential expression of transcription factors is involved in the increased tumor necrosis factor (TNF)-α and interleukin (IL)-6 production by PBMCs of BD patients compared to healthy controls (HCs). Methods Expression of transcription factors was examined by real-time reverse transcriptase-polymerase chain reaction and western blotting. Cytokine production by CD11b+ cells transfected with siRNAs against transcription factors was measured by enzyme-linked immunosorbent assay. Results In the absence of lipopolysaccharide stimulation, the transcript level of CCAAT-enhancer-binding proteins (C/EBP) β was increased in PBMCs from patients with active BD compared to that in PBMCs from patients with stable BD. The C/EBPδ transcript level was higher in PBMCs from patients with active BD than in those from HCs. The activating transcription factor 3 (ATF3) transcript level was increased in PBMCs from patients with stable BD compared to that in PBMCs from HCs. siRNAs targeting C/EBPβ and C/EBPδ significantly reduced the production of IL-6 and TNF-α in lipopolysaccharide-stimulated CD11b+ cells from patients with BD as well as from HCs. Conclusion We found differential expression of C/EBPβ, C/EBPδ, and ATF3 in PBMCs from patients with BD depending on disease activity, indicating the involvement of these molecules in BD pathogenesis.

  10. Resistin Enhances Inflammatory Cytokine Production in Coronary Artery Tissues by Activating the NF-κB Signaling

    PubMed Central

    Gao, Fang; Si, Feifei; Feng, Siqi; Liu, Ruixi

    2016-01-01

    Purpose. Kawasaki disease (KD) is a systemic vasculitis and is a leading cause of coronary artery lesions (CALs) in childhood. Our previous study has shown higher levels of serum Resistin in KD patients with coronary aneurysm. This study aimed at examining the association of Resistin with inflammatory cytokine expression in mouse model of coronary arteritis and determining the potential mechanisms. Methods. C57BL/6 mice were injected with Lactobacillus cell wall extract (LCWE) to induce coronary arteritis. The relative levels of Resistin, TNF-α, IL-1β, and MMP-9 expression and inflammatory infiltrates in the coronary arteries were determined longitudinally by quantitative RT-PCR, ELISA, and histology. The effect of TLR4 and NF-κB activation on Resistin-induced TNF-α and IL-1β expression in human coronary artery endothelium cells (HCAECs) was examined by quantitative RT-PCR. Results. Higher levels of Resistin, TNF-α, IL-1β, and MMP-9 expression were associated with the degrees of inflammatory infiltrates in the coronary artery walls of the LCWE-injected mice. Resistin enhanced TNF-α and IL-1β expression in HCAECs at 18 or 24 hours after stimulation. Pretreatment with anti-TLR4 attenuated Resistin-enhanced IL-1β, but not TNF-α, expression and pretreatment with parthenolide or QNZ demolished Resistin-enhanced TNF-α expression in HACECs. Pretreatment with parthenolide, but not QNZ, blocked Resistin-enhanced IL-1β expression in HCAECs. Conclusion. Resistin may enhance inflammation by cross-talking with TLR4/NF-κB signaling during the development of coronary arteritis in mice. PMID:27800490

  11. Oxygen-glucose deprivation inducing B1 RNA inhibits neuronal cells metabolic activity by NLRP3 and associated proinflammatory cytokines production.

    PubMed

    Wang, Li; Wang, Weihua; Zhang, Lei; Dai, Peng; Wang, Kai; Hui, Hao; Rao, Wei; Peng, Cheng; Yang, Jinghua; Yan, Zhen; Fei, Zhou

    2015-02-19

    Cerebral ischemia occurs when blood flow to part of the brain is obstructed, which can result in oxygen and glucose deprivation (OGD) and neuronal damage. However, the mechanisms remain poorly understood. The present study investigated the production and effects of double-stranded RNA (dsRNA) induced by OGD in neuronal cells. By confocal microscopy, dsRNA containing B1 and B2 RNA, was found accumulating in HT22 cells under OGD treatment. The sequence of B1 RNA was identified and transfected into HT22 cells. Interestingly, B1 RNA induced transcription and expression of NLRP3, interleukin (IL)-1β, and tumor necrosis factor (TNF)-α, which was similar to the effects of OGD treatment. Moreover, HT22 cell growth inhibition and proinflammatory cytokines production induced by OGD and B1 RNA treatment were down-regulated by NLRP3 knock-down. These findings suggest that B1 RNA induced by OGD forms as dsRNA and inhibits neuronal cell metabolic activity by regulating the NLRP3 and associated proinflammatory cytokines production.

  12. Activation of PPARγ inhibits pro-inflammatory cytokines production by upregulation of miR-124 in vitro and in vivo.

    PubMed

    Wang, Dan; Shi, Liuyan; Xin, Wei; Xu, Jiancheng; Xu, Jing; Li, Qi; Xu, Zhi; Wang, Jianchun; Wang, Guansong; Yao, Wei; He, Binfeng; Yang, Yu; Hu, Mingdong

    2017-03-22

    Peroxisome proliferator-activated receptor gamma (PPARγ) and miR-124 have been reported to play important roles in regulation of inflammation. However, the underlying anti-inflammatory mechanisms remain not well understood. In the present study, we demonstrated that the expression level of PPARγ is positively correlated with that of miR-124 in patients with sepsis. Activation of PPARγ upregulates miR-124 and in turn inhibits miR-124 target gene. PPARγ bound directly to PPRE in the miR-124 promoter region, and enhanced the promoter transcriptional activity. PPARγ-induced miR-124 is involved in the suppression of pro-inflammatory cytokine in vitro and in vivo. These results suggest that PPARγ-induced miR-124 inhibits the production of pro-inflammatory cytokines is a novel PPARγ anti-inflammatory mechanism and also indicate that miR-124 may be a potential therapeutic target for the treatment of inflammatory diseases.

  13. Cytokine crowdsourcing: multicellular production of TH17-associated cytokines.

    PubMed

    Busman-Sahay, Kathleen O; Walrath, Travis; Huber, Samuel; O'Connor, William

    2015-03-01

    In the 2 decades since its discovery, IL-17A has become appreciated for mounting robust, protective responses against bacterial and fungal pathogens. When improperly regulated, however, IL-17A can play a profoundly pathogenic role in perpetuating inflammation and has been linked to a wide variety of debilitating diseases. IL-17A is often present in a composite milieu that includes cytokines produced by TH17 cells (i.e., IL-17F, IL-21, IL-22, and IL-26) or associated with other T cell lineages (e.g., IFN-γ). These combinatorial effects add mechanistic complexity and more importantly, contribute differentially to disease outcome. Whereas TH17 cells are among the best-understood cell types that secrete IL-17A, they are frequently neither the earliest nor dominant producers. Indeed, non-TH17 cell sources of IL-17A can dramatically alter the course and severity of inflammatory episodes. The dissection of the temporal regulation of TH17-associated cytokines and the resulting net signaling outcomes will be critical toward understanding the increasingly intricate role of IL-17A and TH17-associated cytokines in disease, informing our therapeutic decisions. Herein, we discuss important non-TH17 cell sources of IL-17A and other TH17-associated cytokines relevant to inflammatory events in mucosal tissues.

  14. PSM Peptides of Staphylococcus aureus Activate the p38-CREB Pathway in Dendritic Cells, Thereby Modulating Cytokine Production and T Cell Priming.

    PubMed

    Armbruster, Nicole S; Richardson, Jennifer R; Schreiner, Jens; Klenk, Juliane; Günter, Manina; Kretschmer, Dorothee; Pöschel, Simone; Schenke-Layland, Katja; Kalbacher, Hubert; Clark, Kristopher; Autenrieth, Stella E

    2016-02-01

    The challenging human pathogen Staphylococcus aureus has highly efficient immune evasion strategies for causing a wide range of diseases, from skin and soft tissue to life-threatening infections. Phenol-soluble modulin (PSM) peptides are major pathogenicity factors of community-associated methicillin-resistant S. aureus strains. In previous work, we demonstrated that PSMs in combination with TLR2 ligand from S. aureus induce tolerogenic dendritic cells (DCs) characterized by the production of high amounts of IL-10, but no proinflammatory cytokines. This in turn promotes the activation of regulatory T cells while impairing Th1 response; however, the signaling pathways modulated by PSMs remain elusive. In this study, we analyzed the effects of PSMs on signaling pathway modulation downstream of TLR2. TLR2 stimulation in combination with PSMα3 led to increased and prolonged phosphorylation of NF-κB, ERK, p38, and CREB in mouse bone marrow-derived DCs compared with single TLR2 activation. Furthermore, inhibition of p38 and downstream MSK1 prevented IL-10 production, which in turn reduced the capacity of DCs to activate regulatory T cells. Interestingly, the modulation of the signaling pathways by PSMs was independent of the known receptor for PSMs, as shown by experiments with DCs lacking the formyl peptide receptor 2. Instead, PSMs penetrate the cell membrane most likely by transient pore formation. Moreover, colocalization of PSMs and p38 was observed near the plasma membrane in the cytosol, indicating a direct interaction. Thus, PSMs from S. aureus directly modulate the signaling pathway p38-CREB in DCs, thereby impairing cytokine production and in consequence T cell priming to increase the tolerance toward the pathogen.

  15. Cytokine production capacity in depression and anxiety

    PubMed Central

    Vogelzangs, N; de Jonge, P; Smit, J H; Bahn, S; Penninx, B W

    2016-01-01

    Recent studies have suggested that immune function may be dysregulated in persons with depressive and anxiety disorders. Few studies examined the expression of cytokines in response to ex vivo stimulation of blood by lipopolysaccharide (LPS) to study the innate production capacity of cytokines in depression and anxiety. To investigate this, baseline data from the Netherlands Study of Depression and Anxiety (NESDA) were used, including persons (18–65 years; 66% women) with current (that is, past month; N=591) or remitted (N=354) DSM-IV depressive or anxiety disorders and healthy controls (N=297). Depressive and anxiety symptoms were measured by means of the Inventory of Depressive Symptomatology (IDS) and the Beck Anxiety Inventory (BAI). Using Multi-Analyte Profiling technology, plasma levels of 13 cytokines were assayed after whole blood stimulation by addition of LPS. Basal plasma levels of C-reactive protein, interleukin-6 and tumor necrosis factor-α were also available. A basal and a LPS summary index were created. Results show that LPS-stimulated inflammation was associated with increased odds of current depressive/anxiety disorders (odds ratio (OR)=1.28, P=0.009), as was the case for basal inflammation (OR=1.28, P=0.001). These associations were no longer significant after adjustment for lifestyle and health (OR=1.13, P=0.21; OR=1.07, P=0.45, respectively). After adjustment for lifestyle and health, interleukin-8 was associated with both remitted (OR=1.25, P=0.02) and current (OR=1.28, P=0.005) disorders. In addition, LPS-stimulated inflammation was associated with more severe depressive (β=0.129, P<0.001) and anxiety (β=0.165, P<0.001) symptoms, as was basal inflammation. Unlike basal inflammation, LPS-stimulated inflammation was still associated with (anxiety) symptom severity after adjustment for lifestyle and health (IDS: IL-8, MCP-1, MMP2; BAI: LPS index, IL-6, IL-8, IL-10, IL-18, MCP-1, MMP2, TNF-β). To conclude, lifestyle and health factors may

  16. Resveratrol Suppresses Cytokine Production Linked to FcεRI-MAPK Activation in IgE-Antigen Complex-Exposed Basophilic Mast Cells and Mice.

    PubMed

    Han, Seon-Young; Choi, Yean-Jung; Kang, Min-Kyung; Park, Jung Han Yoon; Kang, Young-Hee

    2015-01-01

    A complicated interplay between resident mast cells and other recruited inflammatory cells contributes to the development and progression of allergic inflammation entailing the promotion of T helper 2 (Th2) cytokine responses. The current study examined whether resveratrol suppressed the production of inflammatory Th2 cytokines in cultured rat basophilic leukemia RBL-2H3 cells. Cells pre-treated with resveratrol nontoxic at 1–25 μM were sensitized with anti-dinitrophenyl (anti-DNP), and subsequently stimulated by dinitrophenyl-human serum albumin (DNP–HSA) antigen. Resveratrol dose-dependently diminished the secretion of interleukin (IL)-3, IL-4, IL-13 as well as tumor necrosis factor (TNF)-α by the antigen stimulation from sensitized cells. It was found that resveratrol mitigated the phosphorylation of p38 MAPK, ERK, and JNK elevated in mast cells exposed to Fc epsilon receptor I (FcεRI)-mediated immunoglobulin E (IgE)-antigen complex. The FcεRI aggregation was highly enhanced on the surface of mast cells following the HSA stimulation, which was retarded by treatment with 1–25 μM resveratrol. The IgE-receptor engagement rapidly induced tyrosine phosphorylation of c-Src-related focal adhesion protein paxillin involved in the cytoskeleton rearrangement. The FcεRI-mediated rapid activation of c-Src and paxillin was attenuated in a dose-dependent manner. In addition, the paxillin activation entailed p38 MAPK and ERK-responsive signaling, but the JNK activation was less involved. Consistently, oral administration of resveratrol reduced the tissue level of phosphorylated paxillin in the dorsal skin of DNP–HSA-challenged mice. The other tyrosine kinase Tyk2-STAT1 signaling was activated in the dorsal epidermis of antigen-exposed mice, which was associated with allergic inflammation. These results showed that resveratrol inhibited Th2 cytokines- and paxillin-linked allergic responses dependent upon MAPK signaling. Therefore, resveratrol may possess the

  17. Mechanisms Underlying the Anti-Inflammatory Effects of Clinacanthus nutans Lindau Extracts: Inhibition of Cytokine Production and Toll-Like Receptor-4 Activation.

    PubMed

    Mai, Chun W; Yap, Kok S I; Kho, Mee T; Ismail, Nor H; Yusoff, Khatijah; Shaari, Khozirah; Chin, Swee Y; Lim, Erin S H

    2016-01-01

    Clinacanthus nutans has had a long history of use in folk medicine in Malaysia and Southeast Asia; mostly in the relief of inflammatory conditions. In this study, we investigated the effects of different extracts of C. nutans upon lipopolysaccharide (LPS) induced inflammation in order to identify its mechanism of action. Extracts of leaves and stem bark of C. nutans were prepared using polar and non-polar solvents to produce four extracts, namely polar leaf extract (LP), non-polar leaf extract (LN), polar stem extract (SP), and non-polar stem extracts (SN). The extracts were standardized by determining its total phenolic and total flavonoid contents. Its anti-inflammatory effects were assessed on LPS induced nitrite release in RAW264.7 macrophages and Toll-like receptor (TLR-4) activation in TLR-4 transfected human embryonic kidney cells (HEK-Blue(TM)-hTLR4 cells). The levels of inflammatory cytokines (TNF-α, IFN-γ, IL-1β, IL-6, IL-12p40, and IL-17) in treated RAW264.7 macrophages were quantified to verify its anti-inflammatory effects. Western blotting was used to investigate the effect of the most potent extract (LP) on TLR-4 related inflammatory proteins (p65, p38, ERK, JNK, IRF3) in RAW264.7 macrophages. All four extracts produced a significant, concentration-dependent reduction in LPS-stimulated nitric oxide, LPS-induced TLR-4 activation in HEK-Blue(TM)-hTLR4 cells and LPS-stimulated cytokines production in RAW264.7 macrophages. The most potent extract, LP, also inhibited all LPS-induced TLR-4 inflammatory proteins. These results provide a basis for understanding the mechanisms underlying the previously demonstrated anti-inflammatory activity of C. nutans extracts.

  18. Mechanisms Underlying the Anti-Inflammatory Effects of Clinacanthus nutans Lindau Extracts: Inhibition of Cytokine Production and Toll-Like Receptor-4 Activation

    PubMed Central

    Mai, Chun W.; Yap, Kok S. I.; Kho, Mee T.; Ismail, Nor H.; Yusoff, Khatijah; Shaari, Khozirah; Chin, Swee Y.; Lim, Erin S. H.

    2016-01-01

    Clinacanthus nutans has had a long history of use in folk medicine in Malaysia and Southeast Asia; mostly in the relief of inflammatory conditions. In this study, we investigated the effects of different extracts of C. nutans upon lipopolysaccharide (LPS) induced inflammation in order to identify its mechanism of action. Extracts of leaves and stem bark of C. nutans were prepared using polar and non-polar solvents to produce four extracts, namely polar leaf extract (LP), non-polar leaf extract (LN), polar stem extract (SP), and non-polar stem extracts (SN). The extracts were standardized by determining its total phenolic and total flavonoid contents. Its anti-inflammatory effects were assessed on LPS induced nitrite release in RAW264.7 macrophages and Toll-like receptor (TLR-4) activation in TLR-4 transfected human embryonic kidney cells (HEK-BlueTM-hTLR4 cells). The levels of inflammatory cytokines (TNF-α, IFN-γ, IL-1β, IL-6, IL-12p40, and IL-17) in treated RAW264.7 macrophages were quantified to verify its anti-inflammatory effects. Western blotting was used to investigate the effect of the most potent extract (LP) on TLR-4 related inflammatory proteins (p65, p38, ERK, JNK, IRF3) in RAW264.7 macrophages. All four extracts produced a significant, concentration-dependent reduction in LPS-stimulated nitric oxide, LPS-induced TLR-4 activation in HEK-BlueTM-hTLR4 cells and LPS-stimulated cytokines production in RAW264.7 macrophages. The most potent extract, LP, also inhibited all LPS-induced TLR-4 inflammatory proteins. These results provide a basis for understanding the mechanisms underlying the previously demonstrated anti-inflammatory activity of C. nutans extracts. PMID:26869924

  19. Kinetics and functional implications of Th1 and Th2 cytokine production following activation of peripheral blood mononuclear cells in primary culture.

    PubMed

    McHugh, S; Deighton, J; Rifkin, I; Ewan, P

    1996-06-01

    The importance of cytokine production in some disease processes is now widely recognized. To investigate temporal relationships between cytokines, we stimulated peripheral blood mononuclear cells (PBMC) in vitro using the T cell mitogen phytohemagglutinin (PHA) and various antigens chosen to induce predominantly Th1 (streptokinase: streptodornase or purified protein derivative) or Th2 (Dermatophagoides pteronyssinus, bee or wasp venom: allergens in sensitive subjects) responses. Cytokine production was measured by sensitive bioassays or enzyme-linked immunosorbent assays. Of the 30 subjects studied, 10 were normal and 20 individuals were allergic to either D. pteronyssinus (n = 10) or bee venom (n = 10) (examined before specific allergen immunotherapy). We examined the temporal profiles of a panel of cytokines produced in primary culture. In PHA-driven cultures, cytokines were found to be sequentially produced in the order interleukin (IL)-2, IL-4, IL-5, IL-3, interferon (IFN)-gamma, IL-10, IL-6, IL-12 and tumor necrosis factor (TNF)-alpha. The response to allergen in allergic patients was predominantly Th2 in nature, with the production of IL-4, IL-5, IL-6 and IL-10, but little or no IFN-gamma. IL-2, IL-3, TNF-alpha and IL-12 were also produced in low amounts. The response of both atopic and normal subjects to recall bacterial antigens was predominantly Th1, with high levels of IFN-gamma, IL-2 and TNF-alpha. The relevance of the order, amount and speed of production, characteristic kinetics (production, consumption, homeostatic regulation) and the cell source of the cytokines are discussed.

  20. Protective effect of gedunin on TLR-mediated inflammation by modulation of inflammasome activation and cytokine production: Evidence of a multitarget compound.

    PubMed

    Borges, Perla Villani; Moret, Katelim Hottz; Raghavendra, Nulgumnalli Manjunathaiah; Maramaldo Costa, Thadeu Estevam; Monteiro, Ana Paula; Carneiro, Alan Brito; Pacheco, Patrícia; Temerozo, Jairo Ramos; Bou-Habib, Dumith Chequer; das Graças Henriques, Maria; Penido, Carmen

    2017-01-01

    Activation of toll-like receptors (TLRs) by pathogen-associated molecular patterns (PAMPs) triggers an innate immune response, via cytokine production and inflammasome activation. Herein, we have investigated the modulatory effect of the natural limonoid gedunin on TLR activation in vitro and in vivo. Intraperitoneal (i.p.) pre- and post-treatments of C57BL/6 mouse with gedunin impaired the influx of mononuclear cells, eosinophils and neutrophils, as well as the production of tumor necrosis factor (TNF)-α, interleukin (IL)-6 and nitric oxide (NO), triggered by lipopolysaccharide (LPS) in mouse pleura. Accordingly, in vitro post-treatment of immortalized murine macrophages with gedunin also impaired LPS-induced production of such mediators. Gedunin diminished LPS-induced expression of the nucleotide-binding domain and leucine-rich repeat protein-3 (NLRP3) on pleural leukocytes in vivo and in immortalized macrophages in vitro. In line with this, gedunin inhibited LPS-induced caspase-1 activation and the production of IL-1β in vivo and in vitro. In addition, gedunin treatment triggered the generation of the anti-inflammatory factors IL-10 and heme oxigenase-1 (HO-1) at resting conditions or upon stimulation. We also demonstrate that gedunin effect is not restricted to TLR4-mediated response, since this compound diminished TNF-α, IL-6, NO, NLRP3 and IL-1β, as well as enhanced IL-10 and HO-1, by macrophages stimulated with the TLR2 and TLR3 agonists, palmitoyl-3-Cys-Ser-(Lys)4 (PAM3) and polyriboinosinic:polyribocytidylic acid (POLY I:C), in vitro. In silico modeling studies revealed that gedunin efficiently docked into caspase-1, TLR2, TLR3 and to the myeloid differentiation protein-2 (MD-2) component of TLR4. Overall, our data demonstrate that gedunin modulates TLR4, TLR3 and TLR2-mediated responses and reveal new molecular targets for this compound.

  1. Green and black tea inhibit cytokine-induced IL-8 production and secretion in AGS gastric cancer cells via inhibition of NF-κB activity.

    PubMed

    Gutierrez-Orozco, Fabiola; Stephens, Brian R; Neilson, Andrew P; Green, Rodney; Ferruzzi, Mario G; Bomser, Joshua A

    2010-10-01

    Consumption of tea is associated with a reduced risk for several gastrointestinal cancers. Inflammatory processes, such as secretion of IL-8 from the gastric epithelium in response to chronic chemokine or antigen exposure, serve both as a chemoattractant for white blood cells and a prerequisite for gastric carcinogenesis. In this study, the gastric adenocarcinoma cell line AGS was used to investigate the effect of green tea extract, black tea extract, and epigallocatechin gallate (EGCG), the most abundant catechin in tea, on cytokine-induced inflammation. AGS cells were stimulated with interleukin-1β (IL-1β) to initiate inflammation, followed by exposure to either tea extracts or EGCG. We found that both green and black tea extracts at concentrations of 20 and 2 µM total catechins, respectively, significantly (p < 0.05) inhibited IL-1β-induced IL-8 production and secretion to a similar extent. Treatment of AGS cells with EGCG (8 µM) produced similar reductions in IL-1β-induced IL-8 production and secretion. Inhibition of NF-κB activity was found to be responsible, in part, for these observed effects. Our findings demonstrate that both green and black tea extracts with distinctly different catechin profiles, are capable of disrupting the molecular link between inflammation and carcinogenesis via inhibition of NF-κB activity in AGS cells.

  2. B cells responses and cytokine production are regulated by their immune microenvironment.

    PubMed

    Vazquez, Monica I; Catalan-Dibene, Jovani; Zlotnik, Albert

    2015-08-01

    The adaptive immune system consists of two types of lymphocytes: T and B cells. These two lymphocytes originate from a common precursor, yet are fundamentally different with B cells mediating humoral immunity while T cells mediate cell mediated immunity. In cytokine production, naïve T cells produce multiple cytokines upon activation while naïve activated B cells do not. B cells are capable of producing cytokines, but their cytokine production depends on their differentiation state and activation conditions. Hence, unlike T cells that can produce a large amount of cytokines upon activation, B cells require specific differentiation and activation conditions to produce cytokines. Many cytokines act on B cells as well. Here, we discuss several cytokines and their effects on B cells including: Interleukins, IL-7, IL-4, IL-6, IL-10, and Interferons, IFN-α, IFN-β, IFN-γ. These cytokines play important roles in the development, survival, differentiation and/or proliferation of B cells. Certain chemokines also play important roles in B cell function, namely antibody production. As an example, we discuss CCL28, a chemokine that directs the migration of plasma cells to mucosal sites. We conclude with a brief overview of B cells as cytokine producers and their likely functional consequences on the immune response.

  3. Blueberries reduce pro-inflammatory cytokine TNF-alpha and IL-6 production in mouse macrophages by inhibiting NF Kappa B activation and the MAPK pathway

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Blueberries (BB) have been reported to attenuate atherosclerosis in apoE deficient (ApoE-/-) mice. The aim of this study was to evaluate the effects of BB in reducing pro-inflammatory cytokine production in mouse macrophages. ApoE-/- mice were fed AIN-93G diet (CD) or CD formulated to contain 1% fre...

  4. In vitro exposure to the herbicide atrazine inhibits T cell activation, proliferation, and cytokine production and significantly increases the frequency of Foxp3+ regulatory T cells.

    PubMed

    Thueson, Lindsay E; Emmons, Tiffany R; Browning, Dianna L; Kreitinger, Joanna M; Shepherd, David M; Wetzel, Scott A

    2015-02-01

    The herbicide atrazine (2-chloro-4-[ethylamino]-6-[isopropylamino]-s-triazine) is the most common water contaminant in the United States. Atrazine is a phosphodiesterase inhibitor and is classified as an estrogen disrupting compound because it elevates estrogen levels via induction of the enzyme aromatase. Previous studies have shown that atrazine exposure alters the function of innate immune cells such as NK cells, DC, mast cells, and macrophages. In this study we have examined the impact of in vitro atrazine exposure on the activation, proliferation, and effector cytokine production by primary murine CD4(+) T lymphocytes. We found that atrazine exposure significantly inhibited CD4(+) T cell proliferation and accumulation as well as the expression of the activation markers CD25 and CD69 in a dose-dependent manner. Interestingly, the effects were more pronounced in cells from male animals. These effects were partially mimicked by pharmacological reagents that elevate intracellular cAMP levels and addition of exogenous rmIL-2 further inhibited proliferation and CD25 expression. Consistent with these findings, atrazine exposure during T cell activation resulted in a 2- to 5-fold increase in the frequency of Foxp3(+) CD4(+) T cells.

  5. Leishmania donovani-specific 25- and 28-kDa urinary proteins activate macrophage effector functions, lymphocyte proliferation and Th1 cytokines production.

    PubMed

    Kumar, Vinod; Gour, Jalaj K; Singh, Nisha; Bajpai, Surabhi; Singh, Rakesh K

    2013-04-01

    Growing incidence of drug resistance against leishmaniasis in endemic areas and limited drug options necessitates the need for a vaccine. Notwithstanding significant leishmanial research in the past decades, a vaccine candidate is far from reality. In this study, we report the potential of two urinary leishmanial proteins to induce macrophage effector functions, inflammatory cytokines production and human lymphocytes proliferation. A total four proteins of molecular mass 25, 28, 54 and 60 kDa were identified in human urine samples. The 25 and 28 kDa proteins significantly induced NADPH oxidase (p<0.001), superoxide dismutase (p<0.001) and inducible nitric oxide synthase (p<0.001) activities in stimulated RAW264.7 macrophages. The release of nitric oxide, tumor necrosis factor-alpha and interleukin (IL)-12 was also significantly (p<0.001) higher in 25 and 28 kDa activated macrophages as compared with cells activated with other two proteins. These two proteins also induced significant (p<0.001) proliferation and release of IFN-γ and IL-12 in human peripheral blood mononuclear cells.

  6. The FcepsilonRIbeta immunoreceptor tyrosine-based activation motif exerts inhibitory control on MAPK and IkappaB kinase phosphorylation and mast cell cytokine production.

    PubMed

    Furumoto, Yasuko; Nunomura, Satoshi; Terada, Tomoyoshi; Rivera, Juan; Ra, Chisei

    2004-11-19

    The high affinity IgE Fc receptor (FcepsilonRI) beta chain functions as a signal amplifier and has been linked to atopy, asthma, and allergy. Herein, we report on a previously unrecognized negative regulatory role for the nonconventional beta chain immunoreceptor tyrosine-based activation motif that contains three tyrosine residues (YX5YX3Y). Degranulation and leukotriene production was found to be impaired in cells expressing the mutated FcepsilonRIbeta immunoreceptor tyrosine-based activation motifs FYY, YYF, FYF, and FFF. In contrast, cytokine synthesis and secretion were enhanced in the YFY and FFF mutants. FcepsilonRI phosphorylation and Lyn kinase co-immunoprecipitation was intact in the YFY mutant but was lost in the FYF and FFF mutants. The phosphorylation of Syk, LAT, phospholipase gamma1/2, and Srchomology 2 domain-containing protein phosphatase 2 was intact, whereas the phosphorylation of SHIP-1 was significantly reduced in the YFY mutant cells. The FYF and FFF mutants were defective in phosphorylating all of these molecules. In contrast, the phosphorylation of ERK, p38 MAPK, IkappaB kinase beta (IKKbeta), and nuclear NFkappaB activity was enhanced in the YFY and FFF mutants. These findings show that the FcepsilonRIbeta functions to both selectively amplify (degranulation and leukotriene secretion) and dampen (lymphokine) mast cell effector responses.

  7. Effect of Buthus martensi Karsch on aromatase activity and cytokine-inducted NOS and NO production in osteoblasts and leukaemic cell line FLG 29.1.

    PubMed

    Jin, Un-Ho; Kim, Kap-Sung; Park, Su-Yeon; Chung, Kang-Hyun; Kim, Dong-Soo; Chang, Young-Chae; Kim, Cheorl-Ho

    2006-01-01

    Among the different scorpion species, Buthus martensi Karsch, a widely distributed scorpion species in Asia especially in Korea, has received a lot of attention. Indeed, over the past decade, more than 70 different peptides, toxins, or homologues have been isolated. It may prove a valuable resource for identifying potential anti-inflammatory and analgesic drugs. The recent observation has suggested that the aromatase is a possible local modulator of bone remodeling in osteoarthritis and osteoporosis. In the present study, therefore, the effect of Buthus martensi Karsch (BMK) extract, traditional immunosuppressive Korean aqua-acupuncture water, on the bone function of human osteoblastic cells was studied. To provide insights into the effect of BMK on aromatase activity in bone-derived cells, we examined the human leukaemic cell line FLG 29.1, which is induced to differentiate toward the osteoclastic phenotype by 12-O-tetradecanoyl-phorbol-13-acetate (TPA) and transforming growth factor (TGF)-beta1, and the primary first-passage osteoblastic cells (hOB). Gene expression of the aromatase was not affected by Buthus martensi Karsch in FLG 29.1 and hOB cells. However, enzyme activity was stimulated in a time-dependent fashion by 10.0 microg/ml BMK and by either 1-50 nM TPA or 0.01-0.5 ng/ml TGF-beta1, with maximal responses after 2-3 hr exposure. On the other hand, BMK strongly inhibited interleukin-1beta (IL-1beta)- and tumor necrosis factor (TNF)alpha-induced Nitricoxide (NO) synthase expression with little effect on constitutive NO synthase expression. BMK extracts (10 microg/ml) inhibited cytokine-induced iNOS and nNOS expression. BMK (10 microg/ml) did not affect the ecNOS expression, indicating the extracts are not working on the constitutive NOS expression. BMK strongly inhibited the cytokine-induced NO production (p < 0.01). BMK also showed significant inhibition on NO production in both induced by TNF-alpha+IL-1beta. NO donors, sodium nitroprusside, and NONOate

  8. KSRP: A Checkpoint for Inflammatory Cytokine Production in Astrocytes

    PubMed Central

    LI, XUELIN; LIN, WEI-JYE; CHEN, CHING-YI; SI, YING; ZHANG, XIAOWEN; LU, LIANG; SUSWAM, ESTHER; ZHENG, LEI; KING, PETER H.

    2013-01-01

    Chronic inflammation in the central nervous system (CNS) is a central feature of many neurodegenerative and autoimmune diseases. As an immunologically competent cell, the astrocyte plays an important role in CNS inflammation. It is capable of expressing a number of cytokines such as tumor necrosis factor alpha (TNF-α) and interleukin-1 beta (IL-1β) that promote inflammation directly and through the recruitment of immune cells. Checkpoints are therefore in place to keep tight control over cytokine production. Adenylate/uridylate-rich elements (ARE) in the 3′ untranslated region of cytokine mRNAs serve as a major checkpoint by regulating mRNA stability and translational efficiency. Here, we examined the impact of KH-type splicing regulatory protein (KSRP), an RNA binding protein which destabilizes mRNAs via the ARE, on cytokine expression and paracrine phenotypes of primary astrocytes. We identified a network of inflammatory mediators, including TNF-α and IL-1β, whose expression increased 2 to 4-fold at the RNA level in astrocytes isolated from KSRP−/− mice compared to littermate controls. Upon activation, KSRP−/− astrocytes produced TNF-α and IL-1β at levels that exceeded control cells by 15-fold or more. Conditioned media from KSRP−/− astrocytes induced chemotaxis and neuronal cell death in vitro. Surprisingly, we observed a prolongation of half-life in only a subset of mRNA targets and only after selective astrocyte activation. Luciferase reporter studies indicated that KSRP regulates cytokine gene expression at both transcriptional and post-transcriptional levels. Our results outline a critical role for KSRP in regulating pro-inflammatory mediators and have implications for a wide range of CNS inflammatory and autoimmune diseases. PMID:22847996

  9. Vinpocetine Reduces Carrageenan-Induced Inflammatory Hyperalgesia in Mice by Inhibiting Oxidative Stress, Cytokine Production and NF-κB Activation in the Paw and Spinal Cord

    PubMed Central

    Ruiz-Miyazawa, Kenji W.; Zarpelon, Ana C.; Pinho-Ribeiro, Felipe A.; Pavão-de-Souza, Gabriela F.; Casagrande, Rubia; Verri, Waldiceu A.

    2015-01-01

    Vinpocetine is a safe nootropic agent used for neurological and cerebrovascular diseases. The anti-inflammatory activity of vinpocetine has been shown in cell based assays and animal models, leading to suggestions as to its utility in analgesia. However, the mechanisms regarding its efficacy in inflammatory pain treatment are still not completely understood. Herein, the analgesic effect of vinpocetine and its anti-inflammatory and antioxidant mechanisms were addressed in murine inflammatory pain models. Firstly, we investigated the protective effects of vinpocetine in overt pain-like behavior induced by acetic acid, phenyl-p-benzoquinone (PBQ) and formalin. The intraplantar injection of carrageenan was then used to induce inflammatory hyperalgesia. Mechanical and thermal hyperalgesia were evaluated using the electronic von Frey and the hot plate tests, respectively, with neutrophil recruitment to the paw assessed by a myeloperoxidase activity assay. A number of factors were assessed, both peripherally and in the spinal cord, including: antioxidant capacity, reduced glutathione (GSH) levels, superoxide anion, tumor necrosis factor alpha (TNF-α) and interleukin 1 beta (IL-1β) levels, as well as nuclear factor kappa B (NF-κB) activation. Vinpocetine inhibited the overt pain-like behavior induced by acetic acid, PBQ and formalin (at both phases), as well as the carrageenan-induced mechanical and thermal hyperalgesia and associated neutrophil recruitment. Both peripherally and in the spinal cord, vinpocetine also inhibited: antioxidant capacity and GSH depletion; increased superoxide anion; IL-1β and TNF-α levels; and NF-κB activation. As such, vinpocetine significantly reduces inflammatory pain by targeting oxidative stress, cytokine production and NF-κB activation at both peripheral and spinal cord levels. PMID:25822523

  10. Etomidate Mitigates Lipopolysaccharide-Induced CD14 and TREM-1 Expression, NF-κB Activation, and Pro-inflammatory Cytokine Production in Rat Macrophages.

    PubMed

    Liu, Ming; Zhang, Yu; Xiong, Jun-Yu; Wang, Yan; Lv, Shen

    2016-02-01

    This study was aimed at investigating the effect of etomidate on the viability of rat macrophages and the function of lipopolysaccharide (LPS)-stimulated macrophages as well as the potential mechanisms. Rat macrophages were isolated and treated with different doses of etomidate for 24 h, and their viability was determined by the CCK-8 assay. Furthermore, macrophages were treated with, or without, 1 μg/ml of LPS, and/or 2.5 or 5 μM etomidate in the presence or absence of a TREM-1 inhibitor (LP17, 100 ng/ml), and the levels of TNF-α, IL-6, CD14, and TREM-1 in the different groups of cells were determined by quantitative RT-PCR, ELISA, and Western blot assays. The levels of NF-κB activation in the different groups of cells were analyzed by an electrophoretic mobility shift assay (EMSA). Etomidate at 31.25 μM or a low dose did not affect the viability of rat macrophages, while etomidate at higher doses reduced the viability of macrophages in vitro. Treatment with 2.5 or 5 μM etomidate or with LP17 alone did not affect the levels of TNF-α, IL-6, CD-14, and TREM-1 in macrophages. Treatment with etomidate significantly mitigated LPS-stimulated TNF-α, IL-6, CD-14, and TREM-1 expression (p < 0.05 for all) and inhibited LPS-induced NF-κB activation in macrophages in vitro. However, treatment with both etomidate and LP17 did not enhance the inhibitory effects in macrophages. Hence, etomidate mitigates LPS-up-regulated pro-inflammatory cytokine production and inhibits LPS-enhanced CD14 and TREM-1 expression and NF-κB activation in macrophages.

  11. Diverse age-related effects of Bacopa monnieri and donepezil in vitro on cytokine production, antioxidant enzyme activities, and intracellular targets in splenocytes of F344 male rats.

    PubMed

    Priyanka, Hannah P; Singh, Ran Vijay; Mishra, Miti; ThyagaRajan, Srinivasan

    2013-02-01

    Aged people are more prone to developing neurodegenerative and infectious diseases, autoimmune disorders, and cancer due to impairment of neuroendocrine-immune functions. Neuronal degeneration and immunosuppression aided by increased generation of reactive oxygen species combined with loss of antioxidant enzyme activities promote the aging process. Bacopa monnieri (brahmi), an Ayurvedic herb, and donepezil, a reversible acetylcholinesterase inhibitor, have been used to reverse cognitive dysfunctions in several neurodegenerative diseases. The aim of this study was to investigate the effects of in vitro incubation of lymphocytes from spleens of young (3-month-old), early middle-aged (8- to 9-month-old), and old (18-month-old) F344 rats with brahmi (0.001%, 0.01%, 0.05%, 0.1%, and 1%) and donepezil (5, 10, 25, 50, and 100 μg/ml) on Concanavalin (Con A)-induced proliferation of T lymphocytes and cytokine production, and the activities of antioxidant enzymes [superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) and glutathione-S-transferase (GST)]. In addition, the effects of these compounds on the expression of intracellular signaling pathway markers (ERK, p-ERK, CREB, p-CREB, Akt and p-Akt), nitric oxide (NO) production, and the extent of lipid peroxidation were measured in the splenocytes. Age-related decline in Con A-induced proliferation of T lymphocytes was not reversed by treatment with brahmi and donepezil but donepezil alone further reduced the lymphocyte proliferation in young rats. Lower doses of brahmi treatment reversed the age-related decrease in Con A-induced IL-2 and IFN-γ production by the splenocytes while their production by splenocytes was suppressed by treatment with donepezil in the young and early middle-aged rats. An age-associated decline in the activities of SOD, CAT, GPx, and GST was evident in the lymphocytes of spleen. Brahmi enhanced CAT activity of lymphocytes in all the age groups while donepezil increased SOD

  12. Cytokine production and dysregulation in HIV pathogenesis: lessons for development of therapeutics and vaccines.

    PubMed

    Reuter, Morgan A; Pombo, Carolina; Betts, Michael R

    2012-01-01

    Numerous studies have characterized the cytokine modulation observed in human immunodeficiency virus (HIV) infected individuals, from initial infection through chronic disease. Progressive and non-progressive HIV infection models show the cytokine milieu differs in terms of production and responsiveness in these two groups, suggesting an understanding of the role cytokines play during infection is necessary for directing the immune response toward viral control. This review will cover cytokine induction and dysfunction during HIV pathogenesis, with a focus on the interplay between cytokines and transcription factors, T cell activation, and exhaustion. We highlight cytokines that have either vaccine adjuvant or therapeutic potential and discuss the need to identify key factors required for prevention of progression, clearance of infection, or protection from acquisition.

  13. Low level exposure to monomethyl arsonous acid-induced the over-production of inflammation-related cytokines and the activation of cell signals associated with tumor progression in a urothelial cell model

    SciTech Connect

    Escudero-Lourdes, C.; Medeiros, M.K.; Cardenas-Gonzalez, M.C.; Wnek, S.M.; Gandolfi, J.A.

    2010-04-15

    Human bladder cancer has been associated with chronic exposure to arsenic. Chronic exposure of an immortalized non-tumorigenic urothelial cell line (UROtsa cells) to arsenicals has transformed these cells to a malignant phenotype, but the involved mechanisms are not fully understood. Chronic inflammation has been linked with cancer development mainly because many pro-inflammatory cytokines, growth factors as well as angiogenic chemokines have been found in tumors. In this study the chronology of inflammatory cytokines production was profiled in UROtsa cells chronically exposed to the toxic arsenic metabolite, monomethylarsonous acid [50 nM MMA(III)] to know the role of inflammation in cell transformation. Acute 50 nM MMA(III) exposure induced over-production of many pro-inflammatory cytokines as soon as 12 h after acute exposure. The same cytokines remain over-regulated after chronic exposure to 50 nM MMA(III), especially after 3 mo exposure. At 3 mo exposure the sustained production of cytokines like IL-1, IL-6, IL-8 and TNF is coincident with the appearance of characteristics associated with cell transformation seen in other arsenic-UROtsa studies. The sustained and increased activation of NFkappaB and c-Jun is also present along the transformation process and the phosphorylated proteins p38 MAPK and ERK 1/2 are increased also through the time line. Taken together these results support the notion that chronic inflammation is associated within MMA(III)-induced cell transformation and may act as a promoting factor in UROtsa cell transformation.

  14. Saponins from Sanguisorba officinalis Improve Hematopoiesis by Promoting Survival through FAK and Erk1/2 Activation and Modulating Cytokine Production in Bone Marrow

    PubMed Central

    Chen, Xin; Li, Bogang; Gao, Yue; Ji, Jianxin; Wu, Zhongliu; Chen, Shuang

    2017-01-01

    Radix Sanguisorbae, the root of Sanguisorba officinalis L. is used as traditional Chinese medicine. In recent decades, it has been reported to be clinically effective against myelosuppression induced by chemotherapy and/ or radiotherapy. However, the underlining mechanism has not been well studied. In this work, we evaluated the hematopoietic effect of total saponins from S. officinalis L. on myelosuppressive mice induced by cyclophosphamide and by60Co-γ-irradiation and confirmed the therapeutic effect. Then, we found total saponins and their characteristic constituents Ziyuglycoside I and Ziyuglycoside II can inhibit apoptosis of TF-1 cells caused by cytokine deprivation, and promote survival of mouse bone marrow nuclear cells through focal adhesion kinase (FAK) and extracellular signal-regulated kinase 1/2 (Erk1/2) activation in vitro. In addition, they can down-regulate macrophage inflammatory protein 2 (MIP-2), platelet factor 4 (PF4) and P-selectin secretion, which are reported to be suppressive to hematopoiesis, both in vitro and in vivo. These results suggest that promotion of survival through FAK and Erk1/2 activation and inhibition of suppressive cytokines in the bone marrow is likely to be the pharmacological mechanism underlying the hematopoietic effect of saponins from S. officinalis L. PMID:28360858

  15. Natural product HTP screening for attenuation of cytokine-induced neutrophil chemo attractants (CINCs) and NO2- in LPS/IFNγ activated glioma cells.

    PubMed

    Mazzio, Elizabeth A; Bauer, David; Mendonca, Patricia; Taka, Equar; Soliman, Karam F A

    2017-01-15

    Chronic and acute central nervous system (CNS) inflammation are contributors toward neurological injury associated with head trauma, stroke, infection, Parkinsons or Alzheimers disease. CNS inflammatory illnesses can also contribute toward risk of developing glioblastoma multiforme (GBM). With growing public interest in complementary and alternative medicines (CAMs), we conduct a high throughput (HTP) screening of >1400 natural herbs, plants and over the counter (OTC) products for anti-inflammatory effects on lipopolysaccharide (LPS)/interferon gamma (IFNγ) activated C6 glioma cells. Validation studies were performed showing a pro-inflammatory profile of [LPS 3 µg/ml/ IFNγ 3 ng/ml] consistent with greater release [>8.5 fold] of MCP-1, NO2-, cytokine-induced neutrophil chemo-attractants (CINC) 1, CINC 2a and CINC3. The data show no changes to the following, IL-13, TNF-a, fracktaline, leptin, LIX, GM-CSF, ICAM1, L-Selectin, activin A, agrin, IL-1α, MIP-3a, B72/CD86, NGF, IL-1b, MMP-8, IL-1 R6, PDGF-AA, IL-2, IL-4, prolactin R, RAGE, IL-6, Thymus Chemokine-1, CNTF,IL-10 or TIMP-1. A HTP screening was conducted, where we employ an in vitro efficacy index (iEI) defined as the ratio of toxicity (LC50)/anti-inflammatory potency (IC50). The iEI was precautionary to ensure biological effects were occurring in fully viable cells (ratio > 3.8) independent of toxicity. Using NO2- as a guideline molecule, the data show that 1.77% (25 of 1410 tested) had anti-inflammatory effects with iEI ratios >3.8 and IC50s <250µg/ml. These include reference drugs (hydrocortisone, dexamethasone N6-(1-iminoethyl)-l-lysine and NSAIDS: diclofenac, tolfenamic acid), a histone deacetylase inhibitor (apicidin) and the following natural products; Ashwaganda (Withania somnifera), Elecampagne Root (Inula helenium), Feverfew (Tanacetum parthenium), Green Tea (Camellia sinensis), Turmeric Root (Curcuma longa) Ganthoda (Valeriana wallichii), Tansy (Tanacetum vulgare), Maddar Root (Rubia tinctoria

  16. Maximal adjuvant activity of nasally delivered IL-1α requires adjuvant responsive CD11c+ cells and does not correlate with adjuvant-induced in vivo cytokine production1

    PubMed Central

    Thompson, Afton L.; Johnson, Brandi T.; Sempowski, Gregory D.; Gunn, Michael D.; Hou, Baidong; DeFranco, Anthony L.; Staats, Herman F.

    2012-01-01

    IL-1 has been shown to have strong mucosal adjuvant activities, but little is known about its mechanism of action. We vaccinated IL-1R1 bone marrow chimeric mice to determine if IL-1R1 expression on stromal cells or hematopoietic cells was sufficient for the maximal adjuvant activity of nasally delivered IL-1α as determined by the acute induction of cytokine responses and induction of Bacillus anthracis lethal factor (LF)-specific adaptive immunity. Cytokine and chemokine responses induced by vaccination with IL-1α were predominantly derived from the stromal cell compartment and included G-CSF, IL-6, IL-13, MCP-1, and KC. Nasal vaccination of Il1r1−/− mice given wild-type bone marrow (WT→KO) and WT→WT mice with LF + IL-1α induced maximal adaptive immune responses, while vaccination of wild-type mice given Il1r1−/− bone marrow (KO→WT) resulted in significantly decreased production of LF-specific serum IgG, IgG subclasses, lethal toxin-neutralizing antibodies, and mucosal IgA compared to WT→KO and WT→WT mice (p < 0.05). IL-1α adjuvant activity was not dependent upon mast cells. However, the ability of IL-1α to induce serum LF-specific IgG2c and lethal toxin-neutralizing antibodies was significantly impaired in CD11c-Myd88−/− mice when compared to WT mice (p < 0.05). Our results suggest that CD11c+ cells must be directly activated by nasally administered IL-1α for maximal adjuvant activity and that, while stromal cells are required for maximal adjuvant-induced cytokine production, the adjuvant-induced stromal cell cytokine responses are not required for effective induction of adaptive immunity. PMID:22345651

  17. Endothelin Regulates Porphyromonas gingivalis-Induced Production of Inflammatory Cytokines

    PubMed Central

    Kim, Ji-Hye; Lee, Dong Eun; Kang, Si-Mook; Lee, So Yun; Choi, Lin; Sun, Ji Su; Kim, Seul Ki; Park, Wonse; Kim, Baek Il; Yoo, Yun-Jung; Chang, Inik; Shin, Dong Min

    2016-01-01

    Periodontitis is a very common oral inflammatory disease that results in the destruction of supporting connective and osseous tissues of the teeth. Although the exact etiology is still unclear, Gram-negative bacteria, especially Porphyromonas gingivalis in subgingival pockets are thought to be one of the major etiologic agents of periodontitis. Endothelin (ET) is a family of three 21-amino acid peptides, ET-1, -2, and -3, that activate G protein-coupled receptors, ETA and ETB. Endothelin is involved in the occurrence and progression of various inflammatory diseases. Previous reports have shown that ET-1 and its receptors, ETA and ETB are expressed in the periodontal tissues and, that ET-1 levels in gingival crevicular fluid are increased in periodontitis patients. Moreover, P. gingivalis infection has been shown to induce the production of ET-1 along with other inflammatory cytokines. Despite these studies, however, the functional significance of endothelin in periodontitis is still largely unknown. In this study, we explored the cellular and molecular mechanisms of ET-1 action in periodontitis using human gingival epithelial cells (HGECs). ET-1 and ETA, but not ETB, were abundantly expressed in HGECs. Stimulation of HGECs with P. gingivalis or P. gingivalis lipopolysaccharide increased the expression of ET-1 and ETA suggesting the activation of the endothelin signaling pathway. Production of inflammatory cytokines, IL-1β, TNFα, and IL-6, was significantly enhanced by exogenous ET-1 treatment, and this effect depended on the mitogen-activated protein kinases via intracellular Ca2+ increase, which resulted from the activation of the phospholipase C/inositol 1,4,5-trisphosphate pathway. The inhibition of the endothelin receptor-mediated signaling pathway with the dual receptor inhibitor, bosentan, partially ameliorated alveolar bone loss and immune cell infiltration. These results suggest that endothelin plays an important role in P. gingivalis

  18. Differential Modulation of Lipopolysaccharide-Induced Inflammatory Cytokine Production by and Antioxidant Activity of Fomentariol in RAW264.7 Cells

    PubMed Central

    Seo, Dong-Won; Yi, Young-Joo; Lee, Myeong-Seok

    2015-01-01

    Medicinal mushrooms have been used worldwide to treat cancer and modulate the immune system. Over the last several years, there has been increasing interest in isolating bioactive compounds from medicinal mushrooms and evaluating their health beneficial effects. Fomes fomentarius is used in traditional oriental medicine and is known to possess antioxidant, anti-inflammatory, antidiabetic, and antitumor effects. In the present study, we isolated fomentariol from Fomes fomentarius and investigated its anti-inflammatory effect in murine macrophages (RAW264.7 cells) stimulated with lipopolysaccharides. Fomentariol inhibited the production of nitric oxide and intracellular reactive oxygen species triggered by lipopolysaccharides. Interestingly, fomentariol differentially regulated cytokine production triggered by lipopolysaccharides. Fomentariol effectively suppressed the production of interleukin-1β and interleukin-6 but not tumor necrosis factor-α. The inhibitory effect of fomentariol against nitric oxide, interleukin-1β, and interleukin-6 production was possibly mediated by downregulation of the extracellular signal-regulated kinase signaling pathway. Taken together, our results suggest that fomentariol differentially modulated inflammatory responses triggered by lipopolysaccharides in macrophages and is one of the bioactive compounds that mediate the physiological effects of Fomes fomentarius. PMID:26839505

  19. Diverse Toll-like receptors mediate cytokine production by Fusobacterium nucleatum and Aggregatibacter actinomycetemcomitans in macrophages.

    PubMed

    Park, Se-Ra; Kim, Dong-Jae; Han, Seung-Hyun; Kang, Min-Jung; Lee, Jun-Young; Jeong, Yu-Jin; Lee, Sang-Jin; Kim, Tae-Hyoun; Ahn, Sang-Gun; Yoon, Jung-Hoon; Park, Jong-Hwan

    2014-05-01

    Toll-like receptors (TLRs) orchestrate a repertoire of immune responses in macrophages against various pathogens. Fusobacterium nucleatum and Aggregatibacter actinomycetemcomitans are two important periodontal pathogens. In the present study, we investigated TLR signaling regulating cytokine production of macrophages in response to F. nucleatum and A. actinomycetemcomitans. TLR2 and TLR4 are redundant in the production of cytokines (interleukin-6 [IL-6] and tumor necrosis factor alpha [TNF-α]) in F. nucleatum- and A. actinomycetemcomitans-infected macrophages. The production of cytokines by macrophages in response to F. nucleatum and A. actinomycetemcomitans infection was impaired in MyD88-deficient macrophages. Moreover, cytokine concentrations were lower in MyD88-deficient macrophages than in TLR2/TLR4 (TLR2/4) double-deficient cells. An endosomal TLR inhibitor, chloroquine, reduced cytokine production in TLR2/4-deficient macrophages in response to F. nucleatum and A. actinomycetemcomitans, and DNA from F. nucleatum or A. actinomycetemcomitans induced IL-6 production in bone marrow-derived macrophages (BMDMs), which was abolished by chloroquine. Western blot analysis revealed that TLR2/4 and MyD88 were required for optimal activation of NF-κB and mitogen-activated protein kinases (MAPKs) in macrophages in response to F. nucleatum and A. actinomycetemcomitans, with different kinetics. An inhibitor assay showed that NF-κB and all MAPKs (p38, extracellular signal-regulated kinase [ERK], and Jun N-terminal protein kinase [JNK]) mediate F. nucleatum-induced production of cytokines in macrophages, whereas NF-κB and p38, but not ERK and JNK, are involved in A. actinomycetemcomitans-mediated cytokine production. These findings suggest that multiple TLRs may participate in the cytokine production of macrophages against periodontal bacteria.

  20. Xanthohumol and related prenylated flavonoids inhibit inflammatory cytokine production in LPS-activated THP-1 monocytes: structure-activity relationships and in silico binding to myeloid differentiation protein-2 (MD-2).

    PubMed

    Peluso, Michael R; Miranda, Cristobal L; Hobbs, Deborah J; Proteau, Rosita R; Stevens, Jan Frederik

    2010-10-01

    Xanthohumol (XN) is a prenylated chalcone-type flavonoid found in hops and beer. Our objective of this study was to determine the anti-inflammatory activities of XN, isoxanthohumol (IX), and 15 related prenylated chalcones and flavanones, as well as their structure-activity relationships. The anti-inflammatory activities of the flavonoids were measured by their ability to inhibit lipopolysaccharide (LPS)-induced cytokine production in human monocytic THP-1 cells. The position, number, and length of the prenyl groups had a marked influence on the inhibitory activity of the prenylfavonoids towards MCP-1 and IL-6 production. The α,β-unsaturated carbonyl moiety present in chalcones such as XN was not an absolute requirement for inhibitory activity, as the saturated XN derivative, tetrahydroxanthohumol (TX), showed inhibitory activity comparable to XN. With the aim to determine the mechanism of the observed anti-inflammatory effects, cellular protein levels of Toll-like receptor 4 (TLR4) were measured by Western blot 24 h following coexposure of THP-1 cells to LPS and either XN, TX, or IX. Only XN reduced the cellular TLR4 protein content. Therefore, an additional hypothesis was developed for an anti-inflammatory mechanism that involves the TLR4 coreceptor myeloid differentiation protein-2 (MD-2), which provides the actual binding site for LPS. Molecular docking studies showed that the complementarity of prenylated flavonoids with the hydrophobic MD-2 pocket (indicating goodness of fit) directly predicted their relative ability to inhibit MCP-1 and IL-6 production. In conclusion, prenylated flavonoids may suppress LPS-induced TLR4 activation at least partly by interfering with LPS binding to the TLR4 coreceptor MD-2, and XN (but not other prenylflavonoids) exerts an additional anti-inflammatory effect by downregulating the cellular TLR4 protein content.

  1. Short-term heating reduces the anti-inflammatory effects of fresh raw garlic extracts on the LPS-induced production of NO and pro-inflammatory cytokines by downregulating allicin activity in RAW 264.7 macrophages.

    PubMed

    Shin, Jung-Hye; Ryu, Ji Hyeon; Kang, Min Jung; Hwang, Cho Rong; Han, Jaehee; Kang, Dawon

    2013-08-01

    Garlic has a variety of biologic activities, including anti-inflammatory properties. Although garlic has several biologic activities, some people dislike eating fresh raw garlic because of its strong taste and smell. Therefore, garlic formulations involving heating procedures have been developed. In this study, we investigated whether short-term heating affects the anti-inflammatory properties of garlic. Fresh and heated raw garlic extracts (FRGE and HRGE) were prepared with incubation at 25 °C and 95 °C, respectively, for 2 h. Treatment with FRGE and HRGE significantly reduced the LPS-induced increase in the pro-inflammatory cytokine concentration (TNF-α, IL-1β, and IL-6) and NO through HO-1 upregulation in RAW 264.7 macrophages. The anti-inflammatory effect was greater in FRGE than in HRGE. The allicin concentration was higher in FRGE than in HRGE. Allicin treatment showed reduced production of pro-inflammatory cytokines and NO and increased HO-1 activity. The results show that the decrease in LPS-induced NO and pro-inflammatory cytokines in RAW 264.7 macrophages through HO-1 induction was greater for FRGE compared with HRGE. Additionally, the results indicate that allicin is responsible for the anti-inflammatory effect of FRGE. Our results suggest a potential therapeutic use of allicin in the treatment of chronic inflammatory disease.

  2. Lipopolysaccharide-induced lethality and cytokine production in aged mice.

    PubMed Central

    Tateda, K; Matsumoto, T; Miyazaki, S; Yamaguchi, K

    1996-01-01

    This study was designed to define the lipopolysaccharide (LPS) sensitivity of aged mice in terms of lethality and cytokine production and to determine down-regulating responses of corticosterone and interleukin 10 (IL-10). The 50% lethal doses of LPS in young (6- to 7-week-old) and aged (98- to 102-week-old) mice were 601 and 93 microg per mouse (25.6 and 1.6 mg per kg of body weight), respectively. Aged mice were approximately 6.5-fold more sensitive to the lethal toxicity of LPS in micrograms per mouse (16-fold more sensitive in milligrams per kilogram) than young mice. Levels in sera of tumor necrosis factor-alpha (TNF-alpha) IL-1alpha, and IL-6 after intraperitoneal injection of 100 microg of LPS peaked at 1.5, 3, and 3 h, respectively, and declined thereafter in both groups of mice. However, the peak values of these cytokines were significantly higher in aged than in young mice (P < 0.05). Gamma interferon (IFN-gamma) was detectable at 3 h, and sustained high levels were still detected after 12 h in both age groups. Although there were no significant differences in levels of IFN-gamma in sera from both groups, aged mice showed higher IFN-gamma levels throughout the 3- to 12-h study period. Administration of increasing doses of LPS revealed that aged mice had a lower threshold to IL-1alpha production than young mice. In addition, aged mice were approximately 4-fold more sensitive to the lethal toxicity of exogenous TNF in units per mouse (10-fold more sensitive in units per kilogram) than young mice. With regard to down-regulating factors, corticosterone amounts were similar at basal levels and no differences in kinetics after the LPS challenge were observed, whereas IL-10 levels in sera were significantly higher in aged mice at 1.5 and 3 h than in young mice (P < 0.01). These results indicate that aged mice are more sensitive to the lethal toxicities of LPS and TNF than young mice. We conclude that a relatively activated, or primed, state for LPS

  3. Anti-inflammatory activity of betalain-rich dye of Beta vulgaris: effect on edema, leukocyte recruitment, superoxide anion and cytokine production.

    PubMed

    Martinez, Renata M; Longhi-Balbinot, Daniela T; Zarpelon, Ana C; Staurengo-Ferrari, Larissa; Baracat, Marcela M; Georgetti, Sandra R; Sassonia, Rogério C; Verri, Waldiceu A; Casagrande, Rubia

    2015-04-01

    We have recently developed betalain-rich beetroot (Beta vulgaris) dye (betalain) to be used in food products. Betalain (30-300 mg/kg) intraperitoneal (i.p.) treatment diminished carrageenan (100 µg/paw)-induced paw edema and neutrophil migration to the paw skin tissue. Betalain (100 mg/kg) treatment by subcutaneous or per oral routes also inhibited the carrageenan-induced paw edema. Importantly, the post-treatment with betalain (100 mg/kg, i.p.) significantly inhibited carrageenan- and complete Freund's adjuvant (10 µl/paw)-induced paw edema. Betalain (100 mg/kg) also reduced carrageenan (500 µg/cavity)-induced recruitment of total leukocytes, including mononuclear cells and neutrophils, as well as increasing vascular permeability in the peritoneal cavity. Furthermore, betalain significantly reduced carrageenan-induced superoxide anion, tumor necrosis factor-alpha (TNF-α) and interleukin (IL)-1β levels in the peritoneal fluid, as well as augmenting IL-10 levels. Therefore, this compound presents prominent anti-inflammatory effect on carrageenan-induced paw edema and peritonitis by reducing the production of superoxide anion and the cytokines TNF-α and IL-1β, in addition to increasing IL-10 levels. These results suggest that betalain shows therapeutic potential that could be utilized in the treatment of inflammation-associated diseases.

  4. Combined glutamine and arginine decrease proinflammatory cytokine production by biopsies from Crohn's patients in association with changes in nuclear factor-kappaB and p38 mitogen-activated protein kinase pathways.

    PubMed

    Lecleire, Stéphane; Hassan, Aktham; Marion-Letellier, Rachel; Antonietti, Michel; Savoye, Guillaume; Bôle-Feysot, Christine; Lerebours, Eric; Ducrotté, Philippe; Déchelotte, Pierre; Coëffier, Moïse

    2008-12-01

    Glutamine (Gln) and arginine (Arg) are conditionally essential amino acids with immunomodulatory properties. The aim of the study was to assess the effects of Gln and Arg alone or in combination on cytokine release by cultured colonic biopsies from patients with active Crohn's disease (CD). Ten consecutive patients [mean (range) age 26 (18-39) y] with active colonic CD (mean CD activity index: 383.7 +/- 129.8) were prospectively included in the study. Eight colonic biopsies were obtained via a colonoscopy and incubated during 18 h with low (physiological) or high (pharmacological) doses of Arg (0.1 or 2 mmol/L designated as Arg(low) or Arg(high), respectively) and Gln (0.6 or 10 mmol/L designated as Gln(low) or Gln(high), respectively). The concentrations of cytokines [interleukin (IL)-4, IL-10, IL-8, IL-6, tumor necrosis factor-alpha (TNFalpha), IL-1beta, interferon-gamma) were assessed by ELISA, and nitric oxide (NO) production was evaluated by Griess assay. Nuclear factor (NF)-kappaB p65 subunit, inhibitor of NFkappaB-alpha, and p38 mitogen-activated protein kinase (MAPK) were assessed by immunoblotting. Arg(high)/Gln(high) decreased the production of TNFalpha, IL-1beta, IL-8, and IL-6 (each P < 0.01). Arg(low)/Gln(high) decreased IL-6 and IL-8 production (both P < 0.01), whereas Arg(high)/Gln(low) did not affect cytokine and NO production. Arg(low)/Gln(high) and Arg(high)/Gln(high) decreased NF-kappaB p65 subunit expression, whereas p38 MAPK was decreased only by Arg(high)/Gln(high). Combined pharmacological doses of Arg and Gln decreased TNFalpha and the main proinflammatory cytokines release in active colonic CD biopsies via NF-kappaB and p38 MAPK pathways. These results could be the basis of prospective studies evaluating the effects of enteral supply of combined Arg and Gln during active CD.

  5. Increased production of inflammatory cytokines in mild cognitive impairment.

    PubMed

    Magaki, Shino; Mueller, Claudius; Dickson, Cindy; Kirsch, Wolff

    2007-03-01

    Recent studies indicate that chronic inflammation plays a pathogenic role in both the central nervous system (CNS) and periphery in Alzheimer's disease (AD). We have screened for cytokines differentially produced by peripheral blood mononuclear cells (PBMCs) isolated from subjects with mild cognitive impairment (MCI) and mild AD subjects who had progressed from MCI using a commercially available cytokine array. Following determination of expressed cytokines, we quantified levels of the proinflammatory cytokines TNF-alpha, IL-6, and IL-8, and the anti-inflammatory cytokine IL-10 using flow cytometry. We have found a significant increase in the levels of IL-6, IL-8, and IL-10 produced by PBMCs stimulated for 24 h with phytohemagglutinin (PHA) in MCI subjects compared to healthy elderly controls. However, in PBMCs stimulated for 48 h with lipopolysaccharide (LPS), lower TNF-alpha/IL-10, IL-6/IL-10, and IL-8/IL-10 ratios were seen in MCI subjects. There were no differences in plasma levels of IL-8 between aged controls, MCI, and mild AD, and the levels of circulating IL-6 and IL-10 were below detection limits. Our data indicate that changes in cytokine production by PBMCs may be detected early in MCI, and an alteration of the immune response may precede clinical AD.

  6. Activation of kynurenine pathway in ex vivo fibroblasts from patients with bipolar disorder or schizophrenia: cytokine challenge increases production of 3-hydroxykynurenine.

    PubMed

    Johansson, Anne-Sofie; Owe-Larsson, Björn; Asp, Linnéa; Kocki, Tomasz; Adler, Mats; Hetta, Jerker; Gardner, Renee; Lundkvist, Gabriella B S; Urbanska, Ewa M; Karlsson, Håkan

    2013-11-01

    Accumulating data suggest a causative link between immune stimulation, disturbed metabolism of tryptophan, and pathogenesis of bipolar disorder and schizophrenia. The goal of this study was to examine the production of kynurenic acid (KYNA), 3-hydroxykynurenine (3-HK) and the expression of kynurenine pathway enzymes involved in their synthesis and metabolism in cultured skin fibroblasts obtained from patients with bipolar disorder, schizophrenia or from healthy control individuals. The assessment was performed under basal conditions or following treatment with interferon (IFN)-γ, tumor necrosis factor (TNF)-α, interleukin (IL)-1β, IL-6, or their combinations, in cells exposed to exogenous kynurenine. In both groups of patients, the baseline production of KYNA and 3-HK was increased, as compared to control subjects. Case-treatment analyses revealed significant interactions between bipolar case status and IL-1β, IL-6, IFN-γ + TNF-α, or IFN-γ + IL-1β, as well as between schizophrenia case status and IL-1β, IFN-γ + TNF-α, or IFN-γ + IL-1β, in terms of higher 3-HK. Noteworthy, no case-treatment interactions in terms of KYNA production were found. Observed changes did not appear to correlate with the expression of genes encoding kynurenine aminotransferases (KATs), kynureninase (KYNU) or kynurenine-3-monooxygenase (KMO). The single nucleotide polymorphisms (SNPs), rs1053230 and rs2275163, in KMO influenced KYNA levels yet did not explain the case-treatment discrepancies. In conclusion, our present findings indicate the utility of skin-derived fibroblasts for kynurenines research and support the concept of kynurenine pathway alterations in bipolar disorder and schizophrenia. The increase in ratio between neurotoxic 3-HK and neuroinhibitory/neuroprotective KYNA following exposure to cytokines may account for altered neurogenesis and structural abnormalities characteristic for both diseases.

  7. Effects of bucillamine and N-acetyl-l-cysteine on cytokine production and collagen-induced arthritis (CIA)

    PubMed Central

    Tsuji, F; Miyake, Y; Aono, H; Kawashima, Y; Mita, S

    1999-01-01

    We investigated the effects of bucillamine and N-acetyl-l-cysteine (NAC) on cytokine production and CIA. Bucillamine and NAC inhibited NF-κB activation and tumour necrosis factor-alpha (TNF-α) mRNA expression in human monocytic leukaemia cell line THP-1, and cytokine production from monocyte cell lines at concentrations >10−3 m. They also inhibited cytokine production and CIA in mice at a dose of 500 mg/kg. These results suggest that NF-κB inhibitors such as bucillamine and NAC may inhibit cytokine-related diseases, including arthritis. PMID:9933417

  8. Adenosine modulates LPS-induced cytokine production in porcine monocytes.

    PubMed

    Ondrackova, Petra; Kovaru, Hana; Kovaru, Frantisek; Leva, Lenka; Faldyna, Martin

    2013-03-01

    Adenosine plays an important role during inflammation, particularly through modulation of monocyte function. The objective of the present study was to evaluate the effect of synthetic adenosine analogs on cytokine production by porcine monocytes. The LPS-stimulated cytokine production was measured by flow cytometry and quantitative real-time PCR. Adenosine receptor expression was measured by quantitative real-time PCR. The present study demonstrates that adenosine analog N-ethylcarboxyamidoadenosine (NECA) down-regulates TNF-α production and up-regulates IL-8 production by LPS-stimulated porcine monocytes. The effect was more pronounced in CD163(-) subset of monocytes compared to the CD163(+) subset. Although both monocyte subsets express mRNA for A1, A2A, A2B and A3 adenosine receptors, the treatment of monocytes with various adenosine receptor agonists and antagonists proved that the effect of adenosine is mediated preferentially via A2A adenosine receptor. Moreover, the study suggests that the effect of NECA on porcine monocytes alters the levels of the cytokines which could play a role in the differentiation of naive T cells into Th17 cells. The results suggest that adenosine plays an important role in modulation of cytokine production by porcine monocytes.

  9. Kaurenoic acid from Sphagneticola trilobata Inhibits Inflammatory Pain: effect on cytokine production and activation of the NO-cyclic GMP-protein kinase G-ATP-sensitive potassium channel signaling pathway.

    PubMed

    Mizokami, Sandra S; Arakawa, Nilton S; Ambrosio, Sergio R; Zarpelon, Ana C; Casagrande, Rubia; Cunha, Thiago M; Ferreira, Sergio H; Cunha, Fernando Q; Verri, Waldiceu A

    2012-05-25

    Kaurenoic acid [ent-kaur-16-en-19-oic acid (1)] is a diterpene present in several plants including Sphagneticola trilobata. The only documented evidence for its antinociceptive effect is that it inhibits the writhing response induced by acetic acid in mice. Therefore, the analgesic effect of 1 in different models of pain and its mechanisms in mice were investigated further. Intraperitoneal and oral treatment with 1 dose-dependently inhibited inflammatory nociception induced by acetic acid. Oral treatment with 1 also inhibited overt nociception-like behavior induced by phenyl-p-benzoquinone, complete Freund's adjuvant (CFA), and both phases of the formalin test. Compound 1 also inhibited acute carrageenin- and PGE(2)-induced and chronic CFA-induced inflammatory mechanical hyperalgesia. Mechanistically, 1 inhibited the production of the hyperalgesic cytokines TNF-α and IL-1β. Furthermore, the analgesic effect of 1 was inhibited by l-NAME, ODQ, KT5823, and glybenclamide treatment, demonstrating that such activity also depends on activation of the NO-cyclic GMP-protein kinase G-ATP-sensitive potassium channel signaling pathway, respectively. These results demonstrate that 1 exhibits an analgesic effect in a consistent manner and that its mechanisms involve the inhibition of cytokine production and activation of the NO-cyclic GMP-protein kinase G-ATP-sensitive potassium channel signaling pathway.

  10. High efficiency cell-specific targeting of cytokine activity

    NASA Astrophysics Data System (ADS)

    Garcin, Geneviève; Paul, Franciane; Staufenbiel, Markus; Bordat, Yann; van der Heyden, José; Wilmes, Stephan; Cartron, Guillaume; Apparailly, Florence; de Koker, Stefaan; Piehler, Jacob; Tavernier, Jan; Uzé, Gilles

    2014-01-01

    Systemic toxicity currently prevents exploiting the huge potential of many cytokines for medical applications. Here we present a novel strategy to engineer immunocytokines with very high targeting efficacies. The method lies in the use of mutants of toxic cytokines that markedly reduce their receptor-binding affinities, and that are thus rendered essentially inactive. Upon fusion to nanobodies specifically binding to marker proteins, activity of these cytokines is selectively restored for cell populations expressing this marker. This ‘activity-by-targeting’ concept was validated for type I interferons and leptin. In the case of interferon, activity can be directed to target cells in vitro and to selected cell populations in mice, with up to 1,000-fold increased specific activity. This targeting strategy holds promise to revitalize the clinical potential of many cytokines.

  11. Hematopoietic cytokines.

    PubMed

    Metcalf, Donald

    2008-01-15

    The production of hematopoietic cells is under the tight control of a group of hematopoietic cytokines. Each cytokine has multiple actions mediated by receptors whose cytoplasmic domains contain specialized regions initiating the various responses-survival, proliferation, differentiation commitment, maturation, and functional activation. Individual cytokines can be lineage specific or can regulate cells in multiple lineages, and for some cell types, such as stem cells or megakaryocyte progenitors, the simultaneous action of multiple cytokines is required for proliferative responses. The same cytokines control basal and emergency hematopoietic cell proliferation. Three cytokines, erythropoietin, granulocyte colony-stimulating factor, and granulocyte-macrophage colony-stimulating factor, have now been in routine clinical use to stimulate cell production and in total have been used in the management of many millions of patients. In this little review, discussion will be restricted to those cytokines well established as influencing the production of hematopoietic cells and will exclude newer candidate regulators and those active on lymphoid cells. As requested, this account will describe the cytokines in a historical manner, using a sequential format of discovery, understanding, validation, and puzzlement, a sequence that reflects the evolving views on these cytokines over the past 50 years.

  12. Production of MMP-9 and inflammatory cytokines by Trypanosoma cruzi-infected macrophages.

    PubMed

    de Pinho, Rosa Teixeira; da Silva, Wellington Seguins; de Castro Côrtes, Luzia Monteiro; da Silva Vasconcelos Sousa, Periela; de Araujo Soares, Renata Oliveira; Alves, Carlos Roberto

    2014-12-01

    Matrix metalloproteinases (MMPs) constitute a large family of Zn(2+) and Ca(2+) dependent endopeptidases implicated in tissue remodeling and chronic inflammation. MMPs also play key roles in the activation of growth factors, chemokines and cytokines produced by many cell types, including lymphocytes, granulocytes, and, in particular, activated macrophages. Their synthesis and secretion appear to be important in a number of physiological processes, including the inflammatory process. Here, we investigated the interaction between human and mouse macrophages with T. cruzi Colombian and Y strains to characterize MMP-9 and cytokine production in this system. Supernatants and total extract of T. cruzi infected human and mouse macrophages were obtained and used to assess MMP-9 profile and inflammatory cytokines. The presence of metalloproteinase activity was determined by zymography, enzyme-linked immunosorbent assay and immunoblotting assays. The effect of cytokines on MMP-9 production in human macrophages was verified by previous incubation of cytokines on these cells in culture, and analyzed by zymography. We detected an increase in MMP-9 production in the culture supernatants of T. cruzi infected human and mouse macrophages. The addition of IL-1β or TNF-α to human macrophage cultures increased MMP-9 production. In contrast, MMP-9 production was down-modulated when human macrophage cultures were treated with IFN-γ or IL-4 before infection. Human macrophages infected with T. cruzi Y or Colombian strains produced increased levels of MMP-9, which was related to the production of cytokines such as IL-1β, TNF-α and IL-6.

  13. DNA from Porphyromonas gingivalis and Tannerella forsythia induce cytokine production in human monocytic cell lines.

    PubMed

    Sahingur, S E; Xia, X-J; Alamgir, S; Honma, K; Sharma, A; Schenkein, H A

    2010-04-01

    Toll-like receptor 9 (TLR9) expression is increased in periodontally diseased tissues compared with healthy sites indicating a possible role of TLR9 and its ligand, bacterial DNA (bDNA), in periodontal disease pathology. Here, we determine the immunostimulatory effects of periodontal bDNA in human monocytic cells (THP-1). THP-1 cells were stimulated with DNA of two putative periodontal pathogens: Porphyromonas gingivalis and Tannerella forsythia. The role of TLR9 in periodontal bDNA-initiated cytokine production was determined either by blocking TLR9 signaling in THP-1 cells with chloroquine or by measuring IL-8 production and nuclear factor-kappaB (NF-kappaB) activation in HEK293 cells stably transfected with human TLR9. Cytokine production (IL-1beta, IL-6, and TNF-alpha) was increased significantly in bDNA-stimulated cells compared with controls. Chloroquine treatment of THP-1 cells decreased cytokine production, suggesting that TLR9-mediated signaling pathways are operant in the recognition of DNA from periodontal pathogens. Compared with native HEK293 cells, TLR9-transfected cells demonstrated significantly increased IL-8 production (P < 0.001) and NF-kappaB activation in response to bDNA, further confirming the role of TLR9 in periodontal bDNA recognition. The results of PCR arrays demonstrated upregulation of proinflammatory cytokine and NF-kappaB genes in response to periodontal bDNA in THP-1 cells, suggesting that cytokine induction is through NF-kappaB activation. Hence, immune responses triggered by periodontal bacterial nucleic acids may contribute to periodontal disease pathology by inducing proinflammatory cytokine production through the TLR9 signaling pathway.

  14. Differences in Cytokine Production during Aging and Its Relationship with Antimicrobial Peptides Production.

    PubMed

    Castañeda-Delgado, Julio E; Frausto-Lujan, Isabel; González-Curiel, Irma; Montoya-Rosales, Alejandra; Serrano, Carmen J; Torres-Juarez, Flor; Enciso-Moreno, Jose A; Rivas-Santiago, Bruno

    2017-01-01

    Aging is a major health issue due to the increased susceptibility of elderly people to infectious, autoimmune, and cardiovascular diseases. Innate immunity is an important mechanism to avoid primary infections; therefore, decreasing of its activity may lead to development of infections. Antimicrobial peptides (AMPs) are effector molecules of innate immunity that can eliminate microbial invaders. The role that cytokines play in the regulation of these innate immune mechanisms needs to be explored. Serum determinations of Th1, Th2, and Th17 cytokines were performed in order to evaluate their association with AMPs human beta-defensin (HBD)-2 and LL-37 in young adults, elder adults, and elder adults with recurrent infections. Our results showed differences in interleukin (IL)-10 and IL-6 among the different groups. Inverse correlations in serum cytokine levels and HBD-2 production were identified for IL-10, IL-2, IL-4, tumor necrosis factor-α, and IL-6. Also inverse correlations were identified for IL-10, IL-4, and cathelicidin (LL-37). Such results could impact the development of immunomodulators that promote AMP production to prevent and/or contain infectious diseases in this population.

  15. Effects of cobalt chloride on nitric oxide and cytokines/chemokines production in microglia.

    PubMed

    Mou, Yan Hua; Yang, Jing Yu; Cui, Nan; Wang, Ji Ming; Hou, Yue; Song, Shuang; Wu, Chun Fu

    2012-05-01

    The involvement of microglial activation in metal neurotoxicity is becoming increasingly recognized. Some metal ions, such as zinc (II) and manganese (II), have been recently reported as microglial activators to induce the release of inflammatory mediators including cytokines, chemokines and nitric oxide (NO) which are involved in the pathogenesis of neurological diseases. Cobalt is essential for human life. However, excessive cobalt is cytotoxic and neurotoxic. In the present study, we determined cobalt-induced production of NO and cytokines/chemokines in N9 cells, a murine microglial cell line. High levels of cobalt significantly up-regulated iNOS mRNA and protein expression, which resulted in the release of NO. Cobalt induced the production of tumor necrosis factor α (TNF-α) and interleukin-6 (IL-6) in a concentration- and time-dependent manner in both N9 cells and primary mouse microglia and increased lipopolysaccharides (LPS)-induced cytokine production. Further study showed that cobalt induced cytokine production by a mechanism involving both nuclear factor kappa B (NF-κB) and p38 mitogen-activated protein kinase (MAPK) signaling pathways. The involvement of reactive oxygen species (ROS) in microglial activation was also confirmed. These findings suggested that cobalt neurotoxicity should be attributed not only directly to neuronal damage but also indirectly to microglial activation which might potentiate neuronal injury via elevation of proinflammatory mediator levels.

  16. Skin rejuvenation using cosmetic products containing growth factors, cytokines, and matrikines: a review of the literature

    PubMed Central

    Aldag, Caroline; Nogueira Teixeira, Diana; Leventhal, Phillip S

    2016-01-01

    Skin aging is primarily due to alterations in the dermal extracellular matrix, especially a decrease in collagen I content, fragmentation of collagen fibrils, and accumulation of amorphous elastin material, also known as elastosis. Growth factors and cytokines are included in several cosmetic products intended for skin rejuvenation because of their ability to promote collagen synthesis. Matrikines and matrikine-like peptides offer the advantage of growth factor-like activities but better skin penetration due to their much smaller molecular size. In this review, we summarize the commercially available products containing growth factors, cytokines, and matrikines for which there is evidence that they promote skin rejuvenation. PMID:27877059

  17. Diclofenac enhances proinflammatory cytokine-induced phagocytosis of cultured microglia via nitric oxide production

    SciTech Connect

    Kakita, Hiroki; Aoyama, Mineyoshi; Nagaya, Yoshiaki; Asai, Hayato; Hussein, Mohamed Hamed; Suzuki, Mieko; Kato, Shin; Saitoh, Shinji; Asai, Kiyofumi

    2013-04-15

    Influenza-associated encephalopathy (IAE) is a central nervous system complication with a high mortality rate, which is increased significantly by the non-steroidal anti-inflammatory drug diclofenac sodium (DCF). In the present study, we investigated the effects of DCF on brain immune cells (i.e. microglia) stimulated with three proinflammatory cytokines, namely tumor necrosis factor-α, interleukin-1β, and interferon-γ. Similar to previous findings in astrocytes, all three cytokines induced the expression of inducible NO synthase (iNOS), as well as NO production, in microglia. The addition of DCF to the culture system augmented iNOS expression and NO production. Immunocytochemical analysis and the phagocytosis assay revealed that cytokine treatment induced morphological changes to and phagocytosis by the microglia. The addition of DCF to the culture system enhanced microglial activation, as well as the phagocytic activity of cytokine-stimulated microglia. Inhibitors of nuclear factor (NF)-κB inhibited iNOS gene expression in cytokine-stimulated microglia with or without DCF, suggesting that the NF-κB pathway is one of the main signaling pathways involved. The iNOS inhibitor N{sup G}-monomethyl-L-arginine (L-NMMA) reduced both cytokine-induced phagocytosis and phagocytosis induced by the combination of cytokines plus DCF. Furthermore, the NO donor sodium nitroprusside induced phagocytosis, indicating that NO production is a key regulator of microglial phagocytosis. In conclusion, DCF acts synergistically with proinflammatory cytokines to increase the production of NO in microglia, leading to phagocytic activity of the activated microglia. These findings, together with previous observations regarding astrocytes, may explain the significant increase in mortality of IAE patients treated with DCF. - Highlights: ► Influenza-associated encephalopathy (IAE) is associated with a high mortality rate. ► Hyperimmunization in the brain is believed to be responsible for

  18. Chocolate consumption modulates cytokine production in healthy individuals.

    PubMed

    Netea, Stejara A; Janssen, Sam A; Jaeger, Martin; Jansen, Trees; Jacobs, Liesbeth; Miller-Tomaszewska, Gosia; Plantinga, Theo S; Netea, Mihai G; Joosten, Leo A B

    2013-04-01

    Epidemiological studies suggest that chocolate increases the incidence and severity of acne. Here we demonstrate that chocolate consumption primes human blood mononuclear cells from volunteers to release more interleukin-1β (IL-1β) and IL-10 upon stimulation with Propionibacterium acne or Staphylcoccus aureus, the two microorganisms involved in the pathogenesis of acne. In contrast, production of the Th17-derived cytokine IL-22 was inhibited by chocolate. Modulation of inflammation could represent an important mechanism through which chocolate consumption influences acne.

  19. Investigation of Macrophage Differentiation and Cytokine Production in an Undergraduate Immunology Laboratory

    ERIC Educational Resources Information Center

    Berkes, Charlotte; Chan, Leo Li-Ying

    2015-01-01

    We have developed a semester-long laboratory project for an undergraduate immunology course in which students study multiple aspects of macrophage biology including differentiation from progenitors in the bone marrow, activation upon stimulation with microbial ligands, expression of cell surface markers, and modulation of cytokine production. In…

  20. Products from mast cells influence T lymphocyte proliferation and cytokine production--relevant to allergic asthma?

    PubMed

    de Pater-Huijsen, F L; Pompen, M; Jansen, H M; Out, T A

    1997-06-01

    In IgE allergic diseases both mast cells and T lymphocytes play an important role. Whereas mast cels have been implicated in immediate allergic responses, T lymphocytes mediate subsequent late phase responses and chronic inflammation. Here we review possible links between the early mast cell activation and the later T lymphocyte stimulation. Products from mast cells were found to exert effects on T lymphocytes. Human Mast Cell line-1 (HMC-1) mast cells modulated proliferation and cytokine production of a human CD8+ T-cell clone in vitro. Activated mast cells seemed to drive this CD8+ T-cell clone towards a more pronounced T (helper) 1 type of response, simultaneously decreasing T-cell numbers. It is hypothesized that this might be a negative feed back mechanism operating in allergic subjects, by which the Th2-driven IgE production and eosinophilia are counteracted.

  1. Suppression of inflammatory reactions by terpinen-4-ol, a main constituent of tea tree oil, in a murine model of oral candidiasis and its suppressive activity to cytokine production of macrophages in vitro.

    PubMed

    Ninomiya, Kentaro; Hayama, Kazumi; Ishijima, Sanae A; Maruyama, Naho; Irie, Hiroshi; Kurihara, Junichi; Abe, Shigeru

    2013-01-01

    The onset of oral candidiasis is accompanied by inflammatory symptoms such as pain in the tongue, edema or tissue damage and lowers the quality of life (QOL) of the patient. In a murine oral candidiasis model, the effects were studied of terpinen-4-ol (T-4-ol), one of the main constituents of tea tree oil, Melaleuca alternifolia, on inflammatory reactions. When immunosuppressed mice were orally infected with Candida albicans, their tongues showed inflammatory symptoms within 24 h after the infection, which was monitored by an increase of myeloperoxidase activity and macrophage inflammatory protein-2 in their tongue homogenates. Oral treatment with 50 µL of 40 mg/mL terpinen-4-ol 3h after the Candida infection clearly suppressed the increase of these inflammatory parameters. In vitro analysis of the effects of terpinen-4-ol on cytokine secretion of macrophages indicated that 800 µg/mL of this substance significantly inhibited the cytokine production of the macrophages cultured in the presence of heat-killed C. albicans cells. Based on these findings, the role of the anti-inflammatory action of T-4-ol in its therapeutic activity against oral candidiasis was discussed.

  2. The influence of traditional herbal formulas on cytokine activity.

    PubMed

    Burns, J J; Zhao, Lijun; Taylor, Ethan Will; Spelman, Kevin

    2010-11-28

    Many of the botanical "immunomodulators", a class of herbal medicines widely recognized in traditional medical systems such as Chinese Medicine (TCM) and Ayurvedic Medicine, alter immune function and may offer clinically relevant therapeutics or leads to therapeutics. Many of these traditional remedies are prepared from combinations of medicinal plants which may influence numerous molecular pathways. These effects may differ from the sum of effects from the individual plants and therefore, research demonstrating the effects of the formula is crucial for insights into the effects of traditional remedies. In this review we surveyed the primary literature for research that focused on combinations of medicinal plants and effects on cytokine activity. The results demonstrate that many extracts of herb mixtures have effects on at least one cytokine. The most commonly studies cytokines were IL-4, IL-6, IL-10, TNF and IFN-γ. The majority of the formulas researched derived from TCM. The following formulas had activity on at least three cytokines; Chizukit N, CKBM, Daeganghwal-tang, Food Allergy Formula, Gamcho-Sasim-Tang, Hachimi-jio-gan, Herbkines, Hochuekki, Immune System Formula, Jeo-Dang-Tang, Juzen-taiho-to, Kakkon-to, Kan jang, Mao-Bushi-Saishin-to, MSSM-002, Ninjin-youei-to, PG201, Protec, Qing-huo-bai-du-yin, Qingfu Guanjieshu, Sambucol Active Defense, Seng-fu-tang, Shin-Xiao-Xiang, Tien Hsien, Thuja formula, Unkei-to, Vigconic, Wheeze-relief-formula, Xia-Bai-San, Yangyuk-Sanhwa-Tang, Yi-fey Ruenn-hou, and Yuldahansotang. Of the western based combinations, formulas with Echinacea spp. were common and showed multiple activities. Numerous formulas demonstrated activity on both gene and protein expression. The research demonstrates that the reviewed botanical formulas modulate cytokine activity, although the bulk of the research is in vitro. Therapeutic success using these formulas may be partially due to their effects on cytokines. Further study of phytotherapy on

  3. Francisella tularensis Antioxidants Harness Reactive Oxygen Species to Restrict Macrophage Signaling and Cytokine Production*

    PubMed Central

    Melillo, Amanda A.; Bakshi, Chandra Shekhar; Melendez, J. Andrés

    2010-01-01

    Francisella tularensis is the etiologic agent of the highly infectious animal and human disease tularemia. Its extreme infectivity and virulence are associated with its ability to evade immune detection, which we now link to its robust reactive oxygen species-scavenging capacity. Infection of primary human monocyte-derived macrophages with virulent F. tularensis SchuS4 prevented proinflammatory cytokine production in the presence or absence of IFN-γ compared with infection with the attenuated live vaccine strain. SchuS4 infection also blocked signals required for macrophage cytokine production, including Akt phosphorylation, IκBα degradation, and NF-κB nuclear localization and activation. Concomitant with SchuS4-mediated suppression of Akt phosphorylation was an increase in the levels of the Akt antagonist PTEN. Moreover, SchuS4 prevented the H2O2-dependent oxidative inactivation of PTEN compared with a virulent live vaccine strain. Mutation of catalase (katG) sensitized F. tularensis to H2O2 and enhanced PTEN oxidation, Akt phosphorylation, NF-κB activation, and inflammatory cytokine production. Together, these findings suggest a novel role for bacterial antioxidants in restricting macrophage activation through their ability to preserve phosphatases that temper kinase signaling and proinflammatory cytokine production. PMID:20558723

  4. Effects of rhinovirus species on viral replication and cytokine production

    PubMed Central

    Nakagome, Kazuyuki; Bochkov, Yury A.; Ashraf, Shamaila; Brockman-Schneider, Rebecca A.; Evans, Michael D.; Pasic, Thomas R.; Gern, James E.

    2014-01-01

    Background Epidemiological studies provide evidence of differential virulence of rhinovirus (RV) species. We recently reported that RV-A and RV-C induced more severe illnesses than RV-B, suggesting that the biology of RV-B might be different from RV-A or RV-C. Objective To test the hypothesis that RV-B has lower replication and induces lesser cytokine responses than RV-A or RV-C. Methods We cloned full-length cDNA of RV-A16, A36, B52, B72, C2, C15 and C41 from clinical samples, and grew clinical isolates of RV-A7 and B6 in cultured cells. Sinus epithelial cells were differentiated at air-liquid interface. We tested for differences in viral replication in epithelial cells after infection with purified viruses (108 RNA copies) and measured virus load by quantitative RT-PCR. We measured lactate dehydrogenase (LDH) concentration as a marker of cellular cytotoxicity, and cytokine/chemokine secretion by multiplex ELISA. Results At 24 hours post infection, virus load of RV-B (RV-B52, B72, or B6) in adherent cells was lower than that of RV-A or RV-C. The growth kinetics of infection indicated that RV-B types replicate more slowly. Furthermore, RV-B released less LDH than RV-A or RV-C, and induced lower levels of cytokines and chemokines such as CXCL10, even after correction for viral replication. RV-B replicates to lower levels also in primary bronchial epithelial cells. Conclusions Our results indicate that RV-B types have lower and slower replication, and lower cellular cytotoxicity and cytokine/chemokine production compared to RV-A or RV-C. These characteristics may contribute to reduced severity of illnesses that has been observed with RV-B infections. Clinical implications RV-B types replicate at a lower rate and produce less cytokine/chemokine compared to RV-A or RV-C, which may contribute to the clinical observation that RV-B causes less severe illnesses. Capsule summary RV-B types replicate more slowly and to lower levels, and less cytokine/chemokine production

  5. Effect of grepafloxacin on cytokine production in vitro.

    PubMed

    Ono, Y; Ohmoto, Y; Ono, K; Sakata, Y; Murata, K

    2000-07-01

    The effect of a new quinolone antibacterial agent, grepafloxacin, on the production of cytokines was investigated using lipopolysaccharide-stimulated human peripheral blood cells. Grepafloxacin 1-30 mg/L inhibited the production of interleukin 1alpha (IL-1alpha) and IL-1beta, and the expression of IL-1alpha, IL-1beta, tumour necrosis factor alpha (TNFalpha), IL-6 and IL-8 mRNA. These results suggest that the inhibitory effect of grepafloxacin is exerted, in part, at the gene transcription level.

  6. A synthetic derivative of the natural product rocaglaol is a potent inhibitor of cytokine-mediated signaling and shows neuroprotective activity in vitro and in animal models of Parkinson's disease and traumatic brain injury.

    PubMed

    Fahrig, T; Gerlach, I; Horváth, E

    2005-05-01

    Many acute and chronic neurodegenerative diseases are characterized by a localized inflammatory response and constitutive activation of the transcription factors nuclear factor-kappa B (NF-kappa B) and activator protein-1 (AP-1) as well as their upstream activating signaling cascades. Ample evidence indicates the implication of these processes in the pathogenesis of several diseases of the central nervous system. In this study, we show that a synthetic derivative of the natural product rocaglaol (compound A) displays potent anti-inflammatory properties in human endothelial and murine glial cells in vitro. Compound A inhibited cytokine- and lipopolysaccharide-induced release of various cytokines/chemokines and of nitric oxide as well as expression of the adhesion molecule endothelial leukocyte adhesion molecule-1 and the inducible enzymes nitric-oxide synthase and cyclooxygenase-2. As shown by immunocytochemistry and immunoblotting, compound A inhibited NF-kappa B and AP-1 activity in mixed glial cultures. Compound A exhibited neuroprotective activity in vitro and in vivo. 1-Methyl-4-phenylpyridinium-induced damage of mesencephalic dopaminergic neurons was significantly decreased, and long-term treatment of 1-methyl-4-phenyl-1,2,3,6,-tetrahydropyridine-injected mice with compound A significantly and dose-dependently reduced dopaminergic neuronal cell death. In addition, shortterm application of compound A to rats suffering from traumatic brain injury induced by subdural hematoma resulted in a significant reduction of the cerebral infarct volume. These results suggest that by inhibiting NF-kappa B and AP-1 signaling, compound A is able to reduce tissue inflammation and neuronal cell death, resulting in significant neuroprotection in animal models of neurodegeneration.

  7. Cytokine production in patients with cirrhosis and TLR4 polymorphisms

    PubMed Central

    Nieto, Juan Camilo; Sánchez, Elisabet; Román, Eva; Vidal, Silvia; Oliva, Laia; Guarner-Argente, Carlos; Poca, Maria; Torras, Xavier; Juárez, Cándido; Guarner, Carlos; Soriano, German

    2014-01-01

    AIM: To analyze the cytokine production by peripheral blood cells from cirrhotic patients with and without TLR4 D299G and/or T399I polymorphisms. METHODS: The study included nine patients with cirrhosis and TLR4 D299G and/or T399I polymorphisms, and 10 wild-type patients matched for age, sex and degree of liver failure. TLR4 polymorphisms were determined by sequence-based genotyping. Cytokine production by peripheral blood cells was assessed spontaneously and also after lipopolysaccharide (LPS) and lipoteichoic acid (LTA) stimulation. RESULTS: Patients with TLR4 polymorphisms had a higher incidence of previous hepatic encephalopathy than wild-type patients (78% vs 20%, P = 0.02). Spontaneous production of interleukin (IL)-6 and IL-10 was lower in patients with TLR4 polymorphisms than in wild-type patients [IL-6: 888.7 (172.0-2119.3) pg/mL vs 5540.4 (1159.2-26053.9) pg/mL, P < 0.001; IL-10: 28.7 (6.5-177.1) pg/mL vs 117.8 (6.5-318.1) pg/mL, P = 0.02]. However, the production of tumor necrosis factor-α, IL-6 and IL-10 after LPS and LTA stimulation was similar in the two groups. CONCLUSION: TLR4 polymorphisms were associated with a distinctive pattern of cytokine production in cirrhotic patients, suggesting that they play a role in the development of cirrhosis complications. PMID:25516666

  8. STK11/LKB1 deficiency promotes neutrophil recruitment and proinflammatory cytokine production to suppress T cell activity in the lung tumor microenvironment

    PubMed Central

    Koyama, Shohei; Akbay, Esra A.; Li, Yvonne Y.; Aref, Amir R.; Skoulidis, Ferdinandos; Herter-Sprie, Grit S.; Buczkowski, Kevin A.; Liu, Yan; Awad, Mark M.; Denning, Warren L.; Diao, Lixia; Wang, Jing; Parra-Cuentas, Edwin R.; Wistuba, Ignacio I.; Soucheray, Margaret; Thai, Tran C.; Asahina, Hajime; Kitajima, Shunsuke; Altabef, Abigail; Cavanaugh, Jillian D.; Rhee, Kevin; Gao, Peng; Zhang, Haikuo; Fecci, Peter E.; Shimamura, Takeshi; Hellmann, Matthew D.; Heymach, John V.; Hodi, F. Stephen; Freeman, Gordon J.; Barbie, David A.; Dranoff, Glenn; Hammerman, Peter S.; Wong, Kwok-Kin

    2016-01-01

    STK11/LKB1 is among the most commonly inactivated tumor suppressors in non-small cell lung cancer (NSCLC), especially in tumors harboring KRAS mutations. Many oncogenes promote immune escape, undermining the effectiveness of immunotherapies, but it is unclear whether inactivation of tumor suppressor genes such as STK11/LKB1 exert similar effects. In this study, we investigated the consequences of STK11/LKB1 loss on the immune microenvironment in a mouse model of KRAS-driven NSCLC. Genetic ablation of STK11/LKB1 resulted in accumulation of neutrophils with T cell suppressive effects, along with a corresponding increase in the expression of T cell exhaustion markers and tumor-promoting cytokines. The number of tumor-infiltrating lymphocytes was also reduced in LKB1-deficient mouse and human tumors. Furthermore, STK11/LKB1 inactivating mutations were associated with reduced expression of PD-1 ligand PD-L1 in mouse and patient tumors as well as in tumor-derived cell lines. Consistent with these results, PD-1 targeting antibodies were ineffective against Lkb1-deficient tumors. In contrast, treating Lkb1-deficient mice with an IL-6 neutralizing antibody or a neutrophil-depleting antibody yielded therapeutic benefits associated with reduced neutrophil accumulation and proinflammatory cytokine expression. Our findings illustrate how tumor suppressor mutations can modulate the immune milieu of the tumor microenvironment, and they offer specific implications for addressing STK11/LKB1 mutated tumors with PD-1 targeting antibody therapies. PMID:26833127

  9. STK11/LKB1 Deficiency Promotes Neutrophil Recruitment and Proinflammatory Cytokine Production to Suppress T-cell Activity in the Lung Tumor Microenvironment.

    PubMed

    Koyama, Shohei; Akbay, Esra A; Li, Yvonne Y; Aref, Amir R; Skoulidis, Ferdinandos; Herter-Sprie, Grit S; Buczkowski, Kevin A; Liu, Yan; Awad, Mark M; Denning, Warren L; Diao, Lixia; Wang, Jing; Parra-Cuentas, Edwin R; Wistuba, Ignacio I; Soucheray, Margaret; Thai, Tran; Asahina, Hajime; Kitajima, Shunsuke; Altabef, Abigail; Cavanaugh, Jillian D; Rhee, Kevin; Gao, Peng; Zhang, Haikuo; Fecci, Peter E; Shimamura, Takeshi; Hellmann, Matthew D; Heymach, John V; Hodi, F Stephen; Freeman, Gordon J; Barbie, David A; Dranoff, Glenn; Hammerman, Peter S; Wong, Kwok-Kin

    2016-03-01

    STK11/LKB1 is among the most commonly inactivated tumor suppressors in non-small cell lung cancer (NSCLC), especially in tumors harboring KRAS mutations. Many oncogenes promote immune escape, undermining the effectiveness of immunotherapies, but it is unclear whether the inactivation of tumor suppressor genes, such as STK11/LKB1, exerts similar effects. In this study, we investigated the consequences of STK11/LKB1 loss on the immune microenvironment in a mouse model of KRAS-driven NSCLC. Genetic ablation of STK11/LKB1 resulted in accumulation of neutrophils with T-cell-suppressive effects, along with a corresponding increase in the expression of T-cell exhaustion markers and tumor-promoting cytokines. The number of tumor-infiltrating lymphocytes was also reduced in LKB1-deficient mouse and human tumors. Furthermore, STK11/LKB1-inactivating mutations were associated with reduced expression of PD-1 ligand PD-L1 in mouse and patient tumors as well as in tumor-derived cell lines. Consistent with these results, PD-1-targeting antibodies were ineffective against Lkb1-deficient tumors. In contrast, treating Lkb1-deficient mice with an IL6-neutralizing antibody or a neutrophil-depleting antibody yielded therapeutic benefits associated with reduced neutrophil accumulation and proinflammatory cytokine expression. Our findings illustrate how tumor suppressor mutations can modulate the immune milieu of the tumor microenvironment, and they offer specific implications for addressing STK11/LKB1-mutated tumors with PD-1-targeting antibody therapies.

  10. Production and function of cytokines in natural and acquired immunity to Candida albicans infection.

    PubMed Central

    Ashman, R B; Papadimitriou, J M

    1995-01-01

    Host resistance against infections caused by the yeast Candida albicans is mediated predominantly by polymorphonuclear leukocytes and macrophages. Antigens of Candida stimulate lymphocyte proliferation and cytokine synthesis, and in both humans and mice, these cytokines enhance the candidacidal functions of the phagocytic cells. In systemic candidiasis in mice, cytokine production has been found to be a function of the CD4+ T helper (Th) cells. The Th1 subset of these cells, characterized by the production of gamma interferon and interleukin-2, is associated with macrophage activation and enhanced resistance against reinfection, whereas the Th2 subset, which produces interleukins-4, -6, and -10, is linked to the development of chronic disease. However, other models have generated divergent data. Mucosal infection generally elicits Th1-type cytokine responses and protection from systemic challenge, and identification of cytokine mRNA present in infected tissues of mice that develop mild or severe lesions does not show pure Th1- or Th2-type responses. Furthermore, antigens of C. albicans, mannan in particular, can induce suppressor cells that modulate both specific and nonspecific cellular and humoral immune responses, and there is an emerging body of evidence that molecular mimicry may affect the efficiency of anti-Candida responses within defined genetic contexts. PMID:8531890

  11. Cytokine Production in the Serum and Spleen of Mice from Day 6 to 14 of Gestation: Cytokines/Placenta/ Spleen/Serum

    PubMed Central

    Iconomidou, Bessy

    1995-01-01

    Pregnancy, like most biologic phenomena, involves the action of cytokines. These proteins have a short half-life and are believed to exert their effect close to their site of production, where diagnostic tests cannot be easily performed. Here we show that the cytokine content in the maternal serum reflects cytokine production and secretion from maternal spleen cells, which also correlates with production from decidual cells. We show that GM-CSF, IL- 3, and IL-10 are present in the serum at specific time intervals during the first half of murine pregnancy, which correlates with their production from maternal spleen cells. Purified GM-CSF and IL-3 from spleen-cell-culture supernatants are biologically active molecules, able to stimulate placental-cell proliferation. Furthermore, TNF-0, which has been identified in many cases of fetal rejection as well as in labor, is shown to be naturally produced during the second half of pregnancy. Additionally, within the limits of the sensitivity of the technique we have used, the detection of IL-4 and the absence of detectable levels of IL- 2 in the maternal serum strongly comforts the hypothesis that pregnancy is a Th2-dependent phenomenon. The results presented in this paper show that the cytokine profile during pregnancy can be monitored by simple blood tests, which may be of relevance both in the followup of a physiological human pregnancy and to the diagnosis of recurrent abortions due to cytokine imbalance. PMID:8924760

  12. Sirtuin inhibition attenuates the production of inflammatory cytokines in lipopolysaccharide-stimulated macrophages

    SciTech Connect

    Fernandes, Claudia A.; Fievez, Laurence; Neyrinck, Audrey M.; Delzenne, Nathalie M.; Bureau, Fabrice; Vanbever, Rita

    2012-04-20

    Highlights: Black-Right-Pointing-Pointer Lipopolysaccharide-stimulated macrophages were treated with cambinol and sirtinol. Black-Right-Pointing-Pointer Cambinol and sirtinol decreased lipopolysaccharide-induced cytokines. Black-Right-Pointing-Pointer Cambinol decreased NF-{kappa}B activity but had no impact on p38 MAPK activation. Black-Right-Pointing-Pointer Sirtuins are an interesting target for the treatment of inflammatory diseases. -- Abstract: In several inflammatory conditions such as rheumatoid arthritis or sepsis, the regulatory mechanisms of inflammation are inefficient and the excessive inflammatory response leads to damage to the host. Sirtuins are class III histone deacetylases that modulate the activity of several transcription factors that are implicated in immune responses. In this study, we evaluated the impact of sirtuin inhibition on the activation of lipopolysaccharide (LPS)-stimulated J774 macrophages by assessing the production of inflammatory cytokines. The pharmacologic inhibition of sirtuins decreased the production of tumour necrosis factor-alpha (TNF-{alpha}) interleukin 6 (IL-6) and Rantes. The reduction of cytokine production was associated with decreased nuclear factor kappa B (NF-{kappa}B) activity and inhibitor kappa B alpha (I{kappa}B{alpha}) phosphorylation while no impact was observed on the phosphorylation status of p38 mitogen-activated kinase (p38 MAPK). This work shows that sirtuin pharmacologic inhibitors are a promising tool for the treatment of inflammatory conditions.

  13. Persistence of local cytokine production in shigellosis in acute and convalescent stages.

    PubMed Central

    Raqib, R; Lindberg, A A; Wretlind, B; Bardhan, P K; Andersson, U; Andersson, J

    1995-01-01

    Shigella infection is accompanied by an intestinal activation of epithelial cells, T cells, and macrophages within the inflamed colonic mucosa. A prospective study was carried out to elucidate the cytokine pattern in Shigella infection linked to development of immunity and eradication of bacteria from the local site and also to correlate the cytokine profile with histological severity. An indirect immunohistochemical technique was used to determine the production and localization of various cytokines at the single-cell level in cryopreserved rectal biopsies from 24 patients with either Shigella dysenteriae type 1 (n = 18) or Shigella flexneri (n = 6) infection. The histopathological profile included presence of chronic inflammatory cells with or without neutrophils and microulcers in the lamina propria, crypt distortion, branching, and less frequently crypt abscesses. Patients had significantly higher (P < 0.005) numbers of cytokine producing cells for all of the cytokines studied, interleukin-1 alpha (IL-1 alpha), IL-1 beta, IL-1ra, tumor necrosis factor alpha (TNF-alpha), IL-6, IL-8, IL-4, IL-10, gamma interferon, TNF-beta, and transforming growth factor beta 1-3, in the biopsies than the healthy controls (n = 13). The cytokine production profile during the study period was dominated by IL-1 beta, transforming growth factor beta 1-3, IL-4, and IL-10. Significantly increased frequencies of cytokine-producing cells (P < 0.05) were observed for IL-1, IL-6, gamma interferon, and TNF-alpha in biopsies with severe inflammation in comparison with those with mild inflammation. During the acute stage of the disease, 20 of 24 patients exhibited acute inflammation in the rectal biopsies and the cellular infiltration was still extensive 30 days after the onset of diarrhea, although the disease was clinically resolved. In accordance with the histological findings, cytokine production was also upregulated during the convalescent phase; there was no significant difference (P

  14. Reduced expression of monocyte CD200R is associated with enhanced proinflammatory cytokine production in sarcoidosis

    PubMed Central

    Fraser, Simon D.; Sadofsky, Laura R.; Kaye, Paul M.; Hart, Simon P.

    2016-01-01

    In sarcoidosis, the proinflammatory cytokines interferon gamma, tumour necrosis factor and interleukin-6 are released by monocyte-derived macrophages and lymphocytes in the lungs and other affected tissues. Regulatory receptors expressed on monocytes and macrophages act to suppress cytokine production, and reduced expression of regulatory receptors may thus promote tissue inflammation. The aim of this study was to characterise the role of regulatory receptors on blood monocytes in patients with sarcoidosis. Cytokine release in response to stimulation of whole blood was measured in healthy controls and Caucasian non-smoking patients with sarcoidosis who were not taking disease modifying therapy. Expression of the regulatory molecules IL-10R, SIRP-α/β, CD47, CD200R, and CD200L was measured by flow cytometry, and functional activity was assessed using blocking antibodies. Stimulated whole blood and monocytes from patients with sarcoidosis produced more TNF and IL-6 compared with healthy controls. 52.9% of sarcoidosis patients had monocytes characterised by low expression of CD200R, compared with 11.7% of controls (p < 0.0001). Patients with low monocyte CD200R expression produced higher levels of proinflammatory cytokines. In functional studies, blocking the CD200 axis increased production of TNF and IL-6. Reduced expression of CD200R on monocytes may be a mechanism contributing to monocyte and macrophage hyper-activation in sarcoidosis. PMID:27929051

  15. Ndfip1 restricts Th17 cell potency by limiting lineage stability and proinflammatory cytokine production

    PubMed Central

    Kesewa Layman, Awo Akosua; L. Sprout, Stephanie; Phillips, Dylan; Oliver, Paula M.

    2017-01-01

    While Th17 cells can protect against colonization by pathogenic organisms, they also have the potential to become pathogenic and promote autoimmune and inflammatory diseases. Mechanisms that control their pathogenic potential remain poorly understood. Here we show that Ndfip1, a co-activator of the E3 ubiquitin ligase Itch, restricts the frequency and pathogenicity of Th17 cells. Mice lacking Ndfip1 have increased numbers of Th17 cells, and this increase is cell intrinsic. We found that Ndfip1 restricts production of the proinflammatory cytokines in Th17 cells. Increased cytokine production correlated with reduced degradation and accumulation of RORγT. When transferred in vivo, Th17 cells lacking Ndfip1 were more likely to maintain their ability to make IL-17, were more potent proinflammatory cytokine producers, and were powerful inducers of colitis. Together our data support an essential role for Ndfip1 in degrading RORγT and suppressing Th17 lineage stability, proinflammatory cytokine production, and pathogenicity. PMID:28051111

  16. Diclofenac enhances proinflammatory cytokine-induced nitric oxide production through NF-{kappa}B signaling in cultured astrocytes

    SciTech Connect

    Kakita, Hiroki; Aoyama, Mineyoshi Hussein, Mohamed Hamed; Kato, Shin; Suzuki, Satoshi; Ito, Tetsuya; Togari, Hajime; Asai, Kiyofumi

    2009-07-01

    Recently, the number of reports of encephalitis/encephalopathy associated with influenza virus has increased. In addition, the use of a non-steroidal anti-inflammatory drug, diclofenac sodium (DCF), is associated with a significant increase in the mortality rate of influenza-associated encephalopathy. Activated astrocytes are a source of nitric oxide (NO), which is largely produced by inducible NO synthase (iNOS) in response to proinflammatory cytokines. Therefore, we investigated whether DCF enhances nitric oxide production in astrocytes stimulated with proinflammatory cytokines. We stimulated cultured rat astrocytes with three cytokines, interleukin-1{beta}, tumor necrosis factor-{alpha} and interferon-{gamma}, and then treated the astrocytes with DCF or acetaminophen (N-acetyl-p-aminophenol: APAP). iNOS and NO production in astrocyte cultures were induced by proinflammatory cytokines. The addition of DCF augmented NO production, but the addition of APAP did not. NF-{kappa}B inhibitors SN50 and MG132 inhibited iNOS gene expression in cytokine-stimulated astrocytes with or without DCF. Similarly, NF-{kappa}B p65 Stealth small interfering RNA suppressed iNOS gene expression in cytokine-stimulated astrocytes with or without DCF. LDH activity and DAPI staining showed that DCF induces cell damage in cytokine-stimulated astrocytes. An iNOS inhibitor, L-NMMA, inhibited the cytokine- and DCF-induced cell damage. In conclusion, this study demonstrates that iNOS and NO are induced in astrocyte cultures by proinflammatory cytokines. Addition of DCF further augments NO production. This effect is mediated via NF-{kappa}B signaling and leads to cell damage. The enhancement of DCF on NO production may explain the significant increase in the mortality rate of influenza-associated encephalopathy in patients treated with DCF.

  17. T-cell immunity and cytokine production in cosmonauts after long-duration space flights

    NASA Astrophysics Data System (ADS)

    Morukov, B.; Rykova, M.; Antropova, E.; Berendeeva, T.; Ponomaryov, S.; Larina, I.

    2011-04-01

    Long-duration spaceflight effects on T-cell immunity and cytokine production were studied in 12 Russian cosmonauts flown onto the International Space Station. Specific assays were performed before launch and after landing and included analysis of peripheral leukocyte distribution, analysis of T-cell phenotype, expression of activation markers, apoptosis, proliferation of T cells in response to a mitogen, concentrations of cytokines in supernatants of cell cultures. Statistically significant increase was observed in leukocytes', lymphocytes', monocytes' and granulocytes' total number, increase in percentage and absolutely number of CD3 +CD4 +-cells, CD4 +CD45RA +-cells and CD4 +CD45RA +/CD4 +CD45RО + ratio, CD4 +CD25 +Bright regulatory cells ( p<0,05) in peripheral blood after landing. T-lymphocytes' capacity to present CD69 and CD25 on its own surfaces was increased for the majority of crewmembers. Analysis of T-cell response to PHA-stimulation in vitro revealed there were some trends toward reduced proliferation of stimulated T-lymphocytes. There was an apparent post flight decrease in secreted IFN-g for the majority of crewmembers and in most instances there was elevation in secreted IL-10. It revealed depression of IFN-g/IL-10 ratio after flight. Correlation analysis according to Spearman's rank correlation test established significant positive correlations ( p<0.05) between cytokine production and T-cell activation (CD25+, CD38+) and negative correlation ( p<0.05) between cytokine production and number of bulk memory CD4+T-cells (CD45RO+). Thus, these results suggest that T-cell dysfunction can be conditioned by cytokine dysbalance and could lead to development of disease after long-duration space flights.

  18. CP-115,953 stimulates cytokine production by lymphocytes.

    PubMed Central

    Riesbeck, K; Forsgren, A

    1995-01-01

    The cytotoxic quinolone CP-115,953 specifically exerts its inhibitory effect upon eukaryotic topoisomerase II. CP-115,953 stimulates DNA cleavage mediated by topoisomerase II with a potency approximately 600 times greater than that of ciprofloxacin, a quinolone antibacterial agent that currently is in clinical use. Because ciprofloxacin has been reported to strongly enhance interleukin-2 production, we considered it important to study the effect of CP-115,953 on interleukin-2 and gamma interferon (IFN-gamma) mRNA and protein expression in mitogen-stimulated human peripheral blood lymphocytes. For comparison, novobiocin and the antineoplastic drug etoposide were also included in the study. CP-115,953 (25 microM) enhanced interleukin-2 mRNA levels up to 8-fold and IFN-gamma mRNA concentrations up to 6.5-fold. In contrast, ciprofloxacin (282 microM) induced mRNAs for interleukin-2 and IFN-gamma up to 20-fold and 7.8-fold, respectively. However, CP-115,953 showed more prolonged kinetics of IFN-gamma mRNA production than ciprofloxacin. At high concentrations (> or = 141 microM), ciprofloxacin was a greater inducer of interleukin-2 production and exhibited a higher level of stimulatory action than CP-115,953 on IFN-gamma synthesis. At low concentrations, however, CP-115,953 (< or = 25 microM) was more potent than ciprofloxacin in inducing interleukin-2 and IFN-gamma synthesis. Etoposide or novobiocin did not influence cytokine mRNA expression. Thus, among the topoisomerase II inhibitors tested, fluoroquinolones are unique in stimulating cytokine synthesis in lymphocyte cultures. PMID:7726518

  19. In Vivo T Cell Signaling Leading to Apoptosis vs. Cytokine Production

    DTIC Science & Technology

    1995-08-17

    ruled out as an explanation for the T cell depletion caused by high dose anti-CD3 antibody administration. The classical complement-mediated mechanism ...known concerning the mechanisms involved in either anti-CD3 antibody-induced cytokine production or subsequent T cell depletion in vivo. Previous...failed to define the mechanism of deletion or to develop adequate techniques for detecting AICD in vivo. A good model for studying T cell activation is

  20. NKG2D receptor regulates human effector T-cell cytokine production

    PubMed Central

    Barber, Amorette

    2011-01-01

    Although innate immune signals shape the activation of naive T cells, it is unclear how innate signals influence effector T-cell function. This study determined the effects of stimulating the NKG2D receptor in conjunction with the TCR on human effector CD8+ T cells. Stimulation of CD8+ T cells through CD3 and NKG2D simultaneously or through a chimeric NKG2D receptor, which consists of NKG2D fused to the intracellular region of CD3ζ, activated β-catenin and increased expression of β-catenin–induced genes, whereas T cells stimulated through the TCR or a combination of the TCR and CD28 did not. Activation by TCR and NKG2D prevented expression and production of anti-inflammatory cytokines IL-10, IL-9, IL-13, and VEGF-α in a β-catenin– and PPARγ- dependent manner. NKG2D stimulation also modulated the cytokine secretion of T cells activated simultaneously through CD3 and CD28. These data indicate that activating CD8+ T cells through the NKG2D receptor along with the TCR modulates signal transduction and the production of anti-inflammatory cytokines. Thus, human effector T cells alter their function depending on which innate receptors are engaged in conjunction with the TCR complex. PMID:21518928

  1. Cytokine dependent and independent iNKT cell activation

    PubMed Central

    Reilly, Emma C.; Wands, Jack R.; Brossay, Laurent

    2010-01-01

    Invariant NKT (iNKT) cells have been extensively studied throughout the last decade due to their ability to polarize and amplify the downstream immune response. Only recently however, have the various mechanisms underlying NKT cell activation begun to unfold. iNKT cells have the ability to respond as innate immune cells with minimal TCR involvement as well as through direct TCR recognition of glycolipid antigens. Additionally, the existence of several subsets of iNKT cells creates the potential for other unique pathways, which are not yet clearly defined. Here we provide an overview of the known mechanisms of invariant NKT cell activation, focusing on cytokine driven pathways and the resulting cytokine responses. PMID:20554220

  2. Naegleria fowleri induces MUC5AC and pro-inflammatory cytokines in human epithelial cells via ROS production and EGFR activation.

    PubMed

    Cervantes-Sandoval, Isaac; Serrano-Luna, José de Jesús; Meza-Cervantez, Patricia; Arroyo, Rossana; Tsutsumi, Víctor; Shibayama, Mineko

    2009-11-01

    Naegleria fowleri is an amoeboflagellate responsible for the fatal central nervous system (CNS) disease primary amoebic meningoencephalitis (PAM). This amoeba gains access to the CNS by invading the olfactory mucosa and crossing the cribriform plate. Studies using a mouse model of infection have shown that the host secretes mucus during the very early stages of infection, and this event is followed by an infiltration of neutrophils into the nasal cavity. In this study, we investigated the role of N. fowleri trophozoites in inducing the expression and secretion of airway mucin and pro-inflammatory mediators. Using the human mucoepidermal cell line NCI-H292, we demonstrated that N. fowleri induced the expression of the MUC5AC gene and protein and the pro-inflammatory mediators interleukin-8 (IL-8) and interleukin-1 beta (IL-1 beta), but not tumour necrosis factor-alpha or chemokine c-c motif ligand 11 (eotaxin). Since the production of reactive oxygen species (ROS) is a common phenomenon involved in the signalling pathways of these molecules, we analysed if trophozoites were capable of causing ROS production in NCI-H292 cells by detecting oxidation of the fluorescent probe 2,7-dichlorofluorescein diacetate. NCI-H292 cells generated ROS after 15-30 min of trophozoite stimulation. Furthermore, the expression of MUC5AC, IL-8 and IL-1 beta was inhibited in the presence of the ROS scavenger DMSO. In addition, the use of an epidermal growth factor receptor inhibitor decreased the expression of MUC5AC and IL-8, but not IL-1 beta. We conclude that N. fowleri induces the expression of some host innate defence mechanisms, such as mucin secretion (MUC5AC) and local inflammation (IL-8 and IL-1 beta) in respiratory epithelial cells via ROS production and suggest that these innate immune mechanisms probably prevent most PAM infections.

  3. Herbal medicine IMOD suppresses LPS-induced production of proinflammatory cytokines in human dendritic cells

    PubMed Central

    Mirzaee, Saeedeh; Drewniak, Agata; Sarrami-Forooshani, Ramin; Kaptein, Tanja M.; Gharibdoost, Farhad; Geijtenbeek, Teunis B. H.

    2015-01-01

    Traditional medicines that stimulate or modulate the immune system can be used as innovative approaches to treat immunological diseases. The herbal medicine IMOD has been shown to strongly modulate immune responses in several animal studies as well as in clinical trials. However, little is known about the mechanisms of IMOD to modulate immunity. Here we have investigated whether IMOD modulates the immunological function of human dendritic cells (DCs). IMOD alone did not induce DC maturation nor production of cytokines. Notably, IMOD decreased the production of pro-inflammatory cytokines IL-6, IL-12 p70, and TNFα by LPS-activated DCs at both mRNA and protein levels in a dose dependent manner. In contrast, treatment with IMOD did not affect LPS induced-production of the anti-inflammatory cytokine IL-10. Furthermore, IMOD inhibited T cell activation/proliferation by LPS-treated DCs and skewed T-cells responses toward the T helper type 2 polarization. These data strongly indicate that IMOD has a potent immunomodulatory ability that affects TLR signaling and thereby modulates DC function. Insight into the immunomodulatory effect of herbal medicine IMOD may provide innovative strategies to affect the immune system and to help combat various diseases. PMID:25870561

  4. Antioxidant activity of a Lachnum YM226 melanin-iron complex and its influence on cytokine production in mice with iron deficiency anemia.

    PubMed

    Song, Sheng; Yang, Liu; Ye, Ming; Chen, Xue; Shi, Fang; Shaikh, Farnaz

    2016-03-01

    The present study aims to investigate the protective effects of an orally administered Lachnum YM226 melanin-iron complex (LM-Fe) against iron deficiency anemia (IDA) in mice. The IDA mouse model was established by feeding mice with iron-deficient food. Different doses of LM-Fe were given to the anaemic mice via intragastric administration, with FeCl3 and FeSO4 used as positive controls. After the iron supplement administration, it was observed that LM-Fe could significantly improve the decreased haemoglobin (Hb) level, and normalize the serum iron (SI) level, total iron-binding capacity (TIBC) and serum ferritin (SF) of the anaemic mice in a dose-dependent manner. In addition, treatment with LM-Fe significantly increased the antioxidant enzyme activities of superoxidase dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSH-Px) in plasma to normal or better. Furthermore, the levels of tumour necrosis factor-α (TNF-α), interleukin-1β (IL-1β) and interleukin-6 (IL-6) were obviously decreased in the LM-Fe supplemented groups compared with the model group, while the level of interleukin-2 (IL-2) was significantly increased. In conclusion, LM-Fe was efficient at ameliorating the anemia symptoms, improving the activities of antioxidant enzymes and adjusting the immune dysfunction of anaemic mice. Thus, these results demonstrated that LM-Fe might be exploited as an efficient and multifunctional iron supplement.

  5. Decreased production of proinflammatory cytokines by monocytes from individuals presenting Candida-associated denture stomatitis.

    PubMed

    Pinke, Karen Henriette; Freitas, Patrícia; Viera, Narciso Almeida; Honório, Heitor Marques; Porto, Vinicius Carvalho; Lara, Vanessa Soares

    2016-01-01

    Candida-associated denture stomatitis (DS) is the most frequent lesion among denture wearers, especially the elderly. DS is strongly associated with Candida albicans, as well as local and systemic factors, such as impaired immune response. Monocytes are important in the protective immune response against the fungus by the production of cytokines that recruit and activate leukocytes. There are functional changes in these cells with age, and individual alterations involving monocyte response may predispose the host to developing infections by Candida spp. In this study, our aim was to evaluate the production of TNF-α, IL-6, CXCL8, IL-1β, MCP-1 and IL-10 by monocytes from elderly denture wearers with/without DS and elderly or young non-denture wearers. We detected that monocytes from elderly denture wearers with Candida-related denture stomatitis produced lower levels of CXCL-8, IL-6 and MCP-1. This imbalance in cytokine levels was observed in spontaneous or LPS-stimulated production. Therefore, our data suggested that inherent aspects of the host, such as changes in cytokine production by monocytes, might be associated with the development and the persistence of DS irrespective of aging.

  6. Photodynamic therapy induced production of cytokines by latent Epstein Barr virus infected epithelial tumor cells

    NASA Astrophysics Data System (ADS)

    Koon, H. K.; Lo, K. W.; Lung, M. L.; Chang, C. K. C.; Wong, R. N. S.; Mak, N. K.

    2007-02-01

    Photodynamic therapy (PDT) is a method to treat cancer or non-cancer diseases by activation of the light-sensitive photosensitizers. Epstein Barr virus (EBV) has been implicated in the development of certain cancers such as nasopharyngeal carcinoma and B cell lymphoma. This study aims to examine the effects of EBV infection on the production of pro-inflammatory cytokines and chemokines in cells after the photosensitizer Zn-BC-AM PDT treatment. Epithelial tumor cell lines HONE-1 and latent EBV-infected HONE-1 (EBV-HONE-1) cells were used in this study. Cells were treated with the photosensitizer Zn-BC-AM for 24 hours before light irradiation. RT-PCR and quantitative ELISA methods were used for the evaluation of mRNA expression and production of cytokines, respectively. Results show that Zn-BC-AM PDT increases the production of IL-1a and IL-1b in EBV-HONE-1. Over a 10-fold increase in the production of IL-6 was observed in the culture supernatant of Zn-BC-AM PDT-treated HONE-1 cells. PDT-induced IL-6 production was observed in HONE-1 cells. EBV-HONE-1 has a higher background level of IL-8 production than the HONE-1. The production of IL-8 was suppressed in EBV-HONE-1cells after Zn-BC-AM PDT. Our results indicate that the response of HONE-1 cells to Zn-BC-AM PDT depends on the presence of latent EBV infection. Since IL-8 is a cytokine with angiogenic activity, Zn-BC-AM PDT may exert an anti-angiogenic effect through the suppression of IL-8 production by the EBV-infected cells.

  7. Selective modulation of lipopolysaccharide-induced death and cytokine production by various muramyl peptides.

    PubMed Central

    Parant, M A; Pouillart, P; Le Contel, C; Parant, F J; Chedid, L A; Bahr, G M

    1995-01-01

    Pretreatment of animals with the adjuvant muramyl dipeptide enhances both the production of circulating tumor necrosis factor and the sensitivity to the lethal effect of a lipopolysaccharide (LPS) challenge. The present study examined the capacity of various adjuvant muramyl dipeptide derivatives to potentiate responsiveness to LPS administration. Cytokine levels in serum were determined at various time intervals after LPS administration by bioassays and immunoassays; the cytokines examined were tumor necrosis factor, interleukin-1, interleukin-6, and gamma interferon. The time course of cytokine response was not modified by the pretreatment, but most of the levels were strongly enhanced. However, of the four compounds which were found to be potent priming agents, only two caused an increased sensitivity to LPS lethality, showing that elevated titers of cytokines in serum were not correlated with host sensitization. Interestingly, previous studies have shown that these two compounds also display neurobiological properties, implying a possible role of the central nervous system in LPS lethality. However, two hydrophilic derivatives with low activity as priming agents were capable of decreasing the toxicity of LPS when given after the challenge in galactosamine-sensitized mice. These results illustrate the diversity of responses elicited by immunological priming. They raise unanswered questions on the importance of endogenous mediators in the pathophysiological alterations during toxic shock. PMID:7806345

  8. Cytokines induce nitric oxide production in mouse osteoblasts.

    PubMed

    Damoulis, P D; Hauschka, P V

    1994-06-15

    MC3T3-E1 mouse clonal osteogenic cells were incubated with interferon-gamma, interleukin-1 beta, tumor necrosis factor-alpha, and E. coli lipopolysaccharide. TNF alpha, IL-1 beta, and LPS caused a dose- and time-dependent increase of nitrite (NO2-), the stable metabolite of nitric oxide (NO), in conditioned media over 48 hours, while IFN gamma had a minimal effect. Different combinations of the same factors caused a synergistic enhancement of NO2- accumulation, except for IL-1 beta with LPS. The earliest detectable NO2- production was at 6-9 hours, with continued accumulation over 48 hours. NO2- production was inhibited dose-dependently by three arginine analogs known to be specific inhibitors of NO synthase, as well as by actinomycin D, cycloheximide, and dexamethasone; EGTA or indomethacin had a small inhibitory effect. It is concluded that osteoblast-like cells can be induced by proinflammatory cytokines and bacterial endotoxin to produce NO, which can play an important role in bone pathophysiology.

  9. The rLrp of Mycobacterium tuberculosis inhibits proinflammatory cytokine production and downregulates APC function in mouse macrophages via a TLR2-mediated PI3K/Akt pathway activation-dependent mechanism.

    PubMed

    Liu, Yuan; Li, Jia-Yun; Chen, Su-Ting; Huang, Hai-Rong; Cai, Hong

    2016-11-01

    We demonstrate that Mycobacterium tuberculosis recombinant leucine-responsive regulatory protein (rLrp) inhibits lipopolysaccharide (LPS)-induced tumor necrosis factor alpha (TNF-α), interleukin-6, and interleukin-12 production and blocks the nuclear translocation of subunits of the nuclear-receptor transcription factor NF-κB (Nuclear factor-kappa B). Moreover, rLrp attenuated LPS-induced DNA binding and NF-κB transcriptional activity, which was accompanied by the degradation of inhibitory IκBα and a consequent decrease in the nuclear translocation of the NF-κB p65 subunit. RLrp interfered with the LPS-induced clustering of TNF receptor-associated factor 6 and with interleukin-1 receptor-associated kinase 1 binding to TAK1. Furthermore, rLrp did not attenuate proinflammatory cytokines or the expression of CD86 and major histocompatibility complex class-II induced by interferon-gamma in the macrophages of Toll-like receptor 2 deletion (TLR2(-/-)) mice and in protein kinase b (Akt)-depleted mouse cells, indicating that the inhibitory effects of rLrp were dependent on TLR2-mediated activation of the phosphatidylinositol 3-OH kinase (PI3K)/Akt pathway. RLrp could also activate the PI3K/Akt pathway by stimulating the rapid phosphorylation of PI3K, Akt, and glycogen synthase kinase 3 beta in macrophages. In addition, 19 amino acid residues in the N-terminus of rLrp were determined to be important and required for the inhibitory effects mediated by TLR2. The inhibitory function of these 19 amino acids of rLrp raises the possibility that mimetic inhibitory peptides could be used to restrict innate immune responses in situations in which prolonged TLR signaling has deleterious effects. Our study offers new insight into the inhibitory mechanisms by which the TLR2-mediated PI3K/Akt pathway ensures the transient expression of potent inflammatory mediators.

  10. Host Intracellular Signaling Events and Pro-inflammatory Cytokine Production in African Trypanosomiasis

    PubMed Central

    Kuriakose, Shiby M.; Singh, Rani; Uzonna, Jude E.

    2016-01-01

    Pathogens, such as bacteria, viruses, and parasites, possess specific molecules or proteins that are recognized by several host innate immune receptors, leading to the activation of several intracellular signaling molecules and pathways. The magnitude and quality of these events significantly affect the outcome of infection. African trypanosomes, including Trypanosoma congolense, are capable of manipulating the host immune response, including the activity of macrophages, which are the key immune cells that contribute to the immunopathogenesis of African trypanosomiasis. Although it is known that immune hyperactivation and excessive pro-inflammatory cytokine production are the hallmarks of African trypanosomiasis, the mechanisms through which these events are triggered are poorly defined. However, it is known that macrophages may play a significant role in these processes, because phagocytosis of trypanosomes by macrophages initiates intracellular signal transduction cascades that lead to the release of pro-inflammatory cytokines and alteration in cell function. This review highlights recent progress in our understanding of the innate immune receptors, signaling pathways, and transcription factors involved in T. congolense-induced pro-inflammatory cytokine production in macrophages. It will reveal the existence of complex signaling events through which the parasite modulates the host immune response, thus identifying novel targets that could aid in designing strategies to effectively control the disease. PMID:27242788

  11. Phagocytic activity and pro-inflammatory cytokines production by the murine macrophage cell line J774A.1 stimulated by a recombinant BCG (rBCG) expressing the MSP1-C of Plasmodium falciparum.

    PubMed

    Rapeah, S; Dhaniah, M; Nurul, A A; Norazmi, M N

    2010-12-01

    Macrophages are involved in innate immunity against malaria due to their ability to phagocytose infected erythrocytes and produce inflammatory cytokines, which are important for controlling parasite growth during malaria infection. In this study, the ability of a recombinant BCG (rBCG) vaccine expressing the 19-kDa C-terminus of merozoite surface protein-1 (MSP1-C) of Plasmodium falciparum, to stimulate the phagocytic activity and secretion of pro-inflammatory cytokines by the macrophage cell line J774A.1 was measured at varying times. The results demonstrate the ability of the rBCG construct to activate the inflammatory action of macrophages, which is important as a first-line of defence in clearing malaria infections.

  12. The pro-inflammatory cytokine, interleukin-6, enhances the polarization of alternatively activated macrophages.

    PubMed

    Fernando, Maria Ruweka; Reyes, Jose Luis; Iannuzzi, Jordan; Leung, Gabriella; McKay, Derek Mark

    2014-01-01

    Macrophages are important innate immune cells that are associated with two distinct phenotypes: a pro-inflammatory (or classically activated) subset with prototypic macrophage functions such as inflammatory cytokine production and bactericidal activity, and an anti-inflammatory (or alternatively activated (AAM)) subset linked with wound healing and tissue repair processes. In this study, we examined the effect of interlukein-6 on human and murine macrophage polarization. The results indicate that despite being commonly associated with pro-inflammatory functions and being implicated in the pathogenesis/pathophysiology of numerous inflammatory diseases, interleukin-6 can enhance the polarization of AAMs, based on increased expression of hallmark markers: arginase-1, Ym1 and CD206; this effect required the AAM differentiating cytokines, IL-4 and IL-13. Co-treatment of AAMs with IL-6 resulted in spontaneous release of IL-10, suppressed LPS-induced nitric oxide production and inhibited cytokine production by activated CD4+ T cells - immunoregulatory features not observed in the 'parent' IL-4+IL-13-induced AAM. The effect of IL-6 required signal transducer and activator of transcription (STAT)-3, was partially dependent on up-regulation of the IL4Rα chain, and was independent of autocrine IL-10. In the presence of IFNγ, IL-6 promoted the production of IL-1β and TNFα suggesting that this cytokine can enhance the phenotype to which a macrophage has committed. This finding may explain the pleiotrophic nature of IL-6, where it is associated with the perpetuation and enhancement of disease in inflammatory situations, but is also necessary for resolution of inflammation and adequate wound healing to occur in others. Thus, the potential benefit of IL-6 in promoting an AAM, with its' anti-inflammatory and wound healing ability, may need to be considered in immunotherapies aimed at in vivo modulation or inhibition of IL-6.

  13. Glycolysis-dependent histone deacetylase 4 degradation regulates inflammatory cytokine production.

    PubMed

    Wang, Bin; Liu, Ting-Yu; Lai, Chun-Hsiang; Rao, Yan-hua; Choi, Moon-Chang; Chi, Jen-Tsan; Dai, Jian-wu; Rathmell, Jeffrey C; Yao, Tso-Pang

    2014-11-01

    Activation of the inflammatory response is accompanied by a metabolic shift to aerobic glycolysis. Here we identify histone deacetylase 4 (HDAC4) as a new component of the immunometabolic program. We show that HDAC4 is required for efficient inflammatory cytokine production activated by lipopolysaccharide (LPS). Surprisingly, prolonged LPS treatment leads to HDAC4 degradation. LPS-induced HDAC4 degradation requires active glycolysis controlled by GSK3β and inducible nitric oxide synthase (iNOS). Inhibition of GSK3β or iNOS suppresses nitric oxide (NO) production, glycolysis, and HDAC4 degradation. We present evidence that sustained glycolysis induced by LPS treatment activates caspase-3, which cleaves HDAC4 and triggers its degradation. Of importance, a caspase-3-resistant mutant HDAC4 escapes LPS-induced degradation and prolongs inflammatory cytokine production. Our findings identify the GSK3β-iNOS-NO axis as a critical signaling cascade that couples inflammation to metabolic reprogramming and a glycolysis-driven negative feedback mechanism that limits inflammatory response by triggering HDAC4 degradation.

  14. SCIMP is a transmembrane non-TIR TLR adaptor that promotes proinflammatory cytokine production from macrophages

    PubMed Central

    Luo, Lin; Bokil, Nilesh J.; Wall, Adam A.; Kapetanovic, Ronan; Lansdaal, Natalie M.; Marceline, Faustine; Burgess, Belinda J.; Tong, Samuel J.; Guo, Zhong; Alexandrov, Kirill; Ross, Ian L.; Hibbs, Margaret L.; Stow, Jennifer L.; Sweet, Matthew J.

    2017-01-01

    Danger signals activate Toll-like receptors (TLRs), thereby initiating inflammatory responses. Canonical TLR signalling, via Toll/Interleukin-1 receptor domain (TIR)-containing adaptors and proinflammatory transcription factors such as NF-κB, occurs in many cell types; however, additional mechanisms are required for specificity of inflammatory responses in innate immune cells. Here we show that SCIMP, an immune-restricted, transmembrane adaptor protein (TRAP), promotes selective proinflammatory cytokine responses by direct modulation of TLR4. SCIMP is a non-TIR-containing adaptor, binding directly to the TLR4-TIR domain in response to lipopolysaccharide. In macrophages, SCIMP is constitutively associated with the Lyn tyrosine kinase, is required for tyrosine phosphorylation of TLR4, and facilitates TLR-inducible production of the proinflammatory cytokines IL-6 and IL-12p40. Point mutations in SCIMP abrogating TLR4 binding also prevent SCIMP-mediated cytokine production. SCIMP is, therefore, an immune-specific TLR adaptor that shapes host defence and inflammation. PMID:28098138

  15. Thioredoxin Ameliorates Cutaneous Inflammation by Regulating the Epithelial Production and Release of Pro-Inflammatory Cytokines

    PubMed Central

    Tian, Hai; Matsuo, Yoshiyuki; Fukunaga, Atsushi; Ono, Ryusuke; Nishigori, Chikako; Yodoi, Junji

    2013-01-01

    Human thioredoxin-1 (TRX) is a 12-kDa protein with redox-active dithiol in the active site -Cys-Gly-Pro-Cys-. It has been demonstrated that systemic administration and transgenic overexpression of TRX ameliorate inflammation in various animal models, but its anti-inflammatory mechanism is not well characterized. We investigated the anti-inflammatory effects of topically applied recombinant human TRX (rhTRX) in a murine irritant contact dermatitis (ICD) induced by croton oil. Topically applied rhTRX was distributed only in the skin tissues under both non-inflammatory and inflammatory conditions, and significantly suppressed the inflammatory response by inhibiting the production of cytokines and chemokines, such as TNF-α, Il-1β, IL-6, CXCL-1, and MCP-1. In an in vitro study, rhTRX also significantly inhibited the formation of cytokines and chemokines produced by keratinocytes after exposure to croton oil and phorbol 12-myristate 13-acetate. These results indicate that TRX prevents skin inflammation via the inhibition of local formation of inflammatory cytokines and chemokines. As a promising new approach, local application of TRX may be useful for the treatment of various skin and mucosal inflammatory disorders. PMID:24058364

  16. Stress system activity, innate and T helper cytokines, and susceptibility to immune-related diseases.

    PubMed

    Calcagni, Emanuele; Elenkov, Ilia

    2006-06-01

    Associations between stress and health outcomes have now been carefully documented, but the mechanisms by which stress specifically influences disease susceptibility and outcome remain poorly understood. Recent evidence indicates that glucocorticoids (GCs) and catecholamines (CAs), the major stress hormones, inhibit systemically IL-12, TNF-alpha, and INF-gamma, but upregulate IL-10, IL-4, and TGF-beta production. Thus, during an immune and inflammatory response, the activation of the stress system, through induction of a Th2 shift may protect the organism from systemic "overshooting" with T helper lymphocyte 1 (Th1)/proinflammatory cytokines. In certain local responses and under certain conditions, however, stress hormones may actually facilitate inflammation, through induction of IL-1, IL-6, IL-8, IL-18, TNF-alpha, and CRP production, and through activation of the corticotropin-releasing hormone (CRH)/substance P(SP)-histamine axis. Autoimmunity, chronic infections, major depression, and atherosclerosis are characterized by a dysregulation of the pro/anti-inflammatory and Th1/Th2 cytokine balance. Thus, hyperactive or hypoactive stress system, and a dysfunctional neuroendocrine-immune interface associated with abnormalities of the "systemic anti-inflammatory feedback" and/or "hyperactivity" of the local proinflammatory factors may contribute to the pathogenesis of these diseases. Conditions that are associated with significant changes in stress system activity, such as acute or chronic stress, cessation of chronic stress, pregnancy and the postpartum period, or rheumatoid arthritis (RA) through modulation of the systemic or local pro/anti-inflammatory and Th1/Th2 cytokine balance, may suppress or potentiate disease activity and/or progression. Thus, stress hormones-induced inhibition or upregulation of innate and Th cytokine production may represent an important mechanism by which stress affects disease susceptibility, activity, and outcome of various immune

  17. Docosahexaenoic diet supplementation, exercise and temperature affect cytokine production by lipopolysaccharide-stimulated mononuclear cells.

    PubMed

    Capó, Xavier; Martorell, Miquel; Sureda, Antoni; Batle, Juan Miguel; Tur, Josep Antoni; Pons, Antoni

    2016-09-01

    Acute exercise induces changes in peripheral mononuclear cells' (PBMCs) capabilities to produce cytokines. The aim was to investigate the effect of docosahexaenoic acid (DHA) diet supplementation on cytokine production, by lipopolysaccharide (LPS)-stimulated PBMCs after exercise, and the in vitro influence of temperature. Fifteen male soccer players were randomly assigned to a placebo or an experimental group. The experimental group consumed an almond-based beverage enriched with DHA (1.16 g DHA/day) for 8 weeks, whereas the placebo group consumed a similar non-enriched beverage. Blood samples were taken before and after the nutritional intervention in basal conditions and 2 h after acute exercise. Nutritional intervention significantly increased the DHA content in erythrocytes only in experimental group (from 34 ± 3.6 to 43 ± 3.6 nmols DHA/10(9) erythrocytes). Exercise significantly increased Toll-like receptor 4 (TLR4) in PBMCs but only in the placebo group (203 %). Exercise also significantly increased IL6, IL8, VEGF, INFγ, TNFα, IL1α, IL1β, MCP1, and EGG production rates by LPS-stimulated PBMCs, and this response was attenuated by DHA supplementation. Temperature but not DHA also affected the pattern of cytokine production increasing IL6, IL8, IL1β, and MCP1 synthesis. The higher change was evidenced in IL1β increasing the production rate at 39.5 °C from 3.19 ± 0.77 to 22.4 ± 6.1 pg/h 10(6) PBMC in placebo and from 2.36 ± 0.11 to 10.6 ± 0.38 pg/h 10(6) PBMC in the supplemented group. The profile of affected cytokines differs between temperature and exercise, suggesting a different PBMC activation pathway. DHA diet supplementation only attenuated cytokine production after exercise and not that induced by temperature.

  18. A proinflammatory cytokine interleukin-32beta promotes the production of an anti-inflammatory cytokine interleukin-10.

    PubMed

    Kang, Jeong-Woo; Choi, Seung-Chul; Cho, Min-Chul; Kim, Hee-Jong; Kim, Jae-Hwa; Lim, Jong-Seok; Kim, Soo-Hyun; Han, Jae-Yong; Yoon, Do-Young

    2009-09-01

    A new proinflammatory cytokine interleukin-32 (IL-32) has six isoforms. Although IL-32 can be detected in sera from patients suffering from Crohn's disease and rheumatoid arthritis, it is unclear which isoforms are involved. To this end, we investigated the functions of the most abundant IL-32beta by generating K562-IL-32beta stable cell lines. This report confirms, using IL-32 small interfering RNA, that IL-32beta induces an anti-inflammatory cytokine IL-10 in K562-IL-32beta cells and U937 promonocytic cells, which express endogenous IL-32beta upon phorbol 12-myristate 13-acetate (PMA) treatment, and monocyte-derived dendritic cells (DC) upon lipopolysaccharide (LPS) treatment. Interleukin-32beta was induced in monocyte-derived macrophages by LPS and in monocyte-derived DC by LPS, poly(I:C), or anti-CD40 antibody, but was not induced by PMA. We showed that IL-32beta expression was increased in a time-dependent manner in monocyte-derived DC upon LPS treatment and peaked at 24 hr. Production of IL-10 was exactly coincident with IL-32beta expression, but IL-1beta and tumour necrosis factor-alpha production peaked at 6 hr after LPS treatment, then steeply declined. Interleukin-12 p40 was induced at 9 hr and gradually increased until 48 hr, at which time IL-32beta and IL-10 were no longer increased. Knock-down of IL-32beta by IL-32 small interfering RNA led to the decrease of IL-10, but the increase of IL-12 in monocyte-derived DC, which means that IL-32beta promotes IL-10 production, but limits IL-12 production. We also showed that IL-10 neutralization increases IL-12, IL-1beta and tumour necrosis factor-alpha production, which implies that IL-10 suppresses such proinflammatory cytokines. Taken together, our results suggest that IL-32beta upregulates the production of an anti-inflammatory cytokine IL-10, and then IL-10 suppresses proinflammatory cytokines.

  19. Cytokine Production Assays Reveal Discriminatory Immune Defects in Adults with Recurrent Infections and Noninfectious Inflammation

    PubMed Central

    van de Veerdonk, Frank L.; Joosten, Leo A. B.; Simon, Anna; van Crevel, Reinout; Kullberg, Bart-Jan; Gyssens, Inge C.; van der Meer, Jos W. M.; van Deuren, Marcel; Netea, Mihai G.

    2014-01-01

    Cytokine production assays have been primarily used in research settings studying novel immunodeficiencies. We sought to determine the diagnostic value of cytokine production assays in patients with recurrent and/or severe infectious diseases (IDs) without known immunodeficiencies and unclassified noninfectious inflammatory disorders (NIIDs). We retrospectively examined cytokine production in whole-blood and peripheral blood mononuclear cell samples from 157 adult patients. A cytokine production rate of <5% of that of healthy controls was considered defective. While monocyte-derived cytokine (tumor necrosis factor alpha [TNF-α], interleukin-1β [IL-1β], and IL-6) production was rarely affected, 30% of all included patients had deficient production of interferon gamma (IFN-γ), IL-17A, or IL-22. Twenty-five percent of the NIID patients displayed defective IFN-γ production, whereas IL-17A production was generally unaffected. In the group of ID patients, defective IFN-γ production was found in 19% and 14% of the patients with viral and bacterial infections, respectively, and in 38%, 24%, and 50% of patients with mycobacterial, mucocutaneous, and invasive fungal infections, respectively. Defective IL-17A and IL-22 production was mainly confined to ID patients with mucocutaneous fungal infections. In conclusion, cytokine production assays frequently detect defective Th1 responses in patients with mycobacterial or fungal infections, in contrast to patients with respiratory tract infections or isolated bacterial infections. Defective IL-17A and IL-22 production was primarily found in patients with fungal infections, while monocyte-derived cytokine production was unaffected. Thus, lymphocyte-derived cytokine production assays are helpful in the diagnostic workup of patients with recurrent infections and suspected immunodeficiencies and have the potential to reveal immune defects that might guide adjunctive immunomodulatory therapy. PMID:24872512

  20. The rLrp of Mycobacterium tuberculosis inhibits proinflammatory cytokine production and downregulates APC function in mouse macrophages via a TLR2-mediated PI3K/Akt pathway activation-dependent mechanism

    PubMed Central

    Liu, Yuan; Li, Jia-Yun; Chen, Su-Ting; Huang, Hai-Rong; Cai, Hong

    2016-01-01

    We demonstrate that Mycobacterium tuberculosis recombinant leucine-responsive regulatory protein (rLrp) inhibits lipopolysaccharide (LPS)-induced tumor necrosis factor alpha (TNF-α), interleukin-6, and interleukin-12 production and blocks the nuclear translocation of subunits of the nuclear-receptor transcription factor NF-κB (Nuclear factor-kappa B). Moreover, rLrp attenuated LPS-induced DNA binding and NF-κB transcriptional activity, which was accompanied by the degradation of inhibitory IκBα and a consequent decrease in the nuclear translocation of the NF-κB p65 subunit. RLrp interfered with the LPS-induced clustering of TNF receptor-associated factor 6 and with interleukin-1 receptor-associated kinase 1 binding to TAK1. Furthermore, rLrp did not attenuate proinflammatory cytokines or the expression of CD86 and major histocompatibility complex class-II induced by interferon-gamma in the macrophages of Toll-like receptor 2 deletion (TLR2−/−) mice and in protein kinase b (Akt)-depleted mouse cells, indicating that the inhibitory effects of rLrp were dependent on TLR2-mediated activation of the phosphatidylinositol 3-OH kinase (PI3K)/Akt pathway. RLrp could also activate the PI3K/Akt pathway by stimulating the rapid phosphorylation of PI3K, Akt, and glycogen synthase kinase 3 beta in macrophages. In addition, 19 amino acid residues in the N-terminus of rLrp were determined to be important and required for the inhibitory effects mediated by TLR2. The inhibitory function of these 19 amino acids of rLrp raises the possibility that mimetic inhibitory peptides could be used to restrict innate immune responses in situations in which prolonged TLR signaling has deleterious effects. Our study offers new insight into the inhibitory mechanisms by which the TLR2-mediated PI3K/Akt pathway ensures the transient expression of potent inflammatory mediators. PMID:26166760

  1. Cytokine treatment of macrophage suppression of T cell activation.

    PubMed

    Silberman, Daniel; Bucknum, Amanda; Kozlowski, Megan; Matlack, Robin; Riggs, James

    2010-01-01

    High Mphi:T cell ratios suppress the immune response to the retroviral superantigen Mls by IFNgamma-triggered production of the arg- and trp-consuming enzymes iNOS and IDO. Attempts to reverse suppression by treatment with pro-inflammatory cytokines revealed that IL-6 improved the T cell response to Mls and the pro-hematopoietic cyokines IL-3 and GM-CSF increased suppression. GM-CSF treatment increased Mphi expression of CD80, a ligand for the immune suppressive B7H1 and CTLA-4 receptors. These results illustrate potential strategies for reversing the suppression of cell-mediated immunity characteristic of the high Mphi:T cell ratios found in many tumors.

  2. Hepatic inflammatory cytokine production can be regulated by modulating sphingomyelinase and ceramide synthase 6.

    PubMed

    Kim, Min Hee; Ahn, Hee Kyung; Lee, Eun-Ji; Kim, Su-Jeong; Kim, Ye-Ryung; Park, Joo-Won; Park, Woo-Jae

    2017-02-01

    Chronic inflammation is associated with the pathogenesis of type 2 diabetes and diabetic complications, and palmitate has been nominated as a candidate for the molecular link between these disorders. Recently, a crucial role of ceramide in inflammation and metabolic diseases has been reported. Therefore, in this study, we investigated whether ceramide formation is involved in palmitate‑induced hepatic inflammation in vitro and in vivo. Ceramide can be generated either by the de novo pathway or by sphingomyelin degradation, and six different ceramide synthases (CerS) determine the specific acyl chain length of ceramide in mammals. We examined the roles of CerS and sphingomyelinases (SMases) in the secretion of inflammatory cytokines, such as tumour necrosis factor (TNF)‑α, interleukin (IL)‑1β, and IL‑6 in Hep3B cells. Among the six CerS, CerS6 overexpression uniquely elevated TNF‑α secretion via p38 mitogen‑activated protein kinase (MAPK) activation. In addition, the treatment of CerS6 overexpressing cells with palmitate synergistically increased cytokine secretion. However, neither palmitate treatment nor CerS6 overexpression altered lipopolysaccharide (LPS)-induced cytokine secretion. Instead, the activation of acidic (A)‑SMase was involved in LPS‑induced cytokine secretion via the MAPK/NF‑κB pathway. Finally, the suppression of ceramide generation via A‑SMase inhibition or de novo ceramide synthesis decreased high‑fat diet‑induced hepatic cytokine production in vivo. On the whole, our results revealed that CerS6 played a role in TNF‑α secretion, and palmitate augmented inflammatory responses in pathophysiological conditions in which CerS6 is overexpressed. In addition, A‑SMase activation was shown to be involved in LPS‑induced inflammatory processes, suggesting that the modulation of CerS6 and A‑SMase may be a therapeutic target for controlling hepatic inflammation.

  3. Antioxidant Defenses of Francisella tularensis Modulate Macrophage Function and Production of Proinflammatory Cytokines.

    PubMed

    Rabadi, Seham M; Sanchez, Belkys C; Varanat, Mrudula; Ma, Zhuo; Catlett, Sally V; Melendez, Juan Andres; Malik, Meenakshi; Bakshi, Chandra Shekhar

    2016-03-04

    Francisella tularensis, the causative agent of a fatal human disease known as tularemia, has been used in the bioweapon programs of several countries in the past, and now it is considered a potential bioterror agent. Extreme infectivity and virulence of F. tularensis is due to its ability to evade immune detection and to suppress the host's innate immune responses. However, Francisella-encoded factors and mechanisms responsible for causing immune suppression are not completely understood. Macrophages and neutrophils generate reactive oxygen species (ROS)/reactive nitrogen species as a defense mechanism for the clearance of phagocytosed microorganisms. ROS serve a dual role; at high concentrations they act as microbicidal effector molecules that destroy intracellular pathogens, and at low concentrations they serve as secondary signaling messengers that regulate the expression of various inflammatory mediators. We hypothesized that the antioxidant defenses of F. tularensis maintain redox homeostasis in infected macrophages to prevent activation of redox-sensitive signaling components that ultimately result in suppression of pro-inflammatory cytokine production and macrophage microbicidal activity. We demonstrate that antioxidant enzymes of F. tularensis prevent the activation of redox-sensitive MAPK signaling components, NF-κB signaling, and the production of pro-inflammatory cytokines by inhibiting the accumulation of ROS in infected macrophages. We also report that F. tularensis inhibits ROS-dependent autophagy to promote its intramacrophage survival. Collectively, this study reveals novel pathogenic mechanisms adopted by F. tularensis to modulate macrophage innate immune functions to create an environment permissive for its intracellular survival and growth.

  4. SN79, a sigma receptor ligand, blocks methamphetamine-induced microglial activation and cytokine upregulation.

    PubMed

    Robson, Matthew J; Turner, Ryan C; Naser, Zachary J; McCurdy, Christopher R; Huber, Jason D; Matsumoto, Rae R

    2013-09-01

    Methamphetamine (METH) abuse is associated with several negative side effects including neurotoxicity in specific brain regions such as the striatum. The precise molecular mechanisms by which METH usage results in neurotoxicity remain to be fully elucidated, with recent evidence implicating the importance of microglial activation and neuroinflammation in damaged brain regions. METH interacts with sigma receptors which are found in glial cells in addition to neurons. Moreover, sigma receptor antagonists have been shown to block METH-induced neurotoxicity in rodents although the cellular mechanisms underlying their neuroprotection remain unknown. The purpose of the current study was to determine if the prototypic sigma receptor antagonist, SN79, mitigates METH-induced microglial activation and associated increases in cytokine expression in a rodent model of METH-induced neurotoxicity. METH increased striatal mRNA and protein levels of cluster of differentiation 68 (CD68), indicative of microglial activation. METH also increased ionized calcium binding adapter molecule 1 (IBA-1) protein expression, further confirming the activation of microglia. Along with microglial activation, METH increased striatal mRNA expression levels of IL-6 family pro-inflammatory cytokines, leukemia inhibitory factor (lif), oncostatin m (osm), and interleukin-6 (il-6). Pretreatment with SN79 reduced METH-induced increases in CD68 and IBA-1 expression, demonstrating its ability to prevent microglial activation. SN79 also attenuated METH-induced mRNA increases in IL-6 pro-inflammatory cytokine family members. The ability of a sigma receptor antagonist to block METH-induced microglial activation and cytokine production provides a novel mechanism through which the neurotoxic effects of METH may be mitigated.

  5. Caspase 3 inactivates biologically active full length interleukin-33 as a classical cytokine but does not prohibit nuclear translocation

    SciTech Connect

    Ali, Shafaqat; Nguyen, Dang Quan; Falk, Werner; Martin, Michael Uwe

    2010-01-15

    IL-33 is a member of the IL-1 family of cytokines with dual function which either activates cells via the IL-33 receptor in a paracrine fashion or translocates to the nucleus to regulate gene transcription in an intracrine manner. We show that full length murine IL-33 is active as a cytokine and that it is not processed by caspase 1 to mature IL-33 but instead cleaved by caspase 3 at aa175 to yield two products which are both unable to bind to the IL-33 receptor. Full length IL-33 and its N-terminal caspase 3 breakdown product, however, translocate to the nucleus. Finally, bioactive IL-33 is not released by cells constitutively or after activation. This suggests that IL-33 is not a classical cytokine but exerts its function in the nucleus of intact cells and only activates others cells via its receptor as an alarm mediator after destruction of the producing cell.

  6. ALPK1 affects testosterone mediated regulation of proinflammatory cytokines production.

    PubMed

    Kuo, Tzer-Min; Yeh, Kun-Tu; Hsu, Hui-Ting; Chiang, Shang-Lun; Chang, Jan-Gowth; Huang, Chung-Ming; Tu, Hung-Pin; Liu, Chiu-Shong; Ko, Ying-Chin

    2015-11-01

    Alpha-protein kinase 1, also known as alpha-kinase 1 (ALPK1), is associated with chronic kidney disease (CKD), myocardial infarction, gout and type 2 diabetes mellitus (DM). In addition to having an inductive effect on the proinflammatory cytokines in monocytic THP1 cells, ALPK1 is expressed abundantly in the mouse testes. Low testosterone levels are commonly associated with arthritis, CKD, type 2 DM, cardiovascular disease and inflammation. The testosterone's anti-inflammatory effect has been demonstrated to reduce proinflammatory cytokines and adhesion molecules. In this study, we found that ALPK1 transgenic mice showed lower levels of testosterone in both the testes and the serum. Decreasing endogenous ALPK1 enhanced testosterone levels and transcripts of testosterone-regulated genes (P450scc, 3beta-HSD, P450C17, 17beta-HSD, StAR, and INSL3) in TM3 Leydig cells. In contrast, increasing testosterone decreased ALPK1 in both TM3 and monocytic THP1 cells. This decrease was accompanied by a reduction of the proinflammatory cytokines. Increased ALPK1 levels attenuated the testosterone effects in THP1 cells. Finally, we also found that ALPK1 increased the release of TNF-alpha and TGF-beta1 in the human embryonic kidney 293 cells, while testosterone inhibited ALPK1 in the primary kidney cells. Taken together, this data suggests that the balance between ALPK1 and testosterone plays a critical role in the testosterone-mediated inhibition of proinflammatory cytokines.

  7. Phototherapy-treated apoptotic tumor cells induce pro-inflammatory cytokines production in macrophage

    NASA Astrophysics Data System (ADS)

    Lu, Cuixia; Wei, Yanchun; Xing, Da

    2014-09-01

    Our previous studies have demonstrated that as a mitochondria-targeting cancer phototherapy, high fluence low-power laser irradiation (HF-LPLI) induces mitochondrial superoxide anion burst, resulting in oxidative damage to tumor cells. In this study, we further explored the immunological effects of HF-LPLI-induced apoptotic tumor cells. When macrophages were co-incubated with apoptotic cells induced by HF-LPLI, we observed the increased levels of TNF-α secretion and NO production in macrophages. Further experiments showed that NF-κB was activated in macrophages after co-incubation with HF-LPLI-induced apoptotic cells, and inhibition of NF-κB activity by pyrrolidinedithiocarbamic acid (PDTC) reduced the elevated levels of TNF-α secretion and NO production. These data indicate that HF-LPLI-induced apoptotic tumor cells induce the secretion of pro-inflammatory cytokines in macrophages, which may be helpful for better understanding the biological effects of cancer phototherapy.

  8. Gomisin N Decreases Inflammatory Cytokine Production in Human Periodontal Ligament Cells.

    PubMed

    Hosokawa, Yoshitaka; Hosokawa, Ikuko; Shindo, Satoru; Ozaki, Kazumi; Matsuo, Takashi

    2017-04-01

    Gomisin N, which is a lignan isolated from Schisandra chinensis, has some pharmacological effects. However, the anti-inflammatory effects of gomisin N on periodontal disease are uncertain. The aim of this study was to examine the effect of gomisin N on inflammatory mediator production in tumor necrosis factor (TNF)-α-stimulated human periodontal ligament cells (HPDLC). Gomisin N inhibited interleukin (IL)-6, IL-8, CC chemokine ligand (CCL) 2, and CCL20 production in TNF-α-stimulated HPDLC in a dose-dependent manner. Moreover, we revealed that gomisin N could suppress extracellular signal-regulated kinase (ERK) and c-Jun N terminal kinase (JNK) phosphorylation in TNF-α-stimulated HPDLC though protein kinase B (Akt) phosphorylation was not suppressed by gomisin N treatment. In summary, gomisin N might exert anti-inflammatory effects by attenuating cytokine production in periodontal ligament cells via inhibiting the TNF-α-stimulated ERK and JNK pathways activation.

  9. Glia activation and cytokine increase in rat hippocampus by kainic acid-induced status epilepticus during postnatal development.

    PubMed

    Rizzi, Massimo; Perego, Carlo; Aliprandi, Marisa; Richichi, Cristina; Ravizza, Teresa; Colella, Daniele; Velískŏvá, Jana; Moshé, Solomon L; De Simoni, M Grazia; Vezzani, Annamaria

    2003-12-01

    In adult rats, status epilepticus (SE) induces cytokine production by glia especially when seizures are associated with neuronal injury. This suggests that cytokines may play a role in seizure-induced neuronal damage. As SE-induced injury is age-specific, we used rats of different ages (with distinct susceptibilities to seizure-induced neuronal injury) to elucidate the role of cytokines in this process. Thus, we investigated the activation of microglia and astrocytes, induction of cytokines, and hippocampal neuronal injury 4 and 24 h following kainic acid-induced SE in postnatal day (PN) 9, 15, and 21 rats. At PN9, there was little activation of microglia and astrocytes at any time point studied. Interleukin-1beta (IL), tumor necrosis factor-alpha (TNF), and IL-6 or the naturally occurring IL-1 receptor antagonist (Ra) mRNA expression did not increase. No evidence of cell injury has been detected. At PN15, immunostaining of microglia and astrocytes was enhanced, but only IL-1beta mRNA expression was increased. These changes were observed 4 h after SE. Scattered injured neurons in CA3 and subiculum, but not in any other region, were present 24 h following SE. At PN21, immunostaining of microglia and astrocytes and the mRNA expression of all cytokines studied was significantly increased already 4 h after SE. At 24 h, many injured neurons were present in CA1 and CA3 regions and in 40% of rats in other forebrain areas. These data show that (i) the pattern of glia activation and cytokine gene transcription induced by SE is age-dependent and (ii) neuronal injury in the hippocampus occurs only when cytokines are induced and their synthesis precedes the appearance of neuronal damage. Thus, cytokine expression in immature brain is associated specifically with cell injury rather than with seizures per se, suggesting that proinflammatory cytokines may contribute to the occurence of SE-induced hippocampal damage.

  10. Necroptosis suppresses inflammation via termination of TNF- or LPS-induced cytokine and chemokine production.

    PubMed

    Kearney, C J; Cullen, S P; Tynan, G A; Henry, C M; Clancy, D; Lavelle, E C; Martin, S J

    2015-08-01

    TNF promotes a regulated form of necrosis, called necroptosis, upon inhibition of caspase activity in cells expressing RIPK3. Because necrosis is generally more pro-inflammatory than apoptosis, it is widely presumed that TNF-induced necroptosis may be detrimental in vivo due to excessive inflammation. However, because TNF is intrinsically highly pro-inflammatory, due to its ability to trigger the production of multiple cytokines and chemokines, rapid cell death via necroptosis may blunt rather than enhance TNF-induced inflammation. Here we show that TNF-induced necroptosis potently suppressed the production of multiple TNF-induced pro-inflammatory factors due to RIPK3-dependent cell death. Similarly, necroptosis also suppressed LPS-induced pro-inflammatory cytokine production. Consistent with these observations, supernatants from TNF-stimulated cells were more pro-inflammatory than those from TNF-induced necroptotic cells in vivo. Thus necroptosis attenuates TNF- and LPS-driven inflammation, which may benefit intracellular pathogens that evoke this mode of cell death by suppressing host immune responses.

  11. Fucoidan modulates cytokine production and migration of THP‑1‑derived macrophages via colony‑stimulating factor‑1.

    PubMed

    Li, Peng; Wang, Huayang; Shao, Qianqian; Kong, Beihua; Qu, Xun

    2017-04-01

    Fucoidan is known for its various biological activities, including immunomodulatory effects on immune cells. However, the effect of fucoidan on the functions of macrophages remains to be elucidated. The present study examined the effects of fucoidan on cytokine production and migration of THP‑1‑derived macrophages and its potential mechanisms. Fucoidan was added during the differentiation process of THP‑1‑derived macrophages along with lipopolysaccharide and interferon‑γ for 42 h, and then macrophages were harvested for functional assays. Fucoidan altered the morphology of THP‑1‑derived macrophages, and also attenuated their migration activity and pro‑inflammatory cytokine production. Additionally, THP‑1‑derived macrophages intensively produced colony‑stimulating factor‑1 (CSF‑1), which was significantly decreased by fucoidan. CSF‑1 neutralizing antibody attenuated the basic production level of pro‑inflammatory cytokines in macrophages. Furthermore, when recombinant human CSF‑1 was added along with fucoidan, the attenuating effects of fucoidan on migration and cytokine production were significantly reversed. In conclusion, the present study suggests that macrophages appear to be a potential target in the immunomodulatory action of fucoidan, and CSF‑1 may be involved in this modulation.

  12. Borrelia burgdorferi Induces the Production and Release of Proinflammatory Cytokines in Canine Synovial Explant Cultures

    PubMed Central

    Straubinger, Reinhard K.; Straubinger, Alix F.; Summers, Brian A.; Erb, Hollis N.; Härter, Luc; Appel, Max J. G.

    1998-01-01

    Canine synovial membrane explants were exposed to high- or low-passage Borrelia burgdorferi for 3, 6, 12, and 24 h. Spirochetes received no treatment, were UV light irradiated for 16 h, or were sonicated prior to addition to synovial explant cultures. In explant tissues, mRNA levels for the proinflammatory cytokines tumor necrosis factor alpha (TNF-α), interleukin-1α (IL-1α), IL-1β, and IL-8 were surveyed semiquantitatively by reverse transcription-PCR. Culture supernatants were examined for numbers of total and motile (i.e., viable) spirochetes, TNF-like and IL-1-like activities, polymorphonuclear neutrophil (PMN) chemotaxis-inducing activities, and IL-8. During exposure to synovial explant tissues, the total number of spirochetes in the supernatants decreased gradually by ∼30%, and the viability also declined. mRNAs for TNF-α, IL-1α, IL-1β, and IL-8 were up-regulated in synovial explant tissues within 3 h after infection with untreated or UV light-irradiated B. burgdorferi, and mRNA levels corresponded to the results obtained with bioassays. During 24 h of coincubation, cultures challenged with untreated or UV light-irradiated spirochetes produced similar levels of TNF-like and IL-1-like activities. In contrast, explant tissues exposed to untreated B. burgdorferi generated significantly higher levels of chemotactic factors after 24 h of incubation than did explant tissues exposed to UV light-treated spirochetes. In identical samples, a specific signal for IL-8 was identified by Western blot analysis. High- and low-passage borreliae did not differ in their abilities to induce proinflammatory cytokines. No difference in cytokine induction between untreated and sonicated high-passage spirochetes was observed, suggesting that fractions of the organism can trigger the production and release of inflammatory mediators. The titration of spirochetes revealed a dose-independent cytokine response, where 103 to 107 B. burgdorferi organisms induced similar TNF

  13. Active immunization by a dengue virus-induced cytokine.

    PubMed Central

    Chaturvedi, U C; Mukerjee, R; Dhawan, R

    1994-01-01

    Dengue type 2 virus (DV)-induced cytotoxic factor (CF) is capable of reproducing various pathological lesions in mice that are seen in human dengue. The present study was undertaken to investigate the protective effect of active immunization of mice with CF. Mice were immunized with 5 microgram of CF and prevention of CF-induced increase in capillary permeability and damage to the blood-brain barrier were studied at weekly intervals, up to 48 weeks, by challenging with 3 microgram of CF. Maximum protection against increase in capillary permeability and damage to the blood-brain barrier was observed in week 4 after immunization. A breakthrough in the protection occurred with higher doses of CF in a dose-dependent manner. Challenge with a lethal intracerebral (i.c.) dose of DV showed significantly prolonged mean survival time and delayed onset of symptoms of sickness in the immunized mice compared with the normal mice, but the titre of the virus in the brain was similar in the two groups. On i.p. challenge with the virus the protection against damage to the blood-brain barrier was 86 +/- 7% at week 4 and 17 +/- 4% at week 26 after immunization. Sera obtained from the immunized mice showed the presence of CF-specific antibodies by ELISA, Western blot, and by neutralization of the cytotoxic activity of CF in vitro. The present study describes successful prevention of a cytokine-induced pathology by specific active immunization. PMID:8187327

  14. INDUCTION OF CYTOKINE PRODUCTION IN CHEETAH (ACINONYX JUBATUS) PERIPHERAL BLOOD MONONUCLEAR CELLS AND VALIDATION OF FELINE-SPECIFIC CYTOKINE ASSAYS FOR ANALYSIS OF CHEETAH SERUM.

    PubMed

    Franklin, Ashley D; Crosier, Adrienne E; Vansandt, Lindsey M; Mattson, Elliot; Xiao, Zhengguo

    2015-06-01

    Peripheral blood mononuclear cells (PBMCs) were isolated from the whole blood of cheetahs (Acinonyx jubatus ; n=3) and stimulated with lipopolysaccharides (LPS) to induce the production of proinflammatory cytokines TNF-α, IL-1β, and IL-6 for establishment of cross-reactivity between these cheetah cytokines and feline-specific cytokine antibodies provided in commercially available Feline DuoSet® ELISA kits (R&D Systems, Inc., Minneapolis, Minnesota 55413, USA). This study found that feline-specific cytokine antibodies bind specifically to cheetah proinflammatory cytokines TNF-α, IL-1β, and IL-6 from cell culture supernatants. The assays also revealed that cheetah PBMCs produce a measurable, cell concentration-dependent increase in proinflammatory cytokine production after LPS stimulation. To enable the use of these kits, which are designed for cell culture supernatants for analyzing cytokine concentrations in cheetah serum, percent recovery and parallelism of feline cytokine standards in cheetah serum were also evaluated. Cytokine concentrations in cheetah serum were approximated based on the use of domestic cat standards in the absence of cheetah standard material. In all cases (for cytokines TNF-α, IL-1β, and IL-6), percent recovery increased as the serum sample dilution increased, though percent recovery varied between cytokines at a given dilution factor. A 1:2 dilution of serum resulted in approximately 45, 82, and 7% recovery of TNF-α, IL-1β, and IL-6 standards, respectively. Adequate parallelism was observed across a large range of cytokine concentrations for TNF-α and IL-1β; however, a significant departure from parallelism was observed between the IL-6 standard and the serum samples (P=0.004). Therefore, based on our results, the Feline DuoSet ELISA (R&D Systems, Inc.) kits are valid assays for the measurement of TNF-α and IL-1β in cheetah serum but should not be used for accurate measurement of IL-6.

  15. Multiple effects of TRAIL in human carcinoma cells: Induction of apoptosis, senescence, proliferation, and cytokine production

    SciTech Connect

    Levina, Vera; Marrangoni, Adele M.; DeMarco, Richard; Gorelik, Elieser; Lokshin, Anna E.

    2008-04-15

    TRAIL is a death ligand that induces apoptosis in malignant but not normal cells. Recently the ability of TRAIL to induce proliferation in apoptosis-resistant normal and malignant cells was reported. In this study, we analyzed TRAIL effects in apoptosis sensitive MCF7, OVCAR3 and H460 human tumor cell lines. TRAIL at low concentrations preferentially induced cell proliferation. At 100 ng/ml, apoptotic death was readily observed, however surviving cells acquired higher proliferative capacity. TRAIL-stimulated production of several cytokines, IL-8, RANTES, MCP-1 and bFGF, and activation of caspases 1 and 8 was essential for this effect. Antibodies to IL-8, RANTES, and bFGF blocked TRAIL-induced cell proliferation and further stimulated apoptosis. For the first time, we report that high TRAIL concentrations induced cell senescence as determined by the altered morphology and expression of several senescence markers: SA-{beta}-gal, p21{sup Waf1/Cip1}, p16{sup INK4a}, and HMGA. Caspase 9 inhibition protected TRAIL-treated cells from senescence, whereas inhibition of caspases 1 and 8 increased the yield of SLP cells. In conclusion, in cultured human carcinoma cells, TRAIL therapy results in three functional outcomes, apoptosis, proliferation and senescence. TRAIL-induced proapoptotic and prosurvival responses correlate with the strength of signaling. TRAIL-induced cytokine production is responsible for its proliferative and prosurvival effects.

  16. The role of stress mediators in modulation of cytokine production by ethanol

    SciTech Connect

    Glover, Mitzi; Cheng Bing; Fan Ruping; Pruett, Stephen

    2009-08-15

    Acute ethanol exposure in humans and in animal models activates the hypothalamic-pituitary-adrenal (HPA) axis and the sympathetic nervous system (SNS); the resultant increases in concentration of neuroendocrine mediators contribute to some of the immunosuppressive effects of ethanol. However, the role of these mediators in the ethanol-induced inhibition of inflammatory responses is not clear. This is complicated by the fact that most inflammatory stimuli also activate the HPA axis and SNS, and it has not been determined if ethanol plus an inflammatory stimulus increases these stress responses. Addressing this issue is the major focus of the study described herein. Complementary approaches were used, including quantitative assessment of the stress response in mice treated with polyinosinic-polycytidylic acid (poly I:C, as an inflammatory stimulus) and inhibition of the production or action of key HPA axis and SNS mediators. Treatment of mice with ethanol shortly before treatment with poly I:C yielded a significant increase in the corticosterone response as compared to the response to poly I:C alone, but the increase was small and not likely sufficient to account for the anti-inflammatory effects of ethanol. Inhibition of catecholamine and glucocorticoid production by adrenalectomy, and inhibition of catecholamine action with a sustained release antagonist (nadalol) supported this conclusion and revealed that 'excess' stress responses associated with ethanol treatment is not the mechanism of suppression of pro-inflammatory cytokine production, but stress-induced corticosterone does regulate production of several of these cytokines, which has not previously been reported.

  17. The cytolethal distending toxin of Aggregatibacter actinomycetemcomitans inhibits macrophage phagocytosis and subverts cytokine production.

    PubMed

    Ando-Suguimoto, Ellen Sayuri; da Silva, Maike Paulino; Kawamoto, Dione; Chen, Casey; DiRienzo, Joseph M; Mayer, Marcia Pinto Alves

    2014-03-01

    Aggregatibacter actinomycetemcomitans is an important periodontal pathogen that can participate in periodontitis and other non-oral infections. The cytolethal distending toxin (Cdt) is among the virulence factors produced by this bacterium. The Cdt is also secreted by several mucosa-associated Gram-negative pathogens and may play a role in perpetuating the infection by modulating the immune response. Although the toxin targets a wide range of eukaryotic cell types little is known about its activity on macrophages which play a key part in alerting the rest of the immune system to the presence of pathogens and their virulence factors. In view of this, we tested the hypothesis that the A. actinomycetemcomitans Cdt (AaCdt) disrupts macrophage function by inhibiting phagocytic activity as well as affecting the production of cytokines. Murine macrophages were co-cultured with either wild-type A. actinomycetemcomitans or a Cdt(-) mutant. Viable counts and qPCR showed that phagocytosis of the wild-type strain was significantly reduced relative to that of the Cdt(-) mutant. Addition of recombinant Aa(r)Cdt to co-cultures along with the Cdt(-) mutant diminished the phagocytic activity similar to that observed with the wild type strain. High concentrations of Aa(r)Cdt resulted in decreased phagocytosis of fluorescent bioparticles. Nitric oxide production was modulated by the presence of Cdt and the levels of IL-1β, IL-12 and IL-10 were increased. Production of TNF-α did not differ in the co-culture assays but was increased by the presence of Aa(r)Cdt. These data suggest that the Cdt may modulate macrophage function in A. actinomycetemcomitans infected sites by impairing phagocytosis and modifying the pro-inflammatory/anti-inflammatory cytokine balance.

  18. MTMR3 risk allele enhances innate receptor-induced signaling and cytokines by decreasing autophagy and increasing caspase-1 activation.

    PubMed

    Lahiri, Amit; Hedl, Matija; Abraham, Clara

    2015-08-18

    Inflammatory bowel disease (IBD) is characterized by dysregulated host:microbial interactions and cytokine production. Host pattern recognition receptors (PRRs) are critical in regulating these interactions. Multiple genetic loci are associated with IBD, but altered functions for most, including in the rs713875 MTMR3/HORMAD2/LIF/OSM region, are unknown. We identified a previously undefined role for myotubularin-related protein 3 (MTMR3) in amplifying PRR-induced cytokine secretion in human macrophages and defined MTMR3-initiated mechanisms contributing to this amplification. MTMR3 decreased PRR-induced phosphatidylinositol 3-phosphate (PtdIns3P) and autophagy levels, thereby increasing PRR-induced caspase-1 activation, autocrine IL-1β secretion, NFκB signaling, and, ultimately, overall cytokine secretion. This MTMR3-mediated regulation required the N-terminal pleckstrin homology-GRAM domain and Cys413 within the phosphatase domain of MTMR3. In MTMR3-deficient macrophages, reducing the enhanced autophagy or restoring NFκB signaling rescued PRR-induced cytokines. Macrophages from rs713875 CC IBD risk carriers demonstrated increased MTMR3 expression and, in turn, decreased PRR-induced PtdIns3P and autophagy and increased PRR-induced caspase-1 activation, signaling, and cytokine secretion. Thus, the rs713875 IBD risk polymorphism increases MTMR3 expression, which modulates PRR-induced outcomes, ultimately leading to enhanced PRR-induced cytokines.

  19. DMSO Represses Inflammatory Cytokine Production from Human Blood Cells and Reduces Autoimmune Arthritis

    PubMed Central

    Elisia, Ingrid; Nakamura, Hisae; Lam, Vivian; Hofs, Elyse; Cederberg, Rachel; Cait, Jessica; Hughes, Michael R.; Lee, Leora; Jia, William; Adomat, Hans H.; Guns, Emma S.; McNagny, Kelly M.; Samudio, Ismael; Krystal, Gerald

    2016-01-01

    Dimethyl sulfoxide (DMSO) is currently used as an alternative treatment for various inflammatory conditions as well as for cancer. Despite its widespread use, there is a paucity of data regarding its safety and efficacy as well as its mechanism of action in human cells. Herein, we demonstrate that DMSO has ex-vivo anti-inflammatory activity using Escherichia coli- (E. coli) and herpes simplex virus-1 (HSV-1)-stimulated whole human blood. Specifically, we found that between 0.5%– 2%, DMSO significantly suppressed the expression of many pro-inflammatory cytokines/chemokines and prostaglandin E2 (PGE2). However, a significant reduction in monocyte viability was also observed at 2% DMSO, suggesting a narrow window of efficacy. Anti-inflammatory concentrations of DMSO suppressed E. coli-induced ERK1/2, p38, JNK and Akt phosphorylation, suggesting DMSO acts on these signaling pathways to suppress inflammatory cytokine/chemokine production. Although DMSO induces the differentiation of B16/F10 melanoma cells in vitro, topical administration of DMSO to mice subcutaneously implanted with B16 melanoma cells was ineffective at reducing tumor growth, DMSO was also found to block mouse macrophages from polarizing to either an M1- or an M2-phenotype, which may contribute to its inability to slow tumor growth. Topical administration of DMSO, however, significantly mitigated K/BxN serum-induced arthritis in mice, and this was associated with reduced levels of pro-inflammatory cytokines in the joints and white blood cell levels in the blood. Thus, while we cannot confirm the efficacy of DMSO as an anti-cancer agent, the use of DMSO in arthritis warrants further investigation to ascertain its therapeutic potential. PMID:27031833

  20. Cytokine production of the neutrophils and macrophages in time of phagocytosis under influence of infrared low-level laser irradiation

    NASA Astrophysics Data System (ADS)

    Rudik, Dmitry V.; Tikhomirova, Elena I.; Tuchina, Elena S.

    2006-08-01

    Influence of infrared low-level laser irradiation (LLLI) on induction of synthesis of some cytokines such as interleykin-1 (Il-1), tumor necrosis factor-α (TNF-α), interferon-γ (INF-γ), interleykin-8 (Il-8) and interleykin-4 (Il-4) by the neutrophils and macrophages in time of bacterial cells phagocytosis that was searched. As the object of analysis we used peritoneal macrophages from white mice and neutrophils from peripheral blood of healthy donors. We used the laser diod with spectrum maximum of 850 nm with doses 300, 900 and 1500 mJ (exposition -60, 180 and 300 s respectively; capacity - 5 mW). We carried out the Enzyme-Linked Immunospot Assay (ELISA) to determine cytokine content during phagocytosis after 3 h and 6 h. We found dynamics in production of the cytokines, which was different for the neutrophils and macrophages. We showed that the infrared LLLI has significant stimulating activity on the proinflammatory cytokines production by neutrophils and macrophages. Moreover we revealed dynamics changing in the Il-8 and Il-4 production.

  1. Macrophage preconditioning with synthetic malaria pigment reduces cytokine production via heme iron-dependent oxidative stress.

    PubMed

    Taramelli, D; Recalcati, S; Basilico, N; Olliaro, P; Cairo, G

    2000-12-01

    Hemozoin (malaria pigment), a polymer of hematin (ferri-protoporphyrin IX) derived from hemoglobin ingested by intraerythrocytic plasmodia, modulates cytokine production by phagocytes. Mouse peritoneal macrophages (PM) fed with synthetic beta-hematin (BH), structurally identical to native hemozoin, no longer produce tumor necrosis factor alpha (TNFalpha) and nitric oxide (NO) in response to lipopolysaccharide (LPS). Impairment of NO synthesis is due to inhibition of inducible nitric oxide synthase (iNOS) production. BH-mediated inhibition of PM functions cannot be ascribed to iron release from BH because neither prevention by iron chelators nor down-regulation of iron-regulatory protein activity was detected. Inhibition appears to be related to pigment-induced oxidative stress because (a) thiol compounds partially restored PM functions, (b) heme oxygenase (HO-1) and catalase mRNA levels were up-regulated, and (c) free radicals production increased in BH-treated cells. The antioxidant defenses of the cells determine the response to BH: microglia cells, which show a lower extent of induction of HO-1 and catalase mRNAs and lower accumulation of oxygen radicals, are less sensitive to the inhibitory effect of BH on cytokine production. Results indicate that BH is resistant to degradation by HO-1 and that heme-iron mediated oxidative stress may contribute to malaria-induced immunosuppression. This study may help correlate the different clinical manifestations of malaria, ranging from uncomplicated to severe disease, with dysregulation of phagocyte functions and promote better therapeutic strategies to counteract the effects of hemozoin accumulation.

  2. Effects of the Commercial Flame Retardant Mixture DE-71 on Cytokine Production by Human Immune Cells

    PubMed Central

    Mynster Kronborg, Thit; Frohnert Hansen, Juliana; Nielsen, Claus Henrik; Ramhøj, Louise; Frederiksen, Marie; Vorkamp, Katrin; Feldt-Rasmussen, Ulla

    2016-01-01

    Introduction Although production of polybrominated diphenyl ethers (PBDEs) is now banned, release from existing products will continue for many years. The PBDEs are assumed to be neurotoxic and toxic to endocrine organs at low concentrations. Their effect on the immune system has not been investigated thoroughly. We aimed to investigate the influence of DE-71 on cytokine production by peripheral blood mononuclear cells (PBMCs) stimulated with Escherichia Coli lipopolysaccharide (LPS) or phytohaemagglutinin-L (PHA-L). Material and Methods PBMCs isolated from healthy donors were pre-incubated with DE-71 at various concentrations and subsequently incubated with the monocyte stimulator LPS, or the T-cell activator PHA-L. Interferon (IFN)-γ, interleukin (IL)-1β, IL-2, IL-4, IL-6, IL-8, IL-10, tumor necrosis factor (TNF)-α, IL-17A, and IL-17F were quantified in the supernatants by Luminex kits. Results At non-cytotoxic concentrations (0.01–10 μg/mL), DE-71 significantly enhanced secretion of IL-1β, IL-6, CXCL8, IL-10, and TNF-α (p<0.001–0.019; n = 6) from LPS-stimulated PBMCs. IFN-γ, TNF-α, IL-17A, and IL-17F (p = <0.001–0.043; n = 6) secretion were enhanced from PHA-L-stimulated PBMCs as well. Secretion of IL-1β, IL-2, IL-10, IL-8 and IL-6 was not significantly affected by DE-71. Conclusions We demonstrate an enhancing effect of DE-71 on cytokine production by normal human PBMCs stimulated with LPS or PHA-L ex vivo. PMID:27128973

  3. The effect of fish oil supplementation on cytokine production in children.

    PubMed

    Vaisman, Nachum; Zaruk, Yahalomit; Shirazi, Idit; Kaysar, Nechemia; Barak, Vivian

    2005-09-01

    The ex vivo production of inflammatory cytokines during fish oil supplementation (n-3 polyunsaturated fatty acids, n-3 PUFA) is a matter of considerable controversy. Studies on human subjects have generally reported decreased lymphocyte proliferation and decreased production of IL-2, interferon-gamma, IL-1beta, IL-6 and TNF-alpha, but other studies showed no effect or even increased production. There are no published reports on ex vivo cytokine production in children on long-term, n-3 PUFA supplementation. The current double-blind study explored cytokine production by peripheral blood mononuclear cells (PBMCs), with and without lipopolysaccharide (LPS) stimulation in children on 12 weeks' supplementation with 300 mg/day of n-3 PUFA. Twenty-one children (aged 8-12 years) were randomized to receive 1 g canola oil (control) or 300 mg n-3 PUFA + 700 mg canola oil in a chocolate spread. Blood was then drawn and PBMCs were separated and cultured for 24 h in a culture medium with or without 10 microg/mL LPS for 5 x 10(6) PBMCs. The pro-inflammatory cytokines, IL-1beta, TNF-alpha and IL-6, and the anti-inflammatory cytokines, IL-10 and IL-1RA, were evaluated by ELISA. The levels of all the cytokines were higher in non-stimulated and LPS-stimulated cultures, from n-3 PUFA-treated subjects as compared to controls. There was no difference in the IL-1beta/IL-1RA ratio between the two groups, with and without LPS stimulation. Nevertheless, the ratio tended to be lower in the treated subjects on both occasions. In conclusion, our results indicate an increased production of both pro-inflammatory and anti-inflammatory cytokines, with and without LPS stimulation, in children on 12 weeks' n-3 PUFA supplementation.

  4. Mint3/Apba3 depletion ameliorates severe murine influenza pneumonia and macrophage cytokine production in response to the influenza virus

    PubMed Central

    Uematsu, Takayuki; Fujita, Tomoko; Nakaoka, Hiroki J.; Hara, Toshiro; Kobayashi, Noritada; Murakami, Yoshinori; Seiki, Motoharu; Sakamoto, Takeharu

    2016-01-01

    Influenza virus (IFV) infection is a common cause of severe pneumonia. Studies have suggested that excessive activation of the host immune system including macrophages is responsible for the severe pathologies mediated by IFV infection. Here, we focused on the X11 protein family member Mint3/Apba3, known to promote ATP production via glycolysis by activating hypoxia inducible factor-1 (HIF-1) in macrophages, and examined its roles in lung pathogenesis and anti-viral defence upon IFV infection. Mint3-deficient mice exhibited improved influenza pneumonia with reduced inflammatory cytokines/chemokine levels and neutrophil infiltration in the IFV-infected lungs without alteration in viral burden, type-I interferon production, or acquired immunity. In macrophages, Mint3 depletion attenuated NF-κB signalling and the resultant cytokine/chemokine production in response to IFV infection by increasing IκBα and activating the cellular energy sensor AMPK, respectively. Thus, Mint3 might represent one of the likely therapeutic targets for the treatment of severe influenza pneumonia without affecting host anti-viral defence through suppressing macrophage cytokine/chemokine production. PMID:27883071

  5. The Effect of Long-Term Exercise on the Production of Osteoclastogenic and Antiosteoclastogenic Cytokines by Peripheral Blood Mononuclear Cells and on Serum Markers of Bone Metabolism

    PubMed Central

    Dykes, Rhesa; Chi, David S.

    2016-01-01

    Although it is recognized that the mechanical stresses associated with physical activity augment bone mineral density and improve bone quality, our understanding of how exercise modulates bone homeostasis at the molecular level is lacking. In a before and after trial involving 43 healthy adults, we measured the effect of six months of supervised exercise training on the spontaneous and phytohemagglutinin-induced production of osteoclastogenic cytokines (interleukin-1α, tumor necrosis factor-α), antiosteoclastogenic cytokines (transforming growth factor-β1 and interleukins 4 and 10), pleiotropic cytokines with variable effects on osteoclastogenesis (interferon-γ, interleukin-6), and T cell growth and differentiation factors (interleukins 2 and 12) by peripheral blood mononuclear cells. We also measured lymphocyte phenotypes and serum markers of bone formation (osteocalcin), bone resorption (C-terminal telopeptides of Type I collagen), and bone homeostasis (25 (OH) vitamin D, estradiol, testosterone, parathyroid hormone, and insulin-like growth factor 1). A combination of aerobic, resistance, and flexibility exercises done on average of 2.5 hours a week attenuated the production of osteoclastogenic cytokines and enhanced the production of antiosteoclastogenic cytokines. These changes were accompanied by a 16% reduction in collagen degradation products and a 9.8% increase in osteocalcin levels. We conclude that long-term moderate intensity exercise exerts a favorable effect on bone resorption by changing the balance between blood mononuclear cells producing osteoclastogenic cytokines and those producing antiosteoclastogenic cytokines. This trial is registered with Clinical Trials.gov Identifier: NCT02765945. PMID:27642534

  6. Polybrominated diphenyl ethers enhance the production of proinflammatory cytokines by the placenta.

    PubMed

    Peltier, M R; Klimova, N G; Arita, Y; Gurzenda, E M; Murthy, A; Chawala, K; Lerner, V; Richardson, J; Hanna, N

    2012-09-01

    Polybrominated diphenyl ether(s) (PBDE) are ubiquitous environmental contaminants that bind and cross the placenta but their effects on pregnancy outcome are unclear. It is possible that environmental contaminants increase the risk of inflammation-mediated pregnancy complications such as preterm birth by promoting a proinflammatory environment at the maternal-fetal interface. We hypothesized that PBDE would reduce IL-10 production and enhance the production of proinflammatory cytokines associated with preterm labor/birth by placental explants. Second-trimester placental explants were cultured in either vehicle (control) or 2 μM PBDE mixture of congers 47, 99 and 100 for 72 h. Cultures were then stimulated with 10(6) CFU/ml heat-killed Escherichia coli for a final 24 h incubation and conditioned medium was harvested for quantification of cytokines and PGE(2). COX-2 content and viability of the treated tissues were then quantified by tissue ELISA and MTT reduction activity, respectively. PBDE pre-treatment reduced E. coli-stimulated IL-10 production and significantly increased E. coli-stimulated IL-1β secretion. PBDE exposure also increased basal and bacteria-stimulated COX-2 expression. Basal, but not bacteria-stimulated PGE(2), was also enhanced by PBDE exposure. No effect of PBDE on viability of the explants cultures was detected. In summary, pre-exposure of placental explants to congers 47, 99, and 100 enhanced the placental proinflammatory response to infection. This may increase the risk of infection-mediated preterm birth by lowering the threshold for bacteria to stimulate a proinflammatory response(s).

  7. Fucoidan delays apoptosis and induces pro-inflammatory cytokine production in human neutrophils.

    PubMed

    Jin, Jun-O; Yu, Qing

    2015-02-01

    Although some immune modulatory effects of fucoidan have been elucidated, the effects of fucoidan on the apoptosis and activation of human neutrophils have not been investigated. In this study, we demonstrated that fucoidan purified from the brown seaweed Undaria pinnatifilda delays spontaneous apoptosis of human neutrophils and induces their activation. Fucoidan treatment inhibited apoptotic nuclei changes and phosphatidyl serine (PS) exposure on neutrophils cultured in vitro for 24h. The delay in neutrophil apoptosis mediated by fucoidan was associated with increased levels of the anti-apoptotic protein Mcl-1 and decreased levels of activated caspase-3. Screening of the signaling pathways by specific inhibitors indicated that fucoidan-induced delay in neutrophil apoptosis was dependent on the activation of PI3K/AKT signaling pathway, whereas MAPK signaling pathway was not critical. In addition, fucoidan enhanced the production of IL-6, IL-8 and TNF-α from neutrophils in an AKT-dependent manner. Taken together, these results demonstrated that fucoidan delays human neutrophil apoptosis and induces their production of pro-inflammatory cytokines. This knowledge could facilitate the development of novel therapeutic strategies for infectious diseases and neutropenia by controlling neutrophil homeostasis and function with fucoidan.

  8. Allergens stimulate store-operated calcium entry and cytokine production in airway epithelial cells

    PubMed Central

    Jairaman, Amit; Maguire, Chelsea H.; Schleimer, Robert P.; Prakriya, Murali

    2016-01-01

    Aberrant immune responses to environmental allergens including insect allergens from house dust mites and cockroaches contribute to allergic inflammatory diseases such as asthma in susceptible individuals. Airway epithelial cells (AECs) play a critical role in this process by sensing the proteolytic activity of allergens via protease-activated receptors (PAR2) to initiate inflammatory and immune responses in the airway. Elevation of cytosolic Ca2+ is an important signaling event in this process, yet the fundamental mechanism by which allergens induce Ca2+ elevations in AECs remains poorly understood. Here we find that extracts from dust mite and cockroach induce sustained Ca2+ elevations in AECs through the activation of Ca2+ release-activated Ca2+ (CRAC) channels encoded by Orai1 and STIM1. CRAC channel activation occurs, at least in part, through allergen mediated stimulation of PAR2 receptors. The ensuing Ca2+ entry then activates NFAT/calcineurin signaling to induce transcriptional production of the proinflammatory cytokines IL-6 and IL-8. These findings highlight a key role for CRAC channels as regulators of allergen induced inflammatory responses in the airway. PMID:27604412

  9. ApoE production in human monocytes and its regulation by inflammatory cytokines.

    PubMed

    Braesch-Andersen, Sten; Paulie, Staffan; Smedman, Christian; Mia, Sohel; Kumagai-Braesch, Makiko

    2013-01-01

    The apoE production by tissue macrophages is crucial for the prevention of atherosclerosis and the aim of this study was to further elucidate how this apolipoprotein is regulated by cytokines present during inflammation. Here we studied apoE production in peripheral blood mononuclear cells (PBMC) and analysis was made with a newly developed apoE ELISpot assay. In PBMC, apoE secretion was restricted to monocytes with classical (CD14(++)CD16(-)) and intermediate (CD14(+)CD16(+)) monocytes being the main producers. As earlier described for macrophages, production was strongly upregulated by TGF-β and downregulated by bacterial lipopolysaccharide (LPS) and the inflammatory cytokines IFN-γ, TNF-α and IL-1β. We could here show that a similar down-regulatory effect was also observed with the type I interferon, IFN-α, while IL-6, often regarded as one of the more prominent inflammatory cytokines, did not affect TGF-β-induced apoE production. The TNF-α inhibitor Enbrel could partly block the down-regulatory effect of IFN-γ, IFN-α and IL-1β, indicating that inhibition of apoE by these cytokines may be dependent on or synergize with TNF-α. Other cytokines tested, IL-2, IL-4, IL-12, IL-13, IL-17A and IL-23, had no inhibitory effect on apoE production. In contrast to the effect on monocytes, apoE production by primary hepatocytes and the hepatoma cell line HepG2 was more or less unaffected by treatment with cytokines or LPS.

  10. Dysfunction of Nrf-2 in CF Epithelia Leads to Excess Intracellular H2O2 and Inflammatory Cytokine Production

    PubMed Central

    Chen, Junnan; Kinter, Michael; Shank, Samuel; Cotton, Calvin; Kelley, Thomas J.; Ziady, Assem G.

    2008-01-01

    Cystic fibrosis is characterized by recurring pulmonary exacerbations that lead to the deterioration of lung function and eventual lung failure. Excessive inflammatory responses by airway epithelia have been linked to the overproduction of the inflammatory cytokine IL-6 and IL-8. The mechanism by which this occurs is not fully understood, but normal IL-1β mediated activation of the production of these cytokines occurs via H2O2 dependent signaling. Therefore, we speculated that CFTR dysfunction causes alterations in the regulation of steady state H2O2. We found significantly elevated levels of H2O2 in three cultured epithelial cell models of CF, one primary and two immortalized. Increases in H2O2 heavily contributed to the excessive IL-6 and IL-8 production in CF epithelia. Proteomic analysis of three in vitro and two in vivo models revealed a decrease in antioxidant proteins that regulate H2O2 processing, by ≥2 fold in CF vs. matched normal controls. When cells are stimulated, differential expression in CF versus normal is enhanced; corresponding to an increase in H2O2 mediated production of IL-6 and IL-8. The cause of this redox imbalance is a decrease by ∼70% in CF cells versus normal in the expression and activity of the transcription factor Nrf-2. Inhibition of CFTR function in normal cells produced this phenotype, while N-acetyl cysteine, selenium, an activator of Nrf-2, and the overexpression of Nrf-2 all normalized H2O2 processing and decreased IL-6 and IL-8 to normal levels, in CF cells. We conclude that a paradoxical decrease in Nrf-2 driven antioxidant responses in CF epithelia results in an increase in steady state H2O2, which in turn contributes to the overproduction of the pro-inflammatory cytokines IL-6 and IL-8. Treatment with antioxidants can ameliorate exaggerated cytokine production without affecting normal responses. PMID:18846238

  11. CSF findings in adrenoleukodystrophy: correlation between measures of cytokines, IgG production, and disease severity.

    PubMed

    Phillips, J P; Lockman, L A; Shapiro, E G; Blazar, B R; Loes, D J; Moser, H W; Krivit, W

    1994-06-01

    The childhood-onset cerebral form of adrenoleukodystrophy has a devastating neurologic prognosis. Unfortunately, there is no early method of distinguishing it from the more benign forms of adrenoleukodystrophy, such as adrenomyeloneuropathy. To evaluate the manner in which this disease entity may be reflected in the cerebrospinal fluid, we studied a consecutive series of 19 patients, all with biochemically proved adrenoleukodystrophy. total protein, immunoglobulin production, cytokine levels, and cerebrospinal fluid pressure were measured. In this single sample of cerebrospinal fluid, a significant correlation existed between clinical stage of the illness and cerebrospinal fluid myelin basic protein. No correlation existed with total protein, cytokines, or measures of immunoglobulin production.

  12. Forced expression of stabilized c-Fos in dendritic cells reduces cytokine production and immune responses in vivo

    SciTech Connect

    Yoshida, Ryoko; Suzuki, Mayu; Sakaguchi, Ryota; Hasegawa, Eiichi; Kimura, Akihiro; Shichita, Takashi; Sekiya, Takashi; Shiraishi, Hiroshi; Shimoda, Kouji; Yoshimura, Akihiko

    2012-06-29

    Highlights: Black-Right-Pointing-Pointer Dendritic cells expressing stabilized c-Fos produced less inflammatory cytokines. Black-Right-Pointing-Pointer Dendritic cells expressing stabilized c-Fos activated T cells less efficiently. Black-Right-Pointing-Pointer Transgenic mice expressing stabilized c-Fos were resistant to EAE model. -- Abstract: Intracellular cyclic adenosine monophosphate (cAMP) suppresses innate immunity by inhibiting proinflammatory cytokine production by monocytic cells. We have shown that the transcription factor c-Fos is responsible for cAMP-mediated suppression of inflammatory cytokine production, and that c-Fos protein is stabilized by IKK{beta}-mediated phosphorylation. We found that S308 is one of the major phosphorylation sites, and that the S308D mutation prolongs c-Fos halflife. To investigate the role of stabilized c-Fos protein in dendritic cells (DCs) in vivo, we generated CD11c-promoter-deriven c-FosS308D transgenic mice. As expected, bone marrow-derived DCs (BMDCs) from these Tg mice produced smaller amounts of inflammatory cytokines, including TNF-{alpha}, IL-12, and IL-23, but higher levels of IL-10, in response to LPS, than those from wild-type (Wt) mice. When T cells were co-cultured with BMDCs from Tg mice, production of Th1 and Th17 cytokines was reduced, although T cell proliferation was not affected. Tg mice demonstrated more resistance to experimental autoimmune encephalomyelitis (EAE) than did Wt mice. These data suggest that c-Fos in DCs plays a suppressive role in certain innate and adaptive immune responses.

  13. A mixture of phytosterols from Dunaliella tertiolecta affects proliferation of peripheral blood mononuclear cells and cytokine production in sheep.

    PubMed

    Caroprese, Mariangela; Albenzio, Marzia; Ciliberti, Maria Giovanna; Francavilla, Matteo; Sevi, Agostino

    2012-11-15

    The aim of this study was to investigate the immunomodulatory and anti-inflammatory role of a mixture of phytosterols extracted from the microalga Dunaliella tertiolecta on peripheral blood mononuclear cells (PBMC) isolated from sheep. PBMC were treated to determine cell proliferation and cytokine production with different sterols: ergosterol (E), a mixture of eleven Algae sterols extracted and purified from D. tertiolecta (Algae Extract, AE), a mixture of ergosterol and 7-dehydroporiferasterol extracted and purified from D. tertiolecta (Purified Extract, PE). Cytokine production (TNF-α, IL-6, IL-1β, and IL-10) was evaluated after cell treatment with Concanavalin A (Con A) and lipopolysaccharide (LPS). The mixture of ergosterol and 7-dehydroporiferasterol extracted and purified from D. tertiolecta showed a suppressive effect on cell proliferation, and a reduction of pro-inflammatory cytokines production. Furthermore, a stimulatory effect on the production of the regulatory cytokine IL-10 was found. The immunosuppressive effect exerted by the mixture of ergosterol and 7-dehydroporiferasterol from D. tertiolecta was dose-dependent both in suppressing cell proliferation and in stimulating IL-10 production. Present results showed that the immunomodulatory and anti-inflammatory activities were more apparent in the purified extract characterized by the mixture of ergosterol and 7-dehydroporiferasterol, and might depend on the existence of a synergic effect of the structures of the two phytosterols. Furthermore, findings from our study suggest that the purified extract characterized by the mixture of ergosterol and 7-dehydroporiferasterol from D. tertiolecta could be used to reduce immune reactions resulting from inflammatory diseases in sheep production systems, and could have innovative implications on the modulation of sheep immune system when used as feed supplements.

  14. Inhibitory effects of amantadine on the production of pro-inflammatory cytokines by stimulated in vitro human blood.

    PubMed

    Kubera, Marta; Maes, Michael; Budziszewska, Bogusława; Basta-Kaim, Agnieszka; Leśkiewicz, Monika; Grygier, Beata; Rogóz, Zofia; Lasoń, Władysław

    2009-01-01

    Treatment with amantadine (AMA), an N-methyl-D-aspartate (NMDA) receptor antagonist and antidepressant drug, increased the antidepressant activity of subsequent drugs in experimental studies and in patients suffering from treatment-resistant depression (TRD). Recent evidence indicates that depression may be accompanied by activation of an inflammatory response. These data indicate that pro-inflammatory cytokines may play a role in the etiology of depression, particularly in TRD. The present in vitro study shows the ability of AMA, used at concentrations between 10(-7) to 10(-5) M, to reduce the production of the pro-inflammatory cytokines, specifically interferon-gamma (IFN-gamma) and tumor necrosis factor-alpha (TNF-alpha). In addition, AMA treatment increased the production of the negative immunoregulator, interleukin-10 (IL-10). Furthermore, the combined treatment of AMA with fluoxetine (FLU), but not imipramine (IMI), had a stronger immunomodulatory effect on cytokine production than AMA alone. The above data provide additional rationale for the treatment of patients suffering from depression with a combination of AMA and a selective serotonin reuptake inhibitor.

  15. The cytokine response to human traumatic brain injury: temporal profiles and evidence for cerebral parenchymal production.

    PubMed

    Helmy, Adel; Carpenter, Keri L H; Menon, David K; Pickard, John D; Hutchinson, Peter J A

    2011-02-01

    The role of neuroinflammation is increasingly being recognised in a diverse range of cerebral pathologies, including traumatic brain injury (TBI). We used cerebral microdialysis and paired arterial and jugular bulb plasma sampling to characterise the production of 42 cytokines after severe TBI in 12 patients over 5 days. We compared two microdialysis perfusates in six patients: central nervous system perfusion fluid and 3.5% human albumin solution (HAS); 3.5% HAS has a superior fluid recovery (95.8 versus 83.3%), a superior relative recovery in 18 of 42 cytokines (versus 8 of 42), and a qualitatively superior recovery profile. All 42 cytokines were recovered from the human brain. Sixteen cytokines showed a stereotyped temporal peak, at least twice the median value for that cytokine over the monitoring period; day 1: tumour necrosis factor, interleukin (IL)7, IL8, macrophage inflammatory protein (MIP)1α, soluble CD40 ligand, GRO, IL1β, platelet derived growth factor (PDGF)-AA, MIP1β, RANTES; day 2: IL1 receptor antagonist (ra). IL6, granulocyte-colony stimulating factor (G-CSF), chemokine CXC motif ligand 10 (IP10); days 4 to 5: IL12p70, IL10. Brain extracellular fluid concentrations were significantly higher than plasma concentrations for 19 cytokines: basic fibroblast growth factor (FGF2), G-CSF, IL1α, IL1β, IL1ra, IL3, IL6, IL8, IL10, IL12p40, IL12p70, IP10, monocyte chemotactic protein (MCP)1, MCP3, MIP1α, MIP1β, PDGF-AA, transforming growth factor (TGF)α and vascular endothelial growth factor. No clear arterio-jugular venous gradients were apparent. These data provide evidence for the cerebral production of these cytokines and show a stereotyped temporal pattern after TBI.

  16. Local cytokine production in a murine model of Escherichia coli pyelonephritis.

    PubMed Central

    Rugo, H S; O'Hanley, P; Bishop, A G; Pearce, M K; Abrams, J S; Howard, M; O'Garra, A

    1992-01-01

    Cytokines may play an important role in the regulation of host defense against local bacterial infections. We have evaluated the local production of cytokines in a BALB/c mouse model of Escherichia coli pyelonephritis. Kidneys, draining lymph nodes, and spleens, were harvested at specific time intervals after bladder inoculation with E. coli corresponding to the stages of renal infection, infiltration, and bacterial clearance seen in this model. The presence of messenger RNA for specific cytokines (interleukins 1 through 6, chemotactic factors, granulocyte and granulocyte macrophage-colony stimulating factor (GM-CSF), tumor necrosis factor (TNF alpha) and beta, IFN gamma, transforming growth factor (TGF beta), and cytokine synthesis inhibitory factor (CSIF)/IL-10) was determined by polymerase chain reaction (PCR) amplification of reverse transcribed RNA. We have demonstrated mRNA encoding IL-1, IL-6, G-CSF, GM-CSF, TNF alpha, H400 (a protein homologous to a family of chemotactic factors and identical to MIP-1 beta), and CSIF/IL-10 in the kidney at 12 h and 1, 2, and 3 d after bacterial challenge. No signal was seen in normal animals or in mice after 5 d. This pattern of cytokine expression was observed only in renal tissues suggesting a localized response. IL-6 was present in the urine at 4 h with rapid resolution to baseline levels by 24 to 48 h. In contrast, IL-6 was not usually detectable in the serum. TNF alpha was not detectable in the serum or urine during the course of the infection. By immunohistochemical staining of kidney sections we have shown that IL-6 is produced predominantly by mesangial cells rather than by the inflammatory infiltrate. This study provides additional evidence utilizing novel techniques that specific cytokines are produced locally in response to bacterial infections. The time course of production demonstrated in this model supports the important role of cytokines in natural host resistance to local infection. Images PMID:1541664

  17. Cross-linked bromelain inhibits lipopolysaccharide-induced cytokine production involving cellular signaling suppression in rats.

    PubMed

    Hou, Rolis Chien-Wei; Chen, Yuh-Shuen; Huang, Jing-Rong; Jeng, Kee-Ching G

    2006-03-22

    Bromelain has been reported to have anti-inflammatory and immunomodulatory effects. It has been cross-linked with organic acids and polysaccharides by gamma irradiation. The cross-linked (CL)-bromelain preparation resisted an acidic environment of pH 3 for 2 h and preserved 80% of its enzyme activity. Pretreatment of rats with CL-bromelain intragastrically for 7 days significantly reduced serum cytokine production induced by injected i.p. with 2.5 mg/kg of lipopolysaccharide (LPS). Bromelain significantly reduced serum glutamate-oxalacetate transaminase induced by LPS. The anti-inflammatory effect of CL-bromelain was correlated with reduced LPS-induced NF-kappaB activity and cyclooxygenase 2 (COX-2) mRNA expression in rat livers. In addition, CL-bromelain dose-dependently inhibited LPS-induced COX-2 mRNA and prostaglandin E2 (PGE2) in BV-2 microglial cells. CL-Bromelain also suppressed the LPS-activated extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and p38 mitogen-activated protein kinase (MAPK). In conclusion, the anti-inflammatory effects of the CL-bromelain preparation in vivo and in vitro suggest its therapeutic potentials.

  18. Chronic blockade of glucocorticoid receptors by RU486 enhances lipopolysaccharide-induced depressive-like behaviour and cytokine production in rats.

    PubMed

    Wang, Donglin; Lin, Wenjuan; Pan, Yuqin; Kuang, Xueying; Qi, Xiaoli; Sun, Han

    2011-05-01

    Although accumulating evidence supports a role for cytokines in the pathophysiology of depression, the cytokine hypothesis of depression is debatable. It has been suggested that neuroendocrine and immune systems acting in concert may have roles in the development and the maintenance of the disease. Glucocorticoid receptor (GR) is the key element which exerts both anti-inflammatory and cytokine-inhibiting effects. Whether functional changes of GR are involved in the pathophysiology of cytokine-induced depression remains elusive. In the present study, the effects of both acute and chronic GR blockade on depressive-like behaviour and cytokine production induced by lipopolysaccharides (LPS), cytokine inducer, were investigated in rats. Acute or chronic blockade of GR was achieved by a single administration or repeated administrations, respectively, of the GR antagonist RU486 (RU). Behavioural measurements, including saccharin preference, locomotor activity, and immobility time, were assessed. The serum levels of proinflammatory cytokines (TNFα, IL-1β, and IFNγ) were determined by ELISA. The results showed that LPS induced significant but transient depressive-like behaviour. Repeated, but not single, administration of RU significantly enhanced and prolonged LPS-induced depressive-like behaviour and an increase in the serum production of TNFα and IFNγ. These results indicate that the effective blockade of GR enhanced the depressive-like behaviour induced by cytokines. Findings from this study suggest that GR dysfunction may be an important contributing factor to the development of cytokine-related depression. These findings add to the growing evidence of mechanisms by which cytokines influence depression.

  19. Alteration of cytokine production during visceral larva migrans by Toxascaris leonina in mice.

    PubMed

    Kang, Shin Ae; Park, Mi-Kyung; Cho, Min Kyoung; Yu, Hak Sun

    2013-10-01

    To determine alteration of immune responses during visceral larva migrans (VLM) caused by Toxascaris leonina at several time points, we experimentally infected mice with embryonated eggs of T. leonina and measured T-helper (Th) cell-related serial cytokine production after infection. At day 5 post infection (PI), most larvae were detected from the lungs, spleen, intestine, and muscle. Expression of thymic stromal lymphopoietin (TSLP) and CCL11 (eotaxin) showed a significant increase in most infected organs, except the intestine. However, expression of the CXCL1 (Gro-α) gene was most highly enhanced in the intestine at day 14 PI. Th1-related cytokine secretion of splenocytes showed increases at day 28 PI, and the level showed a decrease at day 42 PI. Th2-related cytokine secretion of splenocytes also showed an increase after infection; in particular, IL-5 level showed a significant increase at day 14 PI, and the level showed a decrease at day 28 PI. However, levels of Th17-related cytokines, IL-6 and IL-17A, showed gradual increases until day 42 PI. In conclusion, Th1, Th2, and Th17-related cytokine production might be important in immune responses against T. leonina VLM in experimental mice.

  20. Alteration of Cytokine Production during Visceral Larva Migrans by Toxascaris leonina in Mice

    PubMed Central

    Kang, Shin Ae; Park, Mi-Kyung; Cho, Min Kyoung

    2013-01-01

    To determine alteration of immune responses during visceral larva migrans (VLM) caused by Toxascaris leonina at several time points, we experimentally infected mice with embryonated eggs of T. leonina and measured T-helper (Th) cell-related serial cytokine production after infection. At day 5 post infection (PI), most larvae were detected from the lungs, spleen, intestine, and muscle. Expression of thymic stromal lymphopoietin (TSLP) and CCL11 (eotaxin) showed a significant increase in most infected organs, except the intestine. However, expression of the CXCL1 (Gro-α) gene was most highly enhanced in the intestine at day 14 PI. Th1-related cytokine secretion of splenocytes showed increases at day 28 PI, and the level showed a decrease at day 42 PI. Th2-related cytokine secretion of splenocytes also showed an increase after infection; in particular, IL-5 level showed a significant increase at day 14 PI, and the level showed a decrease at day 28 PI. However, levels of Th17-related cytokines, IL-6 and IL-17A, showed gradual increases until day 42 PI. In conclusion, Th1, Th2, and Th17-related cytokine production might be important in immune responses against T. leonina VLM in experimental mice. PMID:24327787

  1. Ceftiofur impairs pro-inflammatory cytokine secretion through the inhibition of the activation of NF-{kappa}B and MAPK

    SciTech Connect

    Ci Xinxin; Song Yu; Zeng Fanqin; Zhang Xuemei; Li Hongyu; Wang Xinrui; Cui Junqing Deng Xuming

    2008-07-18

    Ceftiofur is a new broad-spectrum, third-generation cephalosporin antibiotic for veterinary use. Immunopharmacological studies can provide new information on the immunomodulatory activities of some drugs, including their effect on cytokine productions. For this reason, we investigated the effect of ceftiofur on cytokine productions in vitro. We found that ceftiofur can downregulate tumor necrosis factor-{alpha} (TNF-{alpha}), interleukin-1{beta} (IL-1{beta}), and interleukin-6 (IL-6), but did not affect interleukin-10 (IL-10) production. We further investigated signal transduction mechanisms to determine how ceftiofur affects. RAW 264.7 cells were pretreated with 1, 5, or 10 mg/L of ceftiofur 1 h prior to treatment with 1 mg/L of LPS. Thirty minutes later, cells were harvested and mitogen activated protein kinases (MAPKs) activation was measured by Western blot. Alternatively, cells were fixed and nuclear factor-{kappa}B (NF-{kappa}B) activation was measured using immunocytochemical analysis. Signal transduction studies showed that ceftiofur significantly inhibited extracellular signal-regulated kinase (ERK), p38, and c-jun NH{sub 2}-terminal kinase (JNK) phosphorylation protein expression. Ceftiofur also inhibited p65-NF-{kappa}B translocation into the nucleus. Therefore, ceftiofur may inhibit LPS-induced production of inflammatory cytokines by blocking NF-{kappa}B and MAPKs signaling in RAW264.7 cells.

  2. The Highway to Hell: A RIP Kinase-Directed Shortcut to Inflammatory Cytokine Production.

    PubMed

    Hildebrand, Joanne M; Murphy, James M

    2016-07-19

    RIPK1 and RIPK3 are well-known signaling traffic cops in innate immunity. In this issue of Immunity, Degterev and colleagues show that when they blow the whistle on bacterial infection, they quickly point a white-gloved hand down the express route to inflammatory cytokine production.

  3. Blueberries inhibit proinflammatory cytokine TNF-alpha and IL-6 production in macrophages

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Blueberries (BB) have been reported to attenuate atherosclerosis in apoE deficient (ApoE-/-) mice. However, the underlying mechanisms are not fully understood. In this study, the effect of BB on proinflammatory cytokine production in macrophages was investigated. ApoE-/- mice were fed AIN-93G diet (...

  4. Influence of Phthalates on Cytokine Production in Monocytes and Macrophages: A Systematic Review of Experimental Trials

    PubMed Central

    Hansen, Juliana Frohnert; Bendtzen, Klaus; Boas, Malene; Frederiksen, Hanne; Nielsen, Claus H.; Rasmussen, Åse Krogh; Feldt-Rasmussen, Ulla

    2015-01-01

    Background Phthalates are a group of endocrine disrupting chemicals suspected to influence the immune system. The aim of this systematic review is to summarise the present knowledge on the influence of phthalates on monocyte and macrophage production and secretion of cytokines, an influence which could affect both pro- and anti-inflammatory abilities of these cells. Strategy and Results A systematic search was performed in Medline, Embase and Toxline in June 2013, last updated 3rd of August 2014. Criteria used to select studies were described and published beforehand online on Prospero (http://www.crd.york.ac.uk/NIHR_PROSPERO, registration number CRD42013004236). In vivo, ex vivo and in vitro studies investigating the influence of phthalates on cytokine mRNA expression and cytokine secretion in animals and humans were included. A total of 11 reports, containing 12 studies, were found eligible for inclusion. In these, a total of four different phthalate diesters, six primary metabolites (phthalate monoesters) and seven different cytokines were investigated. Though all studies varied greatly in study design and species sources, four out of five studies that investigated di-2-ethylhexyl phthalate found an increased tumour necrosis factor-α secretion/production from monocytes or macrophages. A summary of cytokine measurements was not possible since few studies were comparable in study design and due to insufficient reporting of raw data for most of the included studies. Conclusion Results from this review have suggested that at least one phthalate (di-2-ethylhexyl phthalate) has the ability to enhance tumour necrosis factor-α production/secretion from monocytes/macrophages in vitro, but also observed ex vivo. Influence of other phthalates on other cytokines has only been investigated in few studies. Thus, in vitro studies on primary human monocytes/macrophages as well as more in vivo studies are needed to confirm or dispute these findings. PMID:25811352

  5. Cytokine-release kinetics of platelet-rich plasma according to various activation protocols

    PubMed Central

    Roh, Y. H.; Kim, W.; Park, K. U.

    2016-01-01

    Objectives This study was conducted to evaluate the cytokine-release kinetics of platelet-rich plasma (PRP) according to different activation protocols. Methods Two manual preparation procedures (single-spin (SS) at 900 g for five minutes; double-spin (DS) at 900 g for five minutes and then 1500 g for 15 minutes) were performed for each of 14 healthy subjects. Both preparations were tested for platelet activation by one of three activation protocols: no activation, activation with calcium (Ca) only, or calcium with a low dose (50 IU per 1 ml PRP) of thrombin. Each preparation was divided into four aliquots and incubated for one hour, 24 hours, 72 hours, and seven days. The cytokine-release kinetics were evaluated by assessing PDGF, TGF, VEGF, FGF, IL-1, and MMP-9 concentrations with bead-based sandwich immunoassay. Results The concentration of cytokine released from PRP varied over time and was influenced by various activation protocols. Ca-only activation had a significant effect on the DS PRPs (where the VEGF, FGF, and IL-1 concentrations were sustained) while Ca/thrombin activation had effects on both SS and DS PRPs (where the PDGF and VEGF concentrations were sustained and the TGF and FGF concentrations were short). The IL-1 content showed a significant increase with Ca-only or Ca/thrombin activation while these activations did not increase the MMP-9 concentration. Conclusion The SS and DS methods differed in their effect on cytokine release, and this effect varied among the cytokines analysed. In addition, low dose of thrombin/calcium activation increased the overall cytokine release of the PRP preparations over seven days, relative to that with a calcium-only supplement or non-activation. Cite this article: Professor J. H. Oh. Cytokine-release kinetics of platelet-rich plasma according to various activation protocols. Bone Joint Res 2016;5:37–45. DOI: 10.1302/2046-3758.52.2000540 PMID:26862077

  6. Engagement of Fas on Macrophages Modulates Poly I:C induced cytokine production with specific enhancement of IP-10.

    PubMed

    Lyons, Caitriona; Fernandes, Philana; Fanning, Liam J; Houston, Aileen; Brint, Elizabeth

    2015-01-01

    Viral double-stranded RNA (dsRNA) is recognised by pathogen recognition receptors such as Toll-Like Receptor 3 (TLR3) and retinoic acid inducible gene-I (RIG-I), and results in cytokine and interferon production. Fas, a well characterised death receptor, has recently been shown to play a role in the inflammatory response. In this study we investigated the role of Fas in the anti-viral immune response. Stimulation of Fas on macrophages did not induce significant cytokine production. However, activation of Fas modified the response of macrophages to the viral dsRNA analogue poly I:C. In particular, poly I:C-induced IP-10 production was significantly enhanced. A similar augmentation of IP-10 by Fas was observed following stimulation with both poly A:U and Sendai virus. Fas activation suppressed poly I:C-induced phosphorylation of the MAP kinases p38 and JNK, while overexpression of the Fas adaptor protein, Fas-associated protein with death domain (FADD), activated AP-1 and inhibited poly I:C-induced IP-10 production. Consistent with an inhibitory role for AP-1 in IP-10 production, mutation of the AP-1 binding site on the IP-10 promoter resulted in augmented poly I:C-induced IP-10. These results demonstrate that engagement of the Fas receptor plays a role in modifying the innate immune response to viral RNA.

  7. Peroxiredoxin-1, a possible target in modulating inflammatory cytokine production in macrophage like cell line RAW264.7.

    PubMed

    Tae Lim, Young; Sup Song, Dong; Joon Won, Tae; Lee, Yun-Jung; Yoo, Jong-Sun; Eun Hyung, Kyeong; Won Yoon, Joo; Park, So-Young; Woo Hwang, Kwang

    2012-06-01

    Peroxiredoxin (PRX), a scavenger of H(2) O(2) and alkyl hydroperoxides in living organisms, protects cells from oxidative stress. Contrary to its known anti-oxidant roles, the involvement of PRX-1 in the regulation of lipopolysaccharide (LPS) signaling is poorly understood, possible immunological functions of PRX-1 having been uncovered only recently. In the present study, it was discovered that the PRX-1 deficient macrophage like cell line (RAW264.7) has anti-inflammatory activity when stimulated by LPS. Treatment with LPS for 3 hrs resulted in increased gene expression of an anti-inflammatory cytokine, interleukin-10 (IL-10), in PRX-1 knock down RAW264.7 cells. Gene expression of pro-inflammatory cytokines IL-1β and tumor necrosis factor- α (TNF-α) did not show notable changes under the same conditions. However, production of these cytokines significantly decreased in PRX-1 knock down RAW264.7 cells with 12 hrs of stimulation. Production of IL-10 was also increased in PRX-1 knock down RAW264.7 cells with 12 hrs of stimulation. We predicted that higher concentrations of IL-10 would result in decreased expression of IL-1β and TNF-α in PRX-1 knock-down cells. This was confirmed by blocking IL-10, which reestablished IL-1β and TNF-α secretion. We also observed that increased concentrations of IL-10 do not affect the NF-κB pathway. Interestingly, STAT3 phosphorylation by LPS stimulation was significantly increased in PRX-1 knockdown RAW264.7 cells. Up-regulation of IL-10 in PRX-1 knockdown cells and the resulting downregulation of proinflammatory cytokine production seem to involve the STAT3 pathway in macrophages. Thus, down-regulation of PRX-1 may contribute to the suppression of adverse effects caused by excessive activation of macrophages through affecting the STAT3 signaling pathway.

  8. "PERFEXT": a direct method for quantitative assessment of cytokine production in vivo at the local level.

    PubMed

    Villavedra, M; Carol, H; Hjulström, M; Holmgren, J; Czerkinsky, C

    1997-05-01

    A method termed "PERFEXT", based on sequential perfusion and detergent extraction of lymphoid and non-lymphoid organs, has been developed for the quantitative measurement of cytokines produced at a local level in a given tissue. In vivo treatment of mice with Staphylococcus enterotoxin B (SEB) or lipopolysaccharide (LPS) served as the model systems. Interleukin-2 (IL2) and interferon-gamma (IFN gamma) levels were monitored by ELISA analysis of extracted samples. After local footpad (FP) injection with SEB, spleen and serum IL2 levels peaked at 2-4 h, while IL2 levels peaked at around 4-8 h in both FP and popliteal lymph nodes. SEB injection resulted in increased IFN gamma levels both in the FP and the draining lymph node. The detection of cytokines in the intestine allows for the application of the method at mucosal sites as well, provided enzyme inhibitors are present during the extraction procedure. After FP injection with LPS, IFN gamma production was significantly increased in the draining lymph node and was detectable in the FP, whereas IL2 was undetectable in any organ examined. IL2 and IFN gamma could also be detected at the site of elicitation of a delayed-type hypersensitivity reaction following local FP challenge. Local cytokine production correlated with the swelling response, whereas cytokine production in the spleen did not. IL2 peaked early, followed by a late increase in IFN gamma production, corresponding to the maximum swelling. This simple method should prove useful for analysing the production of other cytokines in vivo in distinct anatomical compartments.

  9. Acellular components of Chlamydia pneumoniae stimulate cytokine production in human blood mononuclear cells.

    PubMed

    Netea, M G; Selzman, C H; Kullberg, B J; Galama, J M; Weinberg, A; Stalenhoef, A F; Van der Meer, J W; Dinarello, C A

    2000-02-01

    Accumulating evidence suggest that infection with Chlamydia pneumoniae is associated with atherosclerosis, but the mechanisms involved remain unclear. Inflammation is important in the initial phase of atherogenesis, and cytokines are important in the initiation and progression of inflammation. The aim of this study was to assess the capacity of acellular components of C. pneumoniae to stimulate the production of pro-inflammatory cytokines and chemokines. Peripheral blood mononuclear cells were stimulated in vitro with sonicated C. pneumoniae. Significant amounts of TNF-alpha, IL-1, IL-6, IL-8, monocyte chemoattractant protein-1 (MCP-1) and macrophage inflammatory protein-1alpha (MIP-1alpha) were produced. Inhibition of endotoxin using polymyxin B revealed that chlamydial endotoxin plays a minor role in the cytokine induction. Neutralization of TNF by TNF-binding protein and blockade of IL-1 receptors by IL-1 receptor antagonist revealed that TNF, IL-1 and IL-6 production was independent from each other, whereas IL-8 synthesis was strongly dependent on endogenous TNF and IL-1. In contrast, synthesis of MCP-1 and MIP-1alpha was dependent on endogenous TNF, but not IL-1. In conclusion, acellular components of C. pneumoniae are a potent stimulus for cytokine production, and this mechanism may have an important role in the inflammatory aspects of atherogenesis.

  10. Effects of a Spirulina-based dietary supplement on cytokine production from allergic rhinitis patients.

    PubMed

    Mao, T K; Van de Water, J; Gershwin, M E

    2005-01-01

    Spirulina represents a blue-green alga that is widely produced and commercialized as a dietary supplement for modulating immune functions, as well as ameliorating a variety of diseases. We have previously shown that the in vitro culture of Spirulina with human peripheral blood mononuclear cells (PBMCs) modulated the production of cytokines. In the present study, we evaluated the impact of a Spirulina-based dietary supplement (Earthrise Nutritionals, Inc., Irvine, CA) on patients with allergic rhinitis by assessing the production of cytokines [interleukin (IL)-4, interferon (IFN)-gamma, and IL-2] critical in regulating immunoglobulin E-mediated allergy. In a randomized double-blinded crossover study versus placebo, allergic individuals were fed daily with either placebo or Spirulina, at 1,000 mg or 2,000 mg, for 12 weeks. PBMCs isolated before and after the Spirulina feeding were stimulated with phytohemagglutinin (PHA) prior to determining the levels of cytokine from cell culture supernatants. Although Spirulina seemed to be ineffective at modulating the secretion of Th1 cytokines (IFN-gamma and IL-2), we discovered that Spirulina, administered at 2,000 mg/day, significantly reduced IL-4 levels by 32% from PHA-stimulated cells. These results indicate that Spirulina can modulate the Th profile in patients with allergic rhinitis by suppressing the differentiation of Th2 cells mediated, in part, by inhibiting the production of IL-4. To our knowledge, this is the first human feeding study that demonstrates the protective effects of Spirulina towards allergic rhinitis.

  11. Metabolic Alterations Contribute to Enhanced Inflammatory Cytokine Production in Irgm1-deficient Macrophages.

    PubMed

    Schmidt, Elyse A; Fee, Brian E; Henry, Stanley C; Nichols, Amanda G; Shinohara, Mari L; Rathmell, Jeffrey C; MacIver, Nancie J; Coers, Jörn; Ilkayeva, Olga R; Koves, Timothy R; Taylor, Gregory A

    2017-03-17

    The immunity-related GTPases (IRGs) are a family of proteins that are induced by interferon (IFN)-γ and play pivotal roles in immune and inflammatory responses. IRGs ostensibly function as dynamin-like proteins that bind to intracellular membranes and promote remodeling and trafficking of those membranes. Prior studies have shown that loss of Irgm1 in mice leads to increased lethality to bacterial infections as well as enhanced inflammation to non-infectious stimuli; however, the mechanisms underlying these phenotypes are unclear. In the studies reported here, we found that uninfected Irgm1-deficient mice displayed high levels of serum cytokines typifying profound autoinflammation. Similar increases in cytokine production were also seen in cultured, IFN-γ-primed macrophages that lacked Irgm1. A series of metabolic studies indicated that the enhanced cytokine production was associated with marked metabolic changes in the Irgm1-deficient macrophages, including increased glycolysis and an accumulation of long chain acylcarnitines. Cells were exposed to the glycolytic inhibitor, 2-deoxyglucose, or fatty acid synthase inhibitors to perturb the metabolic alterations, which resulted in dampening of the excessive cytokine production. These results suggest that Irgm1 deficiency drives metabolic dysfunction in macrophages in a manner that is cell-autonomous and independent of infectious triggers. This may be a significant contributor to excessive inflammation seen in Irgm1-deficient mice in different contexts.

  12. In vitro and in vivo effects of clove on pro-inflammatory cytokines production by macrophages.

    PubMed

    Rodrigues, T G; Fernandes, A; Sousa, J P B; Bastos, J K; Sforcin, J M

    2009-01-01

    Biological properties of clove have been reported, but little is known about its effect on the immune system. This work was aimed to investigate the effect in vivo of a water-soluble part of hydroalcoholic extract of clove on pro-inflammatory cytokines (IL-1beta and IL-6) production by macrophages of BALB/c mice. The action of the essential oil of clove on the production of these cytokines macrophages was also investigated in vitro. The chemical compositions of the extract and of the oil were also investigated. Treatment of mice with water extract of clove was found to inhibit macrophages to produce both IL-1beta and IL-6. The essential oil of clove also inhibited the production of these cytokines in vitro. Eugenol was found to be the major component of the clove extract and essential oil, and probably is the causative agent of cytokine inhibition. Taken together, these data suggest an anti-inflammatory action of this spice.

  13. Dimerization by a cytokine receptor is necessary for constitutive activation of JAK2V617F.

    PubMed

    Lu, Xiaohui; Huang, Lily Jun-Shen; Lodish, Harvey F

    2008-02-29

    The majority of the BCR-ABL-negative myeloproliferative disorders express the mutant JAK2, JAK2V617F. Previously we showed that constitutive activation of this oncogenic JAK2 mutant in Ba/F3 or 32D cells requires coexpression of a cognate homodimeric cytokine receptor, such as the EpoR. However, overexpression of JAK2V617F in Ba/F3 cells renders them cytokine-independent for growth in the absence of an exogenous cytokine receptor. Here, we demonstrated that JAK2V617F domains required for receptor association are essential for cytokine-independent growth by overexpressed JAK2V617F, suggesting JAK2V617F is binding to an unknown endogenous cytokine receptor(s) for its activation. We further showed that disruption of EpoR dimerization by coexpressing a truncated EpoR disrupted JAK2V617F-mediated transformation, indicating that EpoR dimerization plays an essential role in the activation of JAK2V617F. Interestingly, coexpression of JAK2V617F with EpoR mutants that retain JAK2 binding but are defective in mediating Epo-dependent JAK2 activation due to mutations in a conserved juxtamembrane motif does lead to cytokine-independent activation of JAK2V617F. Overall, these findings confirm that JAK2V617F requires binding to a dimerized cytokine receptor for its activation, and that the key EpoR juxtamembrane regulatory motif essential for Epo-dependent JAK2 activation is not essential for the activation of JAK2V617F. The structure of the activated JAK2V617F is thus likely to be different from that of the activated wild-type JAK2, raising the possibility of developing a specifically targeted therapy for myeloproliferative disorders.

  14. Anti-human cytomegalovirus activity of cytokines produced by CD4+ T-cell clones specifically activated by IE1 peptides in vitro.

    PubMed Central

    Davignon, J L; Castanié, P; Yorke, J A; Gautier, N; Clément, D; Davrinche, C

    1996-01-01

    The control of latent cytomegalovirus (CMV) infections by the immune system is poorly understood. We have previously shown that CD4+ T cells specific for the human CMV major regulatory protein IE1 are frequent in latently infected healthy blood donors. In order to learn about the possible role of these cells, we have developed IE1-specific CD4+ T-cell clones and, in this study, analyzed their epitope specificity and function in vitro. We measured their cytokine production when stimulated with specific IE1 peptides or whole recombinant IE1 protein. Their cytokine profiles, as deduced from gamma interferon (IFN-gamma), tumor necrosis factor alpha (TNF-alpha), and interleukin-4 (IL-4) and IL-6 production, were of the Th0- and Th1-like phenotypes. Supernatants from IE1-specific clones producing IFN-gamma and TNF-alpha were shown to inhibit CMV replication in U373 MG cells. This effect was due, as found by using cytokine-specific neutralizing antibodies, mostly to IFN-gamma, which was secreted at higher levels than TNF-alpha. To better assess the anti-CMV activity of cytokines, recombinant IFN-gamma and TNF-alpha were used and shown to have a synergistic effect on the inhibition of CMV replication and protein expression. Thus, IE1-specific CD4+ T cells display in vitro anti-CMV activity through cytokine secretion and may play a role in the control of in vivo latent infections. PMID:8642638

  15. Inhibition of neddylation represses lipopolysaccharide-induced proinflammatory cytokine production in macrophage cells.

    PubMed

    Chang, Fang-Mei; Reyna, Sara M; Granados, Jose C; Wei, Sung-Jen; Innis-Whitehouse, Wendy; Maffi, Shivani K; Rodriguez, Edward; Slaga, Thomas J; Short, John D

    2012-10-12

    Cullin-RING E3 ligases (CRLs) are a class of ubiquitin ligases that control the proteasomal degradation of numerous target proteins, including IκB, and the activity of these CRLs are positively regulated by conjugation of a Nedd8 polypeptide onto Cullin proteins in a process called neddylation. CRL-mediated degradation of IκB, which normally interacts with and retains NF-κB in the cytoplasm, permits nuclear translocation and transactivation of the NF-κB transcription factor. Neddylation occurs through a multistep enzymatic process involving Nedd8 activating enzymes, and recent studies have shown that the pharmacological agent, MLN4924, can potently inhibit Nedd8 activating enzymes, thereby preventing neddylation of Cullin proteins and preventing the degradation of CRL target proteins. In macrophages, regulation of NF-κB signaling functions as a primary pathway by which infectious agents such as lipopolysaccharides (LPSs) cause the up-regulation of proinflammatory cytokines. Here we have analyzed the effects of MLN4924, and compared the effects of MLN4924 with a known anti-inflammatory agent (dexamethasone), on certain proinflammatory cytokines (TNF-α and IL-6) and the NF-κB signaling pathway in LPS-stimulated macrophages. We also used siRNA to block neddylation to assess the role of this molecular process during LPS-induced cytokine responsiveness. Our results demonstrate that blocking neddylation, either pharmacologically or using siRNA, abrogates the increase in certain proinflammatory cytokines secreted from macrophages in response to LPS. In addition, we have shown that MLN4924 and dexamethasone inhibit LPS-induced cytokine up-regulation at the transcriptional level, albeit through different molecular mechanisms. Thus, neddylation represents a novel molecular process in macrophages that can be targeted to prevent and/or treat the LPS-induced up-regulation of proinflammatory cytokines and the disease processes associated with their up-regulation.

  16. Vagus nerve stimulation inhibits cytokine production and attenuates disease severity in rheumatoid arthritis

    PubMed Central

    Koopman, Frieda A.; Chavan, Sangeeta S.; Miljko, Sanda; Grazio, Simeon; Sokolovic, Sekib; Schuurman, P. Richard; Mehta, Ashesh D.; Levine, Yaakov A.; Faltys, Michael; Zitnik, Ralph; Tracey, Kevin J.; Tak, Paul P.

    2016-01-01

    Rheumatoid arthritis (RA) is a heterogeneous, prevalent, chronic autoimmune disease characterized by painful swollen joints and significant disabilities. Symptomatic relief can be achieved in up to 50% of patients using biological agents that inhibit tumor necrosis factor (TNF) or other mechanisms of action, but there are no universally effective therapies. Recent advances in basic and preclinical science reveal that reflex neural circuits inhibit the production of cytokines and inflammation in animal models. One well-characterized cytokine-inhibiting mechanism, termed the “inflammatory reflex,” is dependent upon vagus nerve signals that inhibit cytokine production and attenuate experimental arthritis severity in mice and rats. It previously was unknown whether directly stimulating the inflammatory reflex in humans inhibits TNF production. Here we show that an implantable vagus nerve-stimulating device in epilepsy patients inhibits peripheral blood production of TNF, IL-1β, and IL-6. Vagus nerve stimulation (up to four times daily) in RA patients significantly inhibited TNF production for up to 84 d. Moreover, RA disease severity, as measured by standardized clinical composite scores, improved significantly. Together, these results establish that vagus nerve stimulation targeting the inflammatory reflex modulates TNF production and reduces inflammation in humans. These findings suggest that it is possible to use mechanism-based neuromodulating devices in the experimental therapy of RA and possibly other autoimmune and autoinflammatory diseases. PMID:27382171

  17. Serum Cytokine Profile in Asian Indian Patients with Takayasu Arteritis and its Association with Disease Activity

    PubMed Central

    Goel, Ruchika; Kabeerdoss, Jayakanthan; Ram, Babu; Prakash, John Antony Jude; Babji, Sudhir; Nair, Aswin; Jeyaseelan, Lakshmanan; Jeyaseelan, Visalakshi; Mathew, John; Balaji, Veeraraghavan; Joseph, George; Danda, Debashish

    2017-01-01

    Background: Arterial inflammation Takayasu arteritis (TA) is an outcome of balance between pro- and anti-inflammatory cytokines. Comprehensive assessment of these cytokines is important for understanding pathogenesis and assessing disease activity. Objective: To study pro- and anti-inflammatory cytokines representing different T-helper cell pathway in serum samples of Asian Indian patients with TA and to assess their association with disease activity. Methods: Consecutive Indian patients with TA were assayed for serum interferon-γ, interleukin-6, interleukin-23, interleukin-17, interleukin-10 and transforming growth factor- β levels at baseline and follow up visit. Patients were grouped into active and stable disease based on Indian Takyasu Arteritis clinical Activity Score-2010. Serum levels of these cytokines between active and stable disease and between baseline and follow up visits were compared by non-parametric tests. Results: Among 32 patients enrolled, 15 were classified as active while 17 as stable disease at baseline. IFN-γ levels were significantly higher in active disease than stable disease (p=0.0129) while other cytokines did not differ significantly between 2 groups. Serum levels of none of the cytokines changed significantly over 2 visits in both responders and non-responders. IL23 levels positively correlate with disease duration ((r=0.999; p<0.005). Modest correlation was observed between IFN-γ and IL23 levels at both baseline and follow up and between IFN-γ and IL-6 and CRP at follow up. Conclusion: IFN-γ levels are raised in active disease in TA and correlates well with other biomarkers of disease activity and proinflammatory cytokines. There is also a direct correlation between Il-23 levels and disease duration.

  18. Effect of Lactobacillus paracasei Culture Filtrates and Artichoke Polyphenols on Cytokine Production by Dendritic Cells

    PubMed Central

    Sisto, Angelo; Luongo, Diomira; Treppiccione, Lucia; De Bellis, Palmira; Di Venere, Donato; Lavermicocca, Paola; Rossi, Mauro

    2016-01-01

    The most recent trend in research on probiotic bacteria aims at the exploitation of bioactive bacterial compounds that are responsible for health-promoting effects and suitable for medical applications. Therefore, the main purpose of this study was to ascertain if the immunomodulatory effects of L. paracasei strains on dendritic cells (DCs) were caused by bacterial metabolites released in the culture medium. For that reason, bacterial strains were grown in two media generally used for the culture of DCs, and the effects of culture filtrates on the maturation of DCs and cytokine production were evaluated. Moreover, to reveal potential synergistic effects on the immunomodulation of DCs, an artichoke phenolic extract (APE) was added to the media before bacterial growth. The experiments pointed out an interesting anti-inflammatory activity of a culture filtrate obtained after growing a probiotic L. paracasei strain in one of the media supplemented with APE. Therefore, this culture filtrate—which combines the anti-inflammatory activity and the other well-known health-promoting properties of artichoke phenolic compounds—could represent the basis for future particular exploitations. PMID:27754398

  19. Aloe vera downregulates LPS-induced inflammatory cytokine production and expression of NLRP3 inflammasome in human macrophages.

    PubMed

    Budai, Marietta M; Varga, Aliz; Milesz, Sándor; Tőzsér, József; Benkő, Szilvia

    2013-12-01

    Aloe vera has been used in traditional herbal medicine as an immunomodulatory agent inducing anti-inflammatory effects. However, its role on the IL-1β inflammatory cytokine production has not been studied. IL-1β production is strictly regulated both at transcriptional and posttranslational levels through the activity of Nlrp3 inflammasome. In this study we aimed to determine the effect of Aloe vera on the molecular mechanisms of Nlrp3 inflammasome-mediated IL-1β production in LPS-activated human THP-1 cells and monocyte-derived macrophages. Our results show that Aloe vera significantly reduced IL-8, TNFα, IL-6 and IL-1β cytokine production in a dose dependent manner. The inhibitory effect was substantially more pronounced in the primary cells. We found that Aloe vera inhibited the expression of pro-IL-1β, Nlrp3, caspase-1 as well as that of the P2X7 receptor in the LPS-induced primary macrophages. Furthermore, LPS-induced activation of signaling pathways like NF-κB, p38, JNK and ERK were inhibited by Aloe vera in these cells. Altogether, we show for the first time that Aloe vera-mediated strong reduction of IL-1β appears to be the consequence of the reduced expression of both pro-IL-1β as well as Nlrp3 inflammasome components via suppressing specific signal transduction pathways. Furthermore, we show that the expression of the ATP sensor P2X7 receptor is also downregulated by Aloe vera that could also contribute to the attenuated IL-1β cytokine secretion. These results may provide a new therapeutic approach to regulate inflammasome-mediated responses.

  20. A new platform for constructing antibody-cytokine fusion proteins (immunocytokines) with improved biological properties and adaptable cytokine activity.

    PubMed

    Gillies, Stephen D

    2013-10-01

    A novel method for constructing immunocytokines has been developed that utilizes fusion of cytokines to the C-terminus of the Ig light chain, rather than fusing to the heavy chain. Such molecules are expressed well in transfected cells, are very stable in normal buffers and have biological properties that are superior to immunocytokines made by fusion to the heavy chain. These properties include longer circulating half-life, increased uptake following subcutaneous dosing and similar or improved antibody effector activities of antibody-dependent cytotoxic activity and complement-dependent cytotoxicity, respectively. Furthermore, the sequestering effect of this fusion junction allows one to adjust intermediate affinity (βγ) interleukin 2 receptor (IL2R) binding and activation by shortening the N-terminus of IL2 at the fusion point. This appears to limit access of the critical contact residue Asp20 of IL2 to the β-chain of βγ IL2R, while maintaining binding and activation of high-affinity (αβγ) IL2R-expressing cells. Several immunocytokine forms with varying degrees of IL2R specificity have been constructed, and some appear to regain their activity for the βγ IL2R when bound to antigen-coated beads. Such molecules may have reduced toxicity in the circulation and enhanced anti-tumor activity.

  1. Immunodepression in sepsis and SIRS assessed by ex vivo cytokine production is not a generalized phenomenon: a review.

    PubMed

    Cavaillon, J M; Adib-Conquy, M; Cloëz-Tayarani, I; Fitting, C

    2001-01-01

    Sepsis and non-infectious systemic inflammatory response syndrome (SIRS) are paradoxically associated with an exacerbated production of cytokines, as assessed by their presence in biological fluids, and a diminished ability of circulating leukocytes to produce cytokine upon in vitro activation. In this review, we depict that the observed cellular hyporeactivity is not a global phenomenon and that some signalling pathways are unaltered and allow the cells to respond normally to certain stimuli. Furthermore, we illustrate that during sepsis and SIRS, cells derived from tissues are either fully responsive to ex vivo stimuli or even primed, in contrast to cells derived from hematopoietic compartments (blood, spleen, etc.) which are hyporeactive. In addition to cytokine production, nuclear factor-kappa B (NF-kappa B) status within leukocytes can be used as a useful marker of hypo- or hyper-reactivity. We illustrate that the immune-depression reported in sepsis and SIRS patients, often revealed by a diminished capacity of leukocytes to respond to lipopolysaccharide, is not a generalized phenomenon and that SIRS is associated with a compartmentalized responsiveness which involves either anergic or primed cells.

  2. Human memory, but not naive, CD4+ T cells expressing transcription factor T-bet might drive rapid cytokine production.

    PubMed

    Yu, Si-fei; Zhang, Yan-nan; Yang, Bin-yan; Wu, Chang-you

    2014-12-19

    We found that after stimulation for a few hours, memory but not naive CD4(+) T cells produced a large amount of IFN-γ; however, the mechanism of rapid response of memory CD4(+) T cells remains undefined. We compared the expression of transcription factors in resting or activated naive and memory CD4(+) T cells and found that T-bet, but not pSTAT-1 or pSTAT-4, was highly expressed in resting memory CD4(+) T cells and that phenotypic characteristics of T-bet(+)CD4(+) T cells were CD45RA(low)CD62L(low) CCR7(low). After short-term stimulation, purified memory CD4(+) T cells rapidly produced effector cytokines that were closely associated with the pre-existence of T-bet. By contrast, resting naive CD4(+) T cells did not express T-bet, and they produced cytokines only after sustained stimulation. Our further studies indicated that T-bet was expressed in the nuclei of resting memory CD4(+) T cells, which might have important implications for rapid IFN-γ production. Our results indicate that the pre-existence and nuclear mobilization of T-bet in resting memory CD4(+) T cells might be a possible transcriptional mechanism for rapid production of cytokines by human memory CD4(+) T cells.

  3. Indomethacin Treatment of Mice with Premalignant Oral Lesions Sustains Cytokine Production and Slows Progression to Cancer

    PubMed Central

    Johnson, Sara D.; Young, M. Rita I.

    2016-01-01

    Current treatment options for head and neck squamous cell carcinoma (HNSCC) patients are often ineffective due to tumor-localized and systemic immunosuppression. Using the 4-NQO mouse model of oral carcinogenesis, this study showed that premalignant oral lesion cells produce higher levels of the immune modulator, PGE2, compared to HNSCC cells. Inhibiting prostaglandin production of premalignant lesion cells with the pan-cyclooxygenase inhibitor indomethacin stimulated their induction of spleen cell cytokine production. In contrast, inhibiting HNSCC prostaglandin production did not stimulate their induction of spleen cell cytokine production. Treatment of mice bearing premalignant oral lesions with indomethacin slowed progression of premalignant oral lesions to HNSCC. Flow cytometric analysis of T cells in the regional lymph nodes of lesion-bearing mice receiving indomethacin treatment showed an increase in lymph node cellularity and in the absolute number of CD8+ T cells expressing IFN-γ compared to levels in lesion-bearing mice receiving diluent control treatment. The cytokine-stimulatory effect of indomethacin treatment was not localized to regional lymph nodes but was also seen in the spleen of mice with premalignant oral lesions. Together, these data suggest that inhibiting prostaglandin production at the premalignant lesion stage boosts immune capability and improves clinical outcomes. PMID:27713748

  4. Altered Cytokine Production By Specific Human Peripheral Blood Cell Subsets Immediately Following Spaceflight

    NASA Technical Reports Server (NTRS)

    Crucian, Brian E.; Cubbage, Michael L.; Sams, Clarence F.

    1999-01-01

    In this study, we have attempted to combine standard immunological assays with the cellular resolving power of the flow cytometer to positively identify the specific cell types involved in spaceflight-induced immune alterations. We have obtained whole blood samples from 27 astronauts collected at three timepoints (L-10, R+0 and R+3) surrounding four recent space shuttle missions. The duration of these missions ranged from 10 to 18 days. Assays performed included serum/urine cortisol, comprehensive subset phenotyping, assessment of cellular activation markers and intracellular cytokine production following mitogenic stimulation. Absolute levels of peripheral granulocytes were significantly elevated following spaceflight, but the levels of circulating lymphocytes and monocytes were unchanged. Lymphocyte subset analysis demonstrated trends towards a decreased percentage of T cells and an increased percentage of B cells. Nearly all of the astronauts exhibited an increased CD4:CD8 ratio, which was dramatic in some individuals. Assessment of memory (CD45RA+) vs. naive (CD45RO+) CD4+ T cell subsets was more ambiguous, with subjects tending to group more as a flight crew. All subjects from one mission demonstrated an increased CD45RA:CD45RO ratio, while all subjects from another Mission demonstrated a decreased ratio. While no significant trend was seen in the monocyte population as defined by scatter, a decreased percentage of the CD14+ CD16+ monocyte subset was seen following spaceflight in all subjects tested. In general, most of the cellular changes described above which were assessed at R+O and compared to L-10 trended to pre-flight levels by R+3. Although no significant differences were seen in the expression of the cellular activation markers CD69 and CD25 following exposure to microgravity, significant alterations were seen in cytokine production in response to mitogenic activation for specific subsets. T cell (CD3+) production of IL-2 was significantly decreased

  5. Modulating phenotype and cytokine production of leucocytic retinal infiltrate in experimental autoimmune uveoretinitis following intranasal tolerance induction with retinal antigens

    PubMed Central

    Laliotou, B.; Dick, A.

    1999-01-01

    BACKGROUND/AIM—Nasal administration of retinal antigens induces systemic tolerance which results in suppression of experimental autoimmune uveoretinitis (EAU) when subsequently exposed to antigen. The aim was to establish if tolerance induction alters retinal infiltrating leucocyte phenotype and cytokine profile in tolerised animals when there is significantly reduced tissue destruction despite immunisation with retinal antigen.
METHODS—Female Lewis rats were tolerised by intranasal administration with retinal extract (RE) before immunisation with RE to induce EAU. Control animals were administered phosphate buffered saline (PBS) intranasally. Post immunisation, daily clinical responses were recorded and at the height of disease, retinas were removed and either infiltrating leucocytes isolated for flow cytometric phenotype assessment and intracellular cytokine production, or chorioretina processed for immunohistochemistry. Fellow eyes were assessed for cytokine mRNA by semiquantitative RT-PCR.
RESULTS—Flow cytometric analysis showed that before clinical onset of EAU there is no evidence of macrophage infiltration and no significant difference in circulating T cell populations within the retina. By day 14 a reduced retinal infiltrate in tolerised animals was observed and in particular a reduction in numbers of "activated" (with respect to CD4 and MHC class II expression) macrophages. Immunohistochemistry confirmed these findings and additionally minimal rod outer segment destruction was observed histologically. Cytokine analysis revealed that both IL-10 mRNA and intracellular IL-10 production was increased in tolerised eyes 7 days post immunisation. Although by day 14 post immunisation, IL-10 production was equivalent in both groups, a reduced percentage of IFN-γ+ macrophages and IFN-γ+ CD4+ T cells with increased percentage of IL-4+ CD4+ T cells were observed in tolerised animals.
CONCLUSIONS—Leucocytic infiltrate is not only reduced in number

  6. [Antistaphylococcal activity of the complex of natural cytokines].

    PubMed

    Koval'chuk, L V; Gankovskaia, L V; Moroz, A F; Avedova, T A; Ukhina, T V

    2004-01-01

    The in vitro study of the action of the complex of natural cytokines (CNC), or preparation Superlymph, on different microbial test strains (Staphylococcus aureus, Escherichia coli, yeast-like fungi of the genus Candida) was carried out. CNC suppressed the growth of S. aureus test culture, depending on the concentration of the preparation. The inhibiting effect was observed in the presence of Ca and Mg bivalent cations in the medium. Under the chosen conditions of the experiment CNC did not inhibit the growth of E. coli test cultures, as well as test cultures of yeast-like fungi of the genus Candida. In addition to its antibacterial effect, Superlymph also produced some effect on the release of cathepsin G, the lysosomal enzyme of human leukocyte granules.

  7. Particulate matter initiates inflammatory cytokine release by activation of capsaicin and acid receptors in a human bronchial epithelial cell line.

    PubMed

    Veronesi, B; Oortgiesen, M; Carter, J D; Devlin, R B

    1999-01-01

    Recent experiments have shown that human bronchial epithelial cells (i.e., BEAS-2B) release pro-inflammatory cytokines (i.e., IL-6 and TNFalpha) in a receptor-mediated fashion in response to the neuropeptides, substance P (SP), calcitonin gene-related protein (CGRP), and the prototype botanical irritant capsaicin. In the present experiments, we examined the relevance of these receptors to particulate matter (PM)-associated cellular inflammation. BEAS-2B cells, exposed to residual oil fly ash particles (ROFA), responded with an immediate (<30 s) increase in intracellular calcium levels ([Ca2+]i), increases of key inflammatory cytokine transcripts (i.e., IL-6, IL-8, TNFalpha) within 2 h exposure, and subsequent release of IL-6 and IL-8 cytokine protein after 4 h exposure. Pretreatment of BEAS-2B cells with pharmacological antagonists selective for the SP or CGRP receptors reduced the ROFA-stimulated IL-6 cytokine production by approximately 25 and 50%, respectively. However, pretreatment of these cells with capsazepine (CPZ), an antagonist for capsaicin (i.e., vanilloid) receptors, inhibited the immediate increases in [Ca2+]i, diminished transcript (i.e., IL-6, IL-8, TNFalpha) levels and reduced IL-6 cytokine release to control levels. BEAS-2B cells exposed to ROFA in calcium-free media failed to demonstrate increases of [Ca2+]i and showed reduced levels of cytokine transcript (i.e., IL-6, IL-8, TNFalpha) and IL-6 release, suggesting that ROFA-stimulated cytokine formation was partially dependent on extracellular calcium sources. A final set of experiments compared the inflammatory properties of the soluble and acidic insoluble components of ROFA. BEAS-2B cells, exposed to ROFA or ROFA that had been filtered through a 0.2-micrometer pore filter, produced equivocal IL-6. BEAS-2B cells exposed to pH 5.0 media for 15 min released moderate amounts of IL-6, 4 h later. This cytokine release could be blocked by amiloride, a pH receptor antagonist, but not by CPZ. BEAS-2B

  8. Cytokine and Eicosanoid Production by Cultured Human Monocytes Exposed to Titanium Particulate Debris

    NASA Astrophysics Data System (ADS)

    Robinson, Timothy M.; Manley, Paul A.; Sims, Paul A.; Albrecht, Ralph; Darien, Benjamin J.

    1999-10-01

    Phagocytosis of particulate wear debris from arthroplasties by macrophages induces an inflammatory response that has been linked to implant loosening and premature failure of artificial joints. Inflammatory mediators released by phagocytic macrophages such as tumor necrosis factor-a (TNF-[alpha]), interleukin-1[beta] (IL-1[beta]), interleukin-6 (IL-6), and prostaglandin E2 (PGE2) are believed to play a central role in the pathogenesis of aseptic loosening. The objective of this study was to characterize titanium alloy particulates that closely match wear debris found around joint arthroplasties and to study their effects on the biosynthesis of inflammatory mediators by cultured monocytes. Peripheral blood monocytes were isolated from healthy human volunteers. Monocytes were cultured in 96-well plates for 24 h, washed, and exposed to three concentrations of titanium particulates and controls from 18Ð24 h. Supernatants were assayed for TNF-[alpha], IL-1[beta], IL-6, and PGE2 activity. Energy dispersive X-ray spectroscopy (EDX) verified the titanium alloy to be Ti6A14V. Scanning electron microscopy (SEM) analysis showed significant titanium particulate heterogeneity with approximately 95% of the particles <1 micrometer in diameter. SEM and EDX technology was useful in the characterization of the titanium particulates utilized for in vitro models of titanium-induced cytokine release by monocytes. Incubation of titanium particulates (in concentrations similar to those found around loosened prosthetic joints) with cultured monocytes significantly increased their production of TNF-[alpha], IL-1[beta], and PGE2.

  9. RNA-Seq Reveals Activation of Both Common and Cytokine-Specific Pathways following Neutrophil Priming

    PubMed Central

    Moots, Robert J.; Edwards, Steven W.

    2013-01-01

    Neutrophils are central to the pathology of inflammatory diseases, where they can damage host tissue through release of reactive oxygen metabolites and proteases, and drive inflammation via secretion of cytokines and chemokines. Many cytokines, such as those generated during inflammation, can induce a similar “primed” phenotype in neutrophils, but it is unknown if different cytokines utilise common or cytokine-specific pathways to induce these functional changes. Here, we describe the transcriptomic changes induced in control human neutrophils during priming in vitro with pro-inflammatory cytokines (TNF-α and GM-CSF) using RNA-seq. Priming led to the rapid expression of a common set of transcripts for cytokines, chemokines and cell surface receptors (CXCL1, CXCL2, IL1A, IL1B, IL1RA, ICAM1). However, 580 genes were differentially regulated by TNF-α and GM-CSF treatment, and of these 58 were directly implicated in the control of apoptosis. While these two cytokines both delayed apoptosis, they induced changes in expression of different pro- and anti-apoptotic genes. Bioinformatics analysis predicted that these genes were regulated via differential activation of transcription factors by TNF-α and GM-CSF and these predictions were confirmed using functional assays: inhibition of NF-κB signalling abrogated the protective effect of TNF-α (but not that of GM-CSF) on neutrophil apoptosis, whereas inhibition of JAK/STAT signalling abrogated the anti-apoptotic effect of GM-CSF, but not that of TNF-α (p<0.05). These data provide the first characterisation of the human neutrophil transcriptome following GM-CSF and TNF-α priming, and demonstrate the utility of this approach to define functional changes in neutrophils following cytokine exposure. This may provide an important, new approach to define the molecular properties of neutrophils after in vivo activation during inflammation. PMID:23554905

  10. Bisphosphonate effects in cancer and inflammatory diseases: in vitro and in vivo modulation of cytokine activities.

    PubMed

    Santini, Daniele; Fratto, Maria E; Vincenzi, Bruno; La Cesa, Annalisa; Dianzani, Caterina; Tonini, Giuseppe

    2004-01-01

    Bisphosphonates are endogenous pyrophosphate analogs in which a carbon atom replaces the central atom of oxygen. They are indicated in non-neoplastic diseases including osteoporosis, corticosteroid-induced bone loss, Paget disease, and in cancer-related diseases such as neoplastic hypercalcemia, multiple myeloma and bone metastases secondary to breast and prostate cancer. There is now extensive in vitro evidence suggesting a direct antitumor effect of bisphosphonates at different levels of action. Some new in vitro and in vivo studies support the cytostatic effects of bisphosphonates on tumor cells, and the effects on the regulation of cell growth, apoptosis, angiogenesis, cell adhesion, and invasion, with particular attention to biological properties. Well designed clinical trials are necessary to investigate whether the antitumor potential of bisphosphonates may be clinically relevant. On the basis of their effects on macrophages, we may divide bisphosphonates into two distinct categories: aminobisphosphonates, which sensitize macrophages to an inflammatory stimulus inducing an acute-phase response, and non-aminobisphosphonates that can be metabolized into macrophages and that may inhibit the inflammatory response of macrophages. There is evidence of aminobisphosphonate-induced pro-inflammatory response, in particular, related to modifications of the cytokine network. Several in vivo studies have demonstrated an acute-phase reaction after the first administration of aminobisphosphonates, with a significant increase in the main pro-inflammatory cytokines. However, a peculiar aspect concerning the action of non-aminobisphosphonates seems to be an anti-inflammatory activity caused by the inhibition of the release of inflammatory mediators from activated macrophages, such as interleukin (IL)-6, tumor necrosis factor-alpha and IL-1. The inhibition of inflammatory responses is demonstrated in both in vivo and in vitro models. This activity suggests the use of non

  11. Soluble antigens from group B streptococci induce cytokine production in human blood cultures.

    PubMed Central

    von Hunolstein, C; Totolian, A; Alfarone, G; Mancuso, G; Cusumano, V; Teti, G; Orefici, G

    1997-01-01

    Group B streptococcal antigens stimulated tumor necrosis factor alpha (TNF-alpha), interleukin-1 (IL-1), and IL-6 production in human blood cultures in a concentration- and time-dependent fashion. The minimal concentrations of type-specific polysaccharides, lipoteichoic acid, and group-specific polysaccharide required to produce these effects were, respectively, 0.01, 1, and 10 microg/ml. Cell separation experiments indicated that monocytes were the cell type mainly responsible for cytokine production. Time course studies indicated that TNF-alpha was released before the other cytokines. TNF-alpha, however, did not appear to directly induce IL-1beta, as shown by blockade experiments with anti-TNF-alpha antibodies. IL-6 levels were moderately but significantly decreased by anti-TNF-alpha. These data indicate that several products from group B streptococci are able to directly stimulate human monocytes to release TNF-alpha, IL-1beta, and IL-6. These findings may be clinically relevant, since proinflammatory cytokines can mediate pathophysiologic changes during sepsis. PMID:9317001

  12. Contributions of cell subsets to cytokine production during normal and impaired wound healing.

    PubMed

    Mirza, Rita E; Koh, Timothy J

    2015-02-01

    The objective of this study was to determine the relative contributions of different cell subsets to the production of cytokines and growth factors during normal and impaired wound healing. Cells were isolated from wounds of non-diabetic and diabetic mice and separated by magnetic sorting into neutrophils/T cells/B cells (NTB cell subset), monocytes/macrophages (Mo/Mp subset) and non-leukocytic cells including keratinocyte/fibroblast/endothelial cells (KFE subset). On both per cell and total contribution bases, the Mo/Mp subset was the dominant producer of pro-inflammatory cytokines interleukin (IL)-1β, tumor necrosis factor (TNF)-α and IL-6 in both non-diabetic and diabetic mice and was a significant producer of vascular endothelial cell growth factor (VEGF)-A, insulin-like growth factor (IGF)-1 and transforming growth factor (TGF)-β1. The NTB subset was also a significant producer of TNF-α and IL-10 whereas the KFE subset contributed significant amounts of VEGF, IGF-1 and TGF-β1. Sustained production of pro-inflammatory cytokines and impaired production of healing-associated factors were evident in each subset in diabetic mice. These data will be useful for further experimental and modeling studies on the role of cell subsets in wound healing as well as for designing therapeutic strategies for improving healing.

  13. Surfactant, but not the size of solid lipid nanoparticles (SLN) influences viability and cytokine production of macrophages.

    PubMed

    Schöler, N; Olbrich, C; Tabatt, K; Müller, R H; Hahn, H; Liesenfeld, O

    2001-06-19

    After intravenous (i.v.) injection, solid lipid nanoparticles (SLN) interact with mononuclear cells. Murine peritoneal macrophages were incubated with SLN formulations consisting of Dynasan 114 coated with different surfactants. The present study was performed to examine the impact of surfactants, which are important surface defining components of SLN, on viability and cytokine production by macrophages. Cytotoxicity, as assessed by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) test, was strongly influenced by the surfactant used being marked with cetylpyridinium chloride- (CPC-) coated SLN at a concentration of 0.001% and further increased at SLN concentrations of 0.01 and 0.1%. All other SLN formulations -- containing Poloxamine 908 (P908), Poloxamer 407 (P407), Poloxamer 188 (P188), Solutol HS15 (HS15), Tween 80 (T80), Lipoid S75 (S75), sodium cholate (SC), or sodium dodecylsulfate (SDS) -- when used at the same concentrations reduced cell viability only slightly. None of the SLN formulations tested induced cytokine production but a concentration-dependent decrease of IL-6 production was observed, which appeared to be associated with cytotoxic effects. IL-12 and TNF-alpha were detected neither in supernatants of macrophages treated with SLN at any concentration nor in those of untreated cells. In contrast to the type of surfactant, the size of SLN was found neither to affect cytotoxicity of SLN nor to result in induction or digression of cytokine production by macrophages. In conclusion, testing the effects of surfactants on SLN on activity of macrophages is a prerequisite prior to in vivo use of SLN.

  14. Activity of cytokine-induced killer cells against bone and soft tissue sarcoma

    PubMed Central

    Sangiolo, Dario; Mesiano, Giulia; Gammaitoni, Loretta; Aglietta, Massimo; Grignani, Giovanni

    2014-01-01

    Cytokine-induced killer (CIK) cells are T lymphocytes expanded ex vivo that are endowed with MHC-independent tumoricidal activity. We have recently demonstrated, in a preclinical setting, that CIK cells are active against autologous bone and soft tissue sarcomas. In particular, CIK cells killed a putative sarcoma stem cell population that may underlie disease relapse and chemoresistance. PMID:25050197

  15. Prior TLR5 induction in human T cells results in a transient potentiation of subsequent TCR-induced cytokine production.

    PubMed

    Tremblay, Mikaela M; Bilal, Mahmood Y; Houtman, Jon C D

    2014-02-01

    Activation of TLRs by components required for pathogen viability results in increased inflammation and an enhanced immune response to infection. Unlike their effects on other immune cells, TLR activation in the absence of T cell antigen receptor (TCR) induction has little effect on T cell activity. Instead, the simultaneous induction of TLR and TCR results in increased cytokine release compared to TCR treatment alone. Thus, the current model states that TLRs alter T cell function only if activated at the same time as the TCR. In this study, we tested the novel hypothesis that prior TLR induction can also alter TCR-mediated functions. We found that human T cells responded to ligands for TLR2 and TLR5. However, only prior TLR5 induction potentiated subsequent TCR-mediated cytokine production in human T cells. This response required at least 24h of TLR5 induction and lasted for approximately 24-36h after removal of a TLR5 ligand. Interestingly, prior TLR5 induction enhanced TCR-mediated activation of Akt without increasing Lck, LAT or ERK kinase phosphorylation. Together, our studies show that TLR5 induction leads to a transient increase in the sensitivity of T cells to TCR stimulation by selectively enhancing TCR-mediated Akt function, highlighting that timeframe when TLR5 can potentiate TCR-induced downstream functions are significantly longer that previously appreciated.

  16. Non-LPS components of Chlamydia pneumoniae stimulate cytokine production through Toll-like receptor 2-dependent pathways.

    PubMed

    Netea, Mihai G; Kullberg, Bart Jan; Galama, Jochem M D; Stalenhoef, Anton F H; Dinarello, Charles A; Van der Meer, Jos W M

    2002-04-01

    Recent studies suggest that infection with Chlamydia pneumoniae is associated with atherosclerosis, and that cytokines play an important role in the initiation and progression of Chlamydia-induced inflammation. When freshly isolated peripheral blood mononuclear cells (PBMC) were stimulated for 24 h with sonicated C. pneumoniae, significant amounts of the pro-inflammatory cytokines TNF-alpha and IL-1beta and of the anti-inflammatory cytokine IL-10 were released into the supernatant. The addition of serum increased cytokine release induced by C. pneumonia two- to fivefold (p < 0.01). This effect was not due to complement, mannose-binding lectin (MBL) or lipopolysaccharide-binding protein (LBP). Incubation of PBMC with either anti-Toll-like receptor 4 (TLR4) or anti-CD14 blocking antibodies did not influence the production of cytokines induced by Chlamydia. The induction of cytokines by C. pneumoniae in macrophages from C3H / HeJ mice, known to have a defective TLR4, was identical to that measured in control macrophages from C3H / HeN mice. In contrast, incubation of PBMC with an anti-TLR2 blocking antibody significantly inhibited the production of TNF by 67 % and of IL-1beta by 72 %. In conclusion, C. pneumoniae stimulates cytokine production in a serum-dependent manner, but independently of complement, MBL and LBP. C. pneumoniae induces the pro-inflammatory cytokines TNF and IL-1beta through TLR2, but not TLR4 and CD14.

  17. Effects of tropism and virulence of Leishmania parasites on cytokine production by infected human monocytes

    PubMed Central

    Meddeb-Garnaoui, A; Zrelli, H; Dellagi, K

    2009-01-01

    The nature of early interactions between Leishmania and macrophages which determine the outcome of infection can be related directly to parasite biological properties. Here we compared the capacity of L. major (Lm) strains, reported to be high (LmHV) and low virulent and (LmLV) in the mouse model and L. infantum (Li) strains, dermotropic (LiD) and viscerotropic (LiV), to infect and modulate cytokine production in human peripheral blood derived monocytes. Monocytes were infected with metacyclic promastigotes for 24, 48 and 72 h. Parasite burden was significantly higher in Lm- than in Li-infected monocytes. LmHV and LiD induced a significantly higher parasite burden than LmLV and LiV respectively. Cytokine production was evaluated in monocytes infected for 24 h. Contrary to interleukin (IL)-12p70, monocyte chemotactic protein-1 and transforming growth factor-β production was increased significantly in infected monocytes with no differences between strains. Lm isolates induced significantly higher quantities of tumour necrosis factor (TNF)-α than Li isolates. Low levels of IL-10 were induced by all Leishmania strains and, interestingly, co-stimulation with lipopolysaccharide (LPS) was accompanied by a dramatic increase in IL-10 production by infected monocytes. In conclusion, Lm isolates displaying different levels of virulence in mice exhibited significant differences in parasite burden but similar abilities to modulate cytokine production in human monocytes. Li strains showed weaker infectivity and TNF-α inducing-capacity compared with Lm strains. The dramatic increase of IL-10 production in infected monocytes co-stimulated by LPS may play a role in disease progression considering the presence of LPS during bacterial superinfections observed during human leishmaniasis. PMID:19040614

  18. Macrophages that have ingested apoptotic cells in vitro inhibit proinflammatory cytokine production through autocrine/paracrine mechanisms involving TGF-beta, PGE2, and PAF.

    PubMed Central

    Fadok, V A; Bratton, D L; Konowal, A; Freed, P W; Westcott, J Y; Henson, P M

    1998-01-01

    Apoptosis in vivo is followed almost inevitably by rapid uptake into adjacent phagocytic cells, a critical process in tissue remodeling, regulation of the immune response, or resolution of inflammation. Phagocytosis of apoptotic cells by macrophages has been suggested to be a quiet process that does not lead to production of inflammatory mediators. Here we show that phagocytosis of apoptotic neutrophils (in contrast to immunoglobulin G-opsonized apoptotic cells) actively inhibited the production of interleukin (IL)-1beta, IL-8, IL-10, granulocyte macrophage colony-stimulating factor, and tumor necrosis factor-alpha, as well as leukotriene C4 and thromboxane B2, by human monocyte-derived macrophages. In contrast, production of transforming growth factor (TGF)-beta1, prostaglandin E2, and platelet-activating factor (PAF) was increased. The latter appeared to be involved in the inhibition of proinflammatory cytokine production because addition of exogenous TGF-beta1, prostaglandin E2, or PAF resulted in inhibition of lipopolysaccharide-stimulated cytokine production. Furthermore, anti-TGF-beta antibody, indomethacin, or PAF receptor antagonists restored cytokine production in lipopolysaccharide-stimulated macrophages that had phagocytosed apoptotic cells. These results suggest that binding and/or phagocytosis of apoptotic cells induces active antiinflammatory or suppressive properties in human macrophages. Therefore, it is likely that resolution of inflammation depends not only on the removal of apoptotic cells but on active suppression of inflammatory mediator production. Disorders in either could result in chronic inflammatory diseases. PMID:9466984

  19. Macrophages that have ingested apoptotic cells in vitro inhibit proinflammatory cytokine production through autocrine/paracrine mechanisms involving TGF-beta, PGE2, and PAF.

    PubMed

    Fadok, V A; Bratton, D L; Konowal, A; Freed, P W; Westcott, J Y; Henson, P M

    1998-02-15

    Apoptosis in vivo is followed almost inevitably by rapid uptake into adjacent phagocytic cells, a critical process in tissue remodeling, regulation of the immune response, or resolution of inflammation. Phagocytosis of apoptotic cells by macrophages has been suggested to be a quiet process that does not lead to production of inflammatory mediators. Here we show that phagocytosis of apoptotic neutrophils (in contrast to immunoglobulin G-opsonized apoptotic cells) actively inhibited the production of interleukin (IL)-1beta, IL-8, IL-10, granulocyte macrophage colony-stimulating factor, and tumor necrosis factor-alpha, as well as leukotriene C4 and thromboxane B2, by human monocyte-derived macrophages. In contrast, production of transforming growth factor (TGF)-beta1, prostaglandin E2, and platelet-activating factor (PAF) was increased. The latter appeared to be involved in the inhibition of proinflammatory cytokine production because addition of exogenous TGF-beta1, prostaglandin E2, or PAF resulted in inhibition of lipopolysaccharide-stimulated cytokine production. Furthermore, anti-TGF-beta antibody, indomethacin, or PAF receptor antagonists restored cytokine production in lipopolysaccharide-stimulated macrophages that had phagocytosed apoptotic cells. These results suggest that binding and/or phagocytosis of apoptotic cells induces active antiinflammatory or suppressive properties in human macrophages. Therefore, it is likely that resolution of inflammation depends not only on the removal of apoptotic cells but on active suppression of inflammatory mediator production. Disorders in either could result in chronic inflammatory diseases.

  20. Immune response to the cestode Hymenolepis nana: cytokine production during infection with eggs or cysts.

    PubMed

    Conchedda, M; Bortoletti, G; Gabriele, F; Wakelin, D; Palmas, C

    1997-03-01

    Analysis of cytokine production (IFN-gamma, IL-2, IL-3, IL-4, IL-5) by in vitro Con A-stimulated mesenteric lymph node cells measured daily after egg or cyst infection of mice with Hymenolepis nana showed that cytokine production varies during parasite development and between different host strains (BALB/c and C3H/He mice). Egg infection stimulates a rapid increase in IFN-gamma, independent of mouse strain. In addition, in BALB/c mice a Th2-like response (IL-4, IL-5 secretion) was stimulated 4-5 days p.i., when the parasites are thought to begin their lumenal phase. After infection with cysts significant increases in IFN-gamma, IL-2, IL-4 and IL-5 were observed at the time when autoinfection with eggs is thought to occur. The level of IFN-gamma paralleled that seen after a primary egg infection. This suggests that there is a predominantly Th1-type response during the tissue phase of H. nana development and that, in BALB/c mice, a Th2 polarization occurs during the first few days of the lumenal phase. The cytokine patterns observed are discussed in relation to host responses during chronic helminth infection.

  1. The Traditional Japanese Formula Keishibukuryogan Inhibits the Production of Inflammatory Cytokines by Dermal Endothelial Cells

    PubMed Central

    Yoshihisa, Yoko; Furuichi, Megumi; Ur Rehman, Mati; Ueda, Chieko; Makino, Teruhiko; Shimizu, Tadamichi

    2010-01-01

    Keishibukuryogan (KBG) is one of the traditional herbal formulations widely administered to patients with blood stagnation for improving blood circulation; currently, it is the most frequently prescribed medicine in Japan. KBG has been reported to improve conjunctional microcirculation. The aim of this study was to evaluate the role of KBG and paeoniflorin, a bioactive compound of KBG, in inhibiting the production of inflammatory cytokines using human dermal microvessel endothelial cells (HDMECs). The authors observed that lipopolysaccharide (LPS; 1 μg/mL) stimulated the secretion of proinflammatory cytokines in HDMECs. KBG treatment (10 mg/mL) significantly suppressed the mRNA levels of migration inhibitory factor (MIF), interleukin (IL)-6, IL-8, and tumor necrosis factor (TNF)-α in LPS-stimulated cultured HDMECs. Similarly, paeoniflorin significantly suppressed the mRNA levels of these cytokines in LPS-stimulated cultured HDMECs. ELISA showed that KBG and paeoniflorin suppressed the production of MIF, IL-6, IL-8, and TNF-α in LPS-stimulated HDMECs. Moreover, KBG and paeoniflorin decreased the expression of cyclooxygenase-2 and inducible nitric oxide synthase (iNOS) in these cells. These results suggest that KBG may be useful for improving microvascular inflammation in patients with skin diseases. PMID:21253500

  2. Recombinant CC16 protein inhibits the production of pro-inflammatory cytokines via NF-κB and p38 MAPK pathways in LPS-activated RAW264.7 macrophages.

    PubMed

    Pang, Min; Yuan, Yangyang; Wang, Dong; Li, Ting; Wang, Dan; Shi, Xiaohong; Guo, Min; Wang, Chunfang; Zhang, Xinri; Zheng, Guoping; Yu, Baofeng; Wang, Hailong

    2017-03-17

    Accumulating evidence indicates that Clara cell protein-16 (CC16) has anti-inflammatory functions, although the involved molecular pathways have not been completely elucidated. Here, we evaluated the effect of recombinant rat CC16 (rCC16) on the expression of tumor necrosis factor alpha (TNF-α), interleukin-6 (IL-6), and IL-8 in lipopolysaccharide (LPS)-stimulated mouse macrophages (RAW264.7 cells) and explored the underlying molecular mechanisms. It was found that rCC16 inhibited LPS-induced TNF-α, IL-6, and IL-8 expression at both the messenger ribonucleicacid (mRNA) level and protein level in a concentration-dependent manner, as demonstrated by real-time reverse transcriptase-polymerase chain reaction and enzyme-linked immunosorbent assay. Such suppressive effects were accompanied by the inhibition of transcriptional activity and the deoxyribonucleic acid binding activity of nuclear factor (NF)-κB but not activator protein (AP)-1. Western blot analysis further revealed that rCC16 inhibited the increase of nuclear NF-κB and the reduction of cytosolic NF-κB, the phosphorylation and reduction of NF-κB inhibitory protein IκBα, and the p38 mitogen-activated protein kinase (MAPK)-dependent NF-κB activation by phosphorylation at Ser276 of its p65 subunit. Furthermore, rCC16 was found to have no effect on the phosphorylation of c-Jun N-terminal kinase, c-Jun, or the nuclear translocation of c-Jun. In addition, reduction of TNF-α, IL-6, and IL-8 were reversed when the level of endogenous uteroglobin-binding protein was reduced by RNA interference in rCC16- and LPS-treated RAW264.7 cells. Our data suggest that rCC16 suppresses LPS-mediated inflammatory mediator TNF-α, IL-6, and IL-8 production by inactivating NF-κB and p38 MAPK but not AP-1 in RAW264.7 cells.

  3. Gliadin-dependent cytokine production in a bidimensional cellular model of celiac intestinal mucosa.

    PubMed

    Vincentini, Olimpia; Maialetti, Francesca; Gonnelli, Elena; Silano, Marco

    2015-11-01

    The downstream cascade of the inflammatory response to gliadin in celiac intestinal mucosa encompasses the early activation of the innate immunity that triggers the adaptive response. Therefore, the in vitro study of the pathogenic mechanism of celiac disease (CD) on enterocytes alone or mucosal T lymphocytes alone does not fully consider all the aspects of gliadin-dependent inflammation. Although the in vitro culture of specimens of intestinal mucosa obtained from celiac patients is the gold standard for the study of CD, this technique presents several technical challenges and the bioptic specimens are not easily available. So, in this paper, we described the gliadin-dependent cytokine production in a bidimensional cellular system, which is able to mimic both the innate and the adaptive steps of the mucosal immune response of CD. In the upper compartment, the intestinal epithelial cells are grown on a filter, and in the lower compartment, the mononuclear cells isolated from peripheral blood of celiac patients are cultured. Cells were apically exposed to the toxic gliadin peptide p31-43 for 3 h and then with the immunodominant gliadin fragment pα-9 for 21 h. The incubation with gliadin peptides resulted in increased levels of IL-15, INF-γ, IL-6, tumor necrosis factor (TNF)-α, IL-1β, and CCL 2, 3 and 4 in the basal supernatants, with respect to cells exposed to medium alone. The p31-43-driven epithelial priming of mucosal response consists of transglutaminase (TG2)-mediated deamidation of the immunostimulatory gliadin peptides, as demonstrated by the inhibition of pα-9 activity, when the system is exposed to blocking anti-TG2 antibody.

  4. Proliferation and TH1/TH2 cytokine production in human peripheral blood mononuclear cells after treatment with cypermethrin and mancozeb in vitro.

    PubMed

    Mandarapu, Rajesh; Ajumeera, Rajanna; Venkatesan, Vijayalakshmi; Prakhya, Balakrishna Murthy

    2014-01-01

    In recent times, human cell-based assays are gaining attention in assessments of immunomodulatory effects of chemicals. In the study here, the possible effects of cypermethrin and mancozeb on lymphocyte proliferation and proinflammatory (tumor necrosis factor (TNF-) α) and immunoregulatory cytokine (interferon- (IFN-) γ, interleukins (IL) 2, 4, 6, and 10) formation in vitro were investigated. Human peripheral blood mononuclear cells (PBMC) were isolated and exposed for 6 hr to noncytotoxic doses (0.45-30 µM) of cypermethrin or mancozeb in the presence of activating rat S9 fraction. Cultures were then further incubated for 48 or 72 hr in fresh medium containing phytohemagglutinin (10 µg/mL) to assess, respectively, effects on cell proliferation (BrdU-ELISA method) and cytokine formation (flow cytometric bead immunoassays). Mancozeb induced dose-dependent increases in lymphocyte proliferation, inhibition of production of TNFα and the TH2 cytokines IL-6 and IL-10, and an increase in IFNγ (TH1 cytokine) production (at least 2-fold compared to control); mancozeb also induced inhibition of IL-4 (TH2) and stimulated IL-2 (TH1) production, albeit only in dose-related manners for each. In contrast, cypermethrin exposure did not cause significant effects on proliferation or cytokine profiles. Further studies are needed to better understand the functional significance of our in vitro findings.

  5. Participation of substance P distribution in the cytokine production of rheumatoid synovium.

    PubMed

    Komuro, H; Tanabe, T; Ogushi, M; Takemura, S; Toda, Y; Morimoto, T; Akagi, S; Ogawa, R

    2000-06-01

    Abstract Based on findings which suggested the involvement of the neuropeptide substance P in the pathogenesis of rheumatoid arthritis (RA), we investigated the mechanism of synovial pannus formation in RA, and examined the interaction between the cytokine production of synovial tissues and the concentration of substance P in the cartilage-pannus junction (CPJ). The CPJ and other peripheral synovial tissues were separately obtained from each part of the synovium from the knee joints of seven RA patients. The concentrations of substance P and the cytokines interleukin (IL)-1β and IL-6 in the CPJ and peripheral synovial tissues were determined by enzyme-linked immunosorbent assays. In addition, synovial cells were isolated from the CPJ and peripheral synovial tissues and treated with substance P or neurokinin-1 receptor antagonist to analyze the changes in cytokine production. The substance P levels were 211.2 and 50.5 pg/mg protein in the CPJ and the peripheral synovium, respectively. The IL-1β and IL-6 levels in the CPJ were 24.6 and 12.8 pg/mg protein, respectively. In the peripheral synovium, these levels were 4.3 and 2.5 pg/mg protein, respectively. In the CPJ, the IL-1β and IL-6 levels in tissue containing a high concentration of substance P (>200 pg/mg protein) were 39.4 and 21.6 pg/mg protein, respectively, and those in tissue containing a low concentration of substance P (≤200 pg/mg protein) were 11.6 and 5.1 pg/mg protein, respectively. Synovial cells from the CPJ produced higher levels of IL-1β and IL-6 than those from peripheral tissues. In addition, treatment of the cells with an NK-1 antagonist significantly reduced the production of these cytokines by the synovial cells. The theory that substance P plays a role in the pathogenesis of RA via the upregulation of cytokine production should be considered in further studies on the immunomodulatory properties of substance P in arthritis.

  6. Controlled meal frequency without caloric restriction alters peripheral blood mononuclear cell cytokine production

    PubMed Central

    2011-01-01

    Background Intermittent fasting (IF) improves healthy lifespan in animals by a mechanism involving reduced oxidative damage and increased resistance to stress. However, no studies have evaluated the impact of controlled meal frequency on immune responses in human subjects. Objective A study was conducted to establish the effects of controlled diets with different meal frequencies, but similar daily energy intakes, on cytokine production in healthy male and female subjects. Design In a crossover study design with an intervening washout period, healthy normal weight middle-age male and female subjects (n = 15) were maintained for 2 months on controlled on-site one meal per day (OMD) or three meals per day (TMD) isocaloric diets. Serum samples and peripheral blood mononuclear cells (PBMCs) culture supernatants from subjects were analyzed for the presence of inflammatory markers using a multiplex assay. Results There were no significant differences in the inflammatory markers in the serum of subjects on the OMD or TMD diets. There was an increase in the capacity of PBMCs to produce cytokines in subjects during the first month on the OMD or TMD diets. Lower levels of TNF-α, IL-17, MCP-1 and MIP-1β were produced by PBMCs from subjects on the OMD versus TMD diet. Conclusions PBMCs of subjects on controlled diets exhibit hypersensitivities to cellular stimulation suggesting that stress associated with altered eating behavior might affect cytokine production by immune cells upon stimulation. Moreover, stimulated PBMCs derived from healthy individuals on a reduced meal frequency diet respond with a reduced capability to produce cytokines. PMID:21385360

  7. Cytosolic dsDNA triggers apoptosis and pro-inflammatory cytokine production in normal human melanocytes.

    PubMed

    Wang, Suiquan; Liu, Dongyin; Ning, Weixuan; Xu, Aie

    2015-04-01

    Considerable evidence implicates that viral infection might be a participant factor in the pathogenesis of vitiligo. However, it is still unclear how viral infection leads to the melanocyte destruction. To elucidate the effects of viral dsDNA on the viability and cytokine synthesis of normal human melanocytes and to explore the underlying mechanisms, primary cultured normal human melanocytes were transfected with poly(dA:dT). The results demonstrated that poly(dA:dT) triggered apoptosis instead of pyroptosis in melanocytes. Knocking down AIM2 or RIG-I by RNA interference partially reduced the poly(dA:dT)-induced LDH release, suggesting the involvement of both nucleic acid sensors in the process of melanocyte death. Poly(dA:dT) induced the expression of pro-inflammatory cytokine genes including IFN-β, TNF-α, IL-6 and IL-8 as well, whereas the pro-inflammatory cytokine production was suppressed by RIG-I siRNA, but not by AIM2 siRNA. Poly(dA:dT) treatment increased the phosphorylation of p38 and JNK and NFκB. Accordingly, NFκB inhibitor Bay 11-7082 and JNK inhibitor SP600125 blocked the induction of the cytokine genes except IFN-β. The production of IL6 and IL8 was also suppressed by p38 inhibitor SB203580. On the contrary, the Poly(dA:dT)-induced melanocyte death was only decreased by SP600125. This study provides the possible mechanism of melanocyte destruction and immuno-stimulation in vitiligo by innate immune response following viral infection.

  8. [Production of recombinant human interleukin-38 and its inhibitory effect on the expression of proinflammatory cytokines in THP-1 cells].

    PubMed

    Yuan, X L; Li, Y; Pan, X H; Zhou, M; Gao, Q Y; Li, M C

    2016-01-01

    Interleukin (IL)-38 is the latest member of the IL-1 cytokine family. However, as a result of lacking efficient method to generate relatively large quantity of IL-38, its precise functions are poorly understood. In the present study, the cloning, expression, purification, and activity analysis of recombinant human IL-38 was described. Human IL-38 cDNA was cloned into the prokaryotic expression vector pET-44. The recombinant IL-38 containing a C-hexahistidine tag was expressed in Escherichia coli BL21 (DE3) which induced by isopropyl-β-D-thiogalactoside. The expressed fusion protein was purified by Ni-NTA affinity chromatography. IL-38 protein was largely found in the soluble fraction. The purified IL-38 appeared a single band on SDS-PAGE, the yield of IL-38 was 4 mg from 1 L of bacterial culture, and the purity was more than 98% with low endotoxin level (<0.1 EU/μg). Western blotting confirmed the identity of the purified protein. Activity analysis showed that IL-38 can inhibit effectively the expression of proinflammatory cytokines, such as tumor necrosis factor-α, IL-1β, IL-17, and monocyte chemoattractant protein-1 in lipopolysaccharide-activated THP-1 cells. The production and characterization of biologically active IL-38 will be beneficial for its potential role in clinical applications.

  9. 5-Hydroxytryptamine modulates cytokine and chemokine production in LPS-primed human monocytes via stimulation of different 5-HTR subtypes.

    PubMed

    Dürk, Thorsten; Panther, Elisabeth; Müller, Tobias; Sorichter, Stephan; Ferrari, Davide; Pizzirani, Cinzia; Di Virgilio, Francesco; Myrtek, Daniel; Norgauer, Johannes; Idzko, Marco

    2005-05-01

    The neurotransmitter 5-hydroxytryptamine (5-HT), commonly known as serotonin, is released at peripheral sites from activated enterochromaffin cells, mast cells and platelets. In this study we analyzed the biological activity and intracellular signaling of 5-HT in human monocytes. By reverse transcription (RT) and PCR, messenger RNA (mRNA) expression of 5-HT receptor 1E (5-HTR(1E)), 5-HTR(2A), 5-HTR(3), 5-HTR(4) and 5-HTR(7) could be revealed. Functional studies showed that 5-HT modulates the release of IL-1beta, IL-6, IL-8/CXCL8, IL-12p40 and tumor necrosis factor-alpha (TNF-alpha), while it has no effect on the production of IL-18 and IFN-gamma in LPS-stimulated human blood monocytes. Moreover, RT and PCR revealed that 5-HT modulated mRNA levels of IL-6 and IL-8/CXCL8, but did not influence mRNA levels of IL-1beta and TNF-alpha. Pharmacological studies with isotype-selective receptor agonists allowed us to show that 5-HTR(3) subtype up-regulates the LPS-induced production of IL-1beta, IL-6 and IL-8/CXCL8, while it was not involved in TNF-alpha and IL-12p40 secretion. Furthermore, activation of the G(s)-coupled 5-HTR(4) and 5-HTR(7) subtypes increased intracellular cyclic AMP (cAMP) and secretion of IL-1beta, IL-6, IL-12p40 and IL-8/CXCL8, while, on the contrary, it inhibited LPS-induced TNF-alpha release. Interestingly, 5-HTR(1) and 5-HTR(2) agonists did not modulate the LPS-induced cytokine production in human monocytes. Our results point to a new role for 5-HT in inflammation by modulating cytokine production in monocytes via activation of 5-HTR(3), 5-HTR(4) and 5-HTR(7) subtypes.

  10. Expression of PD-1/LAG-3 and cytokine production by CD4(+) T cells during infection with Plasmodium parasites.

    PubMed

    Doe, Henrietta T; Kimura, Daisuke; Miyakoda, Mana; Kimura, Kazumi; Akbari, Masoud; Yui, Katsuyuki

    2016-02-01

    CD4(+) T cells play critical roles in protection against the blood stage of malarial infection; however, their uncontrolled activation can be harmful to the host. In this study, in which rodent models of Plasmodium parasites were used, the expression of inhibitory receptors on activated CD4(+) T cells and their cytokine production was compared with their expression in a bacterial and another protozoan infection. CD4(+) T cells from mice infected with P. yoelii 17XL, P yoelii 17XNL, P. chabaudi, P. vinckei and P. berghei expressed the inhibitory receptors, PD-1 and LAG-3, as early as 6 days after infection, whereas those from either Listeria monocytogenes- or Leishmania major-infected mice did not. In response to T-cell receptor stimulation, CD4(+) T cells from mice infected with all the pathogens under study produced high concentrations of IFN-γ. IL-2 production was reduced in mice infected with Plasmodium species, but not in those infected with Listeria or Leishmania. In vitro blockade of the interaction between PD-1 and its ligands resulted in increased IFN-γ production in response to Plasmodium antigens, implying that PD-1 expressed on activated CD4(+) T cells actively inhibits T cell immune responses. Studies using Myd88(-/-), Trif(-/-) and Irf3(-/-) mice showed that induction of these CD4(+) T cells and their ability to produce cytokines is largely independent of TLR signaling. These studies suggest that expression of the inhibitory receptors PD-1 and LAG-3 on CD4(+) T cells and their reduced IL-2 production are common characteristic features of Plasmodium infection.

  11. Differing effects of clarithromycin and azithromycin on cytokine production by murine dendritic cells

    PubMed Central

    Sugiyama, K; Shirai, R; Mukae, H; Ishimoto, H; Nagata, T; Sakamoto, N; Ishii, H; Nakayama, S; Yanagihara, K; Mizuta, Y; Kohno, S

    2007-01-01

    The macrolide antibiotics are now well known to have anti-inflammatory effects. Because dendritic cells (DCs) orchestrate immune responses, we examined the in vitro effects of clarithromycin (CAM), azithromycin (AZM) and midecamycin (MDM) on the expression of co-stimulatory molecules and production of cytokines [interleukin (IL)-10, IL-6, interferon (IFN)-γ, IL-12p40, tumour necrosis factor (TNF)-α] of murine bone marrow-derived DCs by lipopolysaccharide (LPS) stimulation. A 15-membered macrolide, AZM, and a 14-membered macrolide, CAM, significantly enhanced the intensity of a co-stimulatory molecule, CD80, on DCs but not CD86 and CD40. AZM significantly increased the production of IL-10 and CAM significantly inhibited the production of IL-6 by DCs. However, a 16-membered macrolide, MDM, did not have any significant effect on these surface markers and cytokine productions. Moreover, AZM increased IL-10 and CAM decreased IL-2 productions significantly, when naive T cells derived from spleen were co-cultured with DCs treated in advance with LPS and these macrolides. These findings suggest that 14-membered and 15-membered, but not 16-membered macrolides play as anti-inflammatory agents, at least in part, through modulating the functions of DCs. However, each macrolide affects them in different ways. PMID:17302905

  12. Up-regulation of T lymphocyte and antibody production by inflammatory cytokines released by macrophage exposure to multi-walled carbon nanotubes

    NASA Astrophysics Data System (ADS)

    Grecco, Ana Carolina P.; Paula, Rosemeire F. O.; Mizutani, Erica; Sartorelli, Juliana C.; Milani, Ana M.; Longhini, Ana Leda F.; Oliveira, Elaine C.; Pradella, Fernando; Silva, Vania D. R.; Moraes, Adriel S.; Peterlevitz, Alfredo C.; Farias, Alessandro S.; Ceragioli, Helder J.; Santos, Leonilda M. B.; Baranauskas, Vitor

    2011-07-01

    Our data demonstrate that multi-walled carbon nanotubes (MWCNTs) are internalized by macrophages, subsequently activating them to produce interleukin (IL)-12 (IL-12). This cytokine induced the proliferative response of T lymphocytes to a nonspecific mitogen and to ovalbumin (OVA). This increase in the proliferative response was accompanied by an increase in the expression of pro-inflammatory cytokines, such as interferon-gamma (IFNγ), tumor necrosis factor-alpha (TNFα) and IL-6, in mice inoculated with MWCNTs, whether or not they had been immunized with OVA. A decrease in the expression of transforming growth factor-beta (TGFβ) was observed in the mice treated with MWCNTs, whereas the suppression of the expression of both TGFβ and IL-10 was observed in mice that had been both treated and immunized. The activation of the T lymphocyte response by the pro-inflammatory cytokines leads to an increase in antibody production to OVA, suggesting the important immunostimulatory effect of carbon nanotubes.

  13. Up-regulation of T lymphocyte and antibody production by inflammatory cytokines released by macrophage exposure to multi-walled carbon nanotubes.

    PubMed

    Grecco, Ana Carolina P; Paula, Rosemeire F O; Mizutani, Erica; Sartorelli, Juliana C; Milani, Ana M; Longhini, Ana Leda F; Oliveira, Elaine C; Pradella, Fernando; Silva, Vania D R; Moraes, Adriel S; Peterlevitz, Alfredo C; Farias, Alessandro S; Ceragioli, Helder J; Santos, Leonilda M B; Baranauskas, Vitor

    2011-07-01

    Our data demonstrate that multi-walled carbon nanotubes (MWCNTs) are internalized by macrophages, subsequently activating them to produce interleukin (IL)-12 (IL-12). This cytokine induced the proliferative response of T lymphocytes to a nonspecific mitogen and to ovalbumin (OVA). This increase in the proliferative response was accompanied by an increase in the expression of pro-inflammatory cytokines, such as interferon-gamma (IFNγ), tumor necrosis factor-alpha (TNFα) and IL-6, in mice inoculated with MWCNTs, whether or not they had been immunized with OVA. A decrease in the expression of transforming growth factor-beta (TGFβ) was observed in the mice treated with MWCNTs, whereas the suppression of the expression of both TGFβ and IL-10 was observed in mice that had been both treated and immunized. The activation of the T lymphocyte response by the pro-inflammatory cytokines leads to an increase in antibody production to OVA, suggesting the important immunostimulatory effect of carbon nanotubes.

  14. Epithelial neutrophil activating peptide-78: a novel chemotactic cytokine for neutrophils in arthritis.

    PubMed Central

    Koch, A E; Kunkel, S L; Harlow, L A; Mazarakis, D D; Haines, G K; Burdick, M D; Pope, R M; Walz, A; Strieter, R M

    1994-01-01

    We and others have shown that cells obtained from inflamed joints of rheumatoid arthritis (RA) patients produce interleukin-8, a potent chemotactic cytokine for neutrophils (PMNs). However, IL-8 accounted for only 40% of the chemotactic activity for PMNs found in these synovial fluids. Currently, we have examined the production of the novel PMN chemotactic cytokine, epithelial neutrophil activating peptide-78 (ENA-78), using peripheral blood, synovial fluid, and synovial tissue from 70 arthritic patients. RA ENA-78 levels were greater in RA synovial fluid (239 +/- 63 ng/ml) compared with synovial fluid from other forms of arthritis (130 +/- 118 ng/ml) or osteoarthritis (2.6 +/- 1.8 ng/ml) (P < 0.05). RA peripheral blood ENA-78 levels (70 +/- 26 ng/ml) were greater than normal peripheral blood levels (0.12 +/- 0.04 ng/ml) (P < 0.05). Anti-ENA-78 antibodies neutralized 42 +/- 9% (mean +/- SE) of the chemotactic activity for PMNs found in RA synovial fluids. Isolated RA synovial tissue fibroblasts in vitro constitutively produced significant levels of ENA-78, and this production was further augmented when stimulated with tumor necrosis factor-alpha (TNF-alpha). In addition RA and osteoarthritis synovial tissue fibroblasts as well as RA synovial tissue macrophages were found to constitutively produce ENA-78. RA synovial fluid mononuclear cells spontaneously produced ENA-78, which was augmented in the presence of lipopolysaccharide. Immunohistochemical localization of ENA-78 from the synovial tissue of patients with arthritis or normal subjects showed that the predominant cellular source of this chemokine was synovial lining cells, followed by macrophages, endothelial cells, and fibroblasts. Synovial tissue macrophages and fibroblasts were more ENA-78 immunopositive in RA than in normal synovial tissue (P < 0.05). These results, which are the first demonstration of ENA-78 in a human disease state, suggest that ENA-78 may play an important role in the recruitment of PMNs

  15. Toll-like receptor agonists partially restore the production of pro-inflammatory cytokines and type I interferon in Sézary syndrome

    PubMed Central

    Manfrere, Kelly C. G.; Torrealba, Marina P.; Miyashiro, Denis R.; Oliveira, Luanda M. S.; de Carvalho, Gabriel C.; Lima, Josenilson F.; Branco, Anna Claudia C. C.; Pereira, Nátalli Z.; Pereira, Juliana; Sanches, José A.; Sato, Maria N.

    2016-01-01

    Sézary syndrome (SS) carries a poor prognosis, and infections represent the most frequent cause of death in SS patients. Toll-like receptors (TLRs) are a family of innate immune receptors that induce protective immune responses against infections. We sought to evaluate the ability of TLR agonists to induce inflammatory cytokine, Th2 cytokine, and type I interferon (IFN-I) production by peripheral blood mononuclear cells (PBMC) of untreated SS patients. We detected impaired IL-6, IL-10 and IL-13 secretion by PBMC induced by the agonists for TLR5, TLR3, TLR7 and TLR9 in SS patients, while it was partially recovered by TLR2/TLR4 and TLR7/8 agonists TNF secretion was restored following stimulation with TLR2/TLR4 agonists. IFN-γ was scarcely produced upon TLR activation in SS cells, albeit TLR 7/8 (CL097) enhanced their secretion at lower levels than the control group. TLR9 agonist efficiently induced IFN-I in SS patients, although this positive regulation was not observed for other cytokines, in direct contrast to the broad activity of CL097. Among the TLR agonists, TLR4 was able to induce pro-inflammatory, IL-10 and Th2 secretion, while TLR7-8 agonist induced the inflammatory cytokines, IFN-I and IFN-γ. These findings reveal a dysfunctional cytokine response upon both extracellular and intracellular TLR activation in SS patients, which was partially restored by TLRs agonists. PMID:27780938

  16. Altered cytokine production by specific human peripheral blood cell subsets immediately following space flight

    NASA Technical Reports Server (NTRS)

    Crucian, B. E.; Cubbage, M. L.; Sams, C. F.

    2000-01-01

    In this study, flow cytometry was used to positively identify the specific lymphocyte subsets exhibiting space flight-induced alterations in cytokine production. Whole blood samples were collected from 27 astronauts at three points (one preflight, two postflight) surrounding four space shuttle missions. Assays performed included serum/urine stress hormones, white blood cell (WBC) phenotyping, and intracellular cytokine production following mitogenic stimulation. Absolute levels of peripheral granulocytes were significantly elevated following space flight, but the levels of circulating lymphocytes and monocytes were unchanged. Lymphocyte subset analysis demonstrated a decreased percentage of T cells, whereas percentages of B cells and natural killer (NK) cells remained unchanged after flight. Nearly all the astronauts exhibited an increased CD4/CD8 T cell ratio. Assessment of naive (CD45RA+) vs. memory (CD45RO+) CD4+ T cell subsets was ambiguous, and subjects tended to group within specific missions. Although no significant trend was seen in absolute monocyte levels, a significant decrease in the percentage of the CD14+ CD16+ monocytes was seen following space flight in all subjects tested. T cell (CD3+) production of interleukin-2 (IL-2) was significantly decreased after space flight, as was IL-2 production by both CD4+ and CD8+ T cell subsets. Production of interferon-gamma (IFN-gamma) was not altered by space flight for the CD8+ cell subset, but there was a significant decrease in IFN-gamma production for the CD4+ T cell subset. Serum and urine stress hormone analysis indicated significant physiologic stresses in astronauts following space flight. Altered peripheral leukocyte subsets, altered serum and urine stress hormone levels, and altered T cell cytokine secretion profiles were all observed postflight. In addition, there appeared to be differential susceptibility to space flight regarding cytokine secretion by T cell subsets. These alterations may be the

  17. Leishmania pifanoi proteoglycolipid complex P8 induces macrophage cytokine production through Toll-like receptor 4.

    PubMed

    Whitaker, Shanta M; Colmenares, Maria; Pestana, Karen Goldsmith; McMahon-Pratt, Diane

    2008-05-01

    The P8 proteoglycolipid complex (P8 PGLC) is a glyconjugate expressed by Leishmania mexicana complex parasites. We previously have shown that vaccination with P8 PGLC provides protection against cutaneous leishmaniasis in susceptible BALB/c mice. However, the biological importance of this complex remains unknown. Here we show that P8 PGLC localizes to the surface of Leishmania pifanoi amastigotes and that upon exposure to macrophages, P8 PGLC binds and induces inflammatory cytokine and chemokine mRNAs such as tumor necrosis factor alpha and RANTES early after stimulation. Our studies indicate that cytokine and chemokine induction is dependent upon Toll-like receptor 4 (TLR4). Interestingly, key inflammatory cytokines and chemokines (such as interleukin-6 [IL-6], macrophage inflammatory protein 1beta, and beta interferon [IFN-beta]) that can be induced through TLR4 activation were not induced or only slightly upregulated by P8 PGLC. Activation by P8 PGLC does not occur in the presence of TLR4 alone and requires both CD14 and myeloid differentiation protein 2 for signaling; this requirement may be responsible for the limited TLR4 response. This is the first characterization of a TLR4 ligand for Leishmania. In vitro experiments indicate that L. pifanoi amastigotes induce lower levels of cytokines in macrophages in the absence of TLR4; however, notably higher IL-10/IFN-gamma ratios were found for TLR4-deficient mice than for BALB/c mice. Further, increased levels of parasites persist in BALB/c mice deficient in TLR4. Taken together, these results suggest that TLR4 recognition of Leishmania pifanoi amastigotes is important for the control of infection and that this is mediated, in part, through the P8 PGLC.

  18. Effect of leptin on activation and cytokine synthesis in peripheral blood lymphocytes of malnourished infected children

    PubMed Central

    Rodríguez, L; Graniel, J; Ortiz, R

    2007-01-01

    Malnutrition compromises immune function, resulting in reduced resistance to infection. Recent animal and human studies have suggested that leptin is capable of modulating the immune response and that its levels, which are regulated by nutritional status, fall rapidly during starvation. Leptin deficiency is associated with impaired cell-mediated immunity, an increased incidence of infectious disease and an associated increase in mortality. The purpose of this study was to examine the effect of leptin on activation and cytokine production in peripheral blood T cells from malnourished children. The data obtained in the present study demonstrate that leptin produced an increase in the percentage of CD4+ and CD8+ cells producing interleukin (IL)-2 and interferon (IFN)-γ in 24-h cultures. Moreover, leptin decreased the percentage of CD4+ and CD8+ cells producing IL-4 and IL-10, and enhanced activation of circulating T cells when co-stimulated by phorbol 12-myristate 13 acetate (PMA)–ionomycin. Leptin enhanced the expression of activation markers CD69 and CD25 in both CD4+ and CD8+ cells after 5 h of stimulation. In conclusion, the results obtained show that leptin modulates CD4+ and CD8+ cell activation towards a T helper 1 (Th1) phenotype by stimulating the synthesis of IL-2 and IFN-γ. In contrast, leptin decreases IL-4 and IL-10 production. Moreover, leptin enhanced the expression of CD69 and CD25 on CD4+ and CD8+ cells after stimulation with PMA–ionomycin. PMID:17355247

  19. Combined immunomodulator and antimicrobial therapy eliminates polymicrobial sepsis and modulates cytokine production in combined injured mice

    PubMed Central

    Elliott, Thomas B.; Bolduc, David L.; Ledney, G. David; Kiang, Juliann G.; Fatanmi, Oluseyi O.; Wise, Stephen Y.; Romaine, Patricia L. P.; Newman, Victoria L.; Singh, Vijay K.

    2015-01-01

    Purpose: A combination therapy for combined injury (CI) using a non-specific immunomodulator, synthetic trehalose dicorynomycolate and monophosphoryl lipid A (STDCM-MPL), was evaluated to augment oral antimicrobial agents, levofloxacin (LVX) and amoxicillin (AMX), to eliminate endogenous sepsis and modulate cytokine production. Materials and methods: Female B6D2F1/J mice received 9.75 Gy cobalt-60 gamma-radiation and wound. Bacteria were isolated and identified in three tissues. Incidence of bacteria and cytokines were compared between treatment groups. Results: Results demonstrated that the lethal dose for 50% at 30 days (LD50/30) of B6D2F1/J mice was 9.42 Gy. Antimicrobial therapy increased survival in radiation-injured (RI) mice. Combination therapy increased survival after RI and extended survival time but did not increase survival after CI. Sepsis began five days earlier in CI mice than RI mice with Gram-negative species predominating early and Gram-positive species increasing later. LVX plus AMX eliminated sepsis in CI and RI mice. STDCM-MPL eliminated Gram-positive bacteria in CI and most RI mice but not Gram-negative. Treatments significantly modulated 12 cytokines tested, which pertain to wound healing or elimination of infection. Conclusions: Combination therapy eliminates infection and prolongs survival time but does not assure CI mouse survival, suggesting that additional treatment for proliferative-cell recovery is required. PMID:25994812

  20. High insulin and leptin increase resistin and inflammatory cytokine production from human mononuclear cells.

    PubMed

    Tsiotra, Panayoula C; Boutati, Eleni; Dimitriadis, George; Raptis, Sotirios A

    2013-01-01

    Resistin and the proinflammatory cytokines, such as TNF- α , IL-6, and IL-1 β , produced by adipocytes, and macrophages, are considered to be important modulators of chronic inflammation contributing to the development of obesity and atherosclerosis. Human monocyte-enriched mononuclear cells, from ten healthy individuals, were exposed to high concentrations of insulin, leptin, and glucose (alone or in combination) for 24 hours in vitro. Resistin, TNF- α , IL-6, and IL-1 β production was examined and compared to that in untreated cells. High insulin and leptin concentrations significantly upregulated resistin and the cytokines. The subsequent addition of high glucose significantly upregulated resistin and TNF- α mRNA and protein secretion, while it did not have any effect on IL-6 or IL-1 β production. By comparison, exposure to dexamethasone reduced TNF- α , IL-6, and IL-1 β production, while at this time point it increased resistin protein secretion. These data suggest that the expression of resistin, TNF- α , IL-6, and IL-1 β from human mononuclear cells, might be enhanced by the hyperinsulinemia and hyperleptinemia and possibly by the hyperglycemia in metabolic diseases as obesity, type 2 diabetes, and atherosclerosis. Therefore, the above increased production may contribute to detrimental effects of their increased adipocyte-derived circulating levels on systemic inflammation, insulin sensitivity, and endothelial function of these patients.

  1. The effect of Jeo Dang-Tang on cytokines production in the patients with cerebral infarction.

    PubMed

    Jeong, Hyun-Ja; Kang, Sei-Young; Kim, Sang-Yong; Lee, Sang-Gwan; Lee, Sung-Geun; Sung, Kang-Keyng; Kim, Hyung-Min

    2003-11-01

    The herbal formulation "Jeo Dang-Tang" (JDT) has long been used for various cerebrovascular diseases. However, very little has scientific investigation been carried out. The aim of the present study is to investigate the effect of JDT on the production of various cytokines in the patients with cerebral infarction (CI). Peripheral blood mononuclear cells (PBMC) obtained from the patients with CI were cultured for 24h in the presence or absence of lipopolysaccharide (LPS) or phytohemagglutinin (PHA). The amount of interleukin (IL)-4, IL-10 and transforming growth factor (TGF)-1beta, in culture supernatant, was significantly increased in the JDT, LPS or PHA treated cells compared to unstimulated cells (P < 0.05). We also show that increased IL-4, and IL-10 level by LPS or PHA was significantly inhibited by JDT in a dose-dependent manner. Maximal inhibition rate of IL-4 and IL-10 production by JDT was 45 +/- 2% and 51 +/- 5% for LPS-stimulated cell and 41.5 +/- 3% and 70.8 +/- 2% for PHA-stimulated cells, respectively (P < 0.05). On the other hand, JDT significantly increased the LPS or PHA-induced TGF-beta1 production (P < 0.05). These data suggest that JDT has a regulatory effect on the cytokines production, which might explain its beneficial effect in the treatment of CI.

  2. CARD15 gene overexpression reduces effect of etanercept, infliximab, and adalimumab on cytokine secretion from PMA activated U937 cells.

    PubMed

    Teimourian, Shahram; Masoudzadeh, Nooshin

    2015-09-05

    Crohn's disease (CD), a subcategory of inflammatory bowel disease, is an immune-related disorder characterized by inflammation of the gastrointestinal mucosa, which can take place in any region along the alimentary tract. The most important gene involved in the etiology of CD is NOD2/CARD15 located on chromosome 16. It has been shown that CARD15 is overexpressed in monocytes of CD patients. The common treatment for the disease is anti-TNF-alpha drugs, the most hopeful of which are probably infliximab and etanercept. Infliximab rapidly reduces signs and symptoms of active Crohn's disease. In contrast, etanercept shows no such effect. In the present study, we evaluated the effects of the CARD15 gene overexpression in monocytic cell line U937 in the production of anti-inflammatory cytokine, IL-10, and proinflammatory cytokine, Il-1 beta, produced after incubation with infliximab, adalimumab, and etanercept separately. Our results show that infliximab and adalimumab significantly decreased IL-10 and IL-1beta secretion levels. However, etanercept inhibition of secretion was less compared with infliximab or adalimumab. In all three cases, suppression of cytokine production is reduced by CARD15 overexpression.

  3. [Cytokines in bone diseases. What is cytokine?].

    PubMed

    Murakami, Yousuke; Kohsaka, Hitoshi

    2010-10-01

    Cytokines have an essential role for cell-cell communication. They can regulate cell proliferation, differentiation, survival, and function. Interaction of cell surface receptor with cytokines is necessary for control of physiological responses. Activation of cytokine receptors transduces specific signal in the receptor-expressing cells, resulting that cytokines can regulate specific cell population. Thus, cytokines contribute directly or indirectly to morphogenesis, host defense and immune response, play critical roles for homeostasis and development.

  4. A correlation of pregnancy term, disease activity, serum female hormones, and cytokines in uveitis

    PubMed Central

    Chan, C-C; Reed, G F; Kim, Y; Agrón, E; Buggage, R R

    2004-01-01

    Background/aims: Pregnancy and the postpartum period are associated with the activity of autoimmune diseases including uveitis. Although the exact mechanism is unknown, hormones are reported to alter inflammatory cytokines and influence disease activity. The authors studied ocular inflammation, female hormones, and serum cytokine levels during and after pregnancy. Methods: A prospective, observational case study was conducted. Four pregnant women in their first trimester with chronic non-infectious uveitis were followed monthly until 6 months after delivery. Serum female hormones (oestrogen, progesterone, prolactin) and various cytokines (IL-2, IL-4, IL-5, IL-6, IL-10, IFN-γ, and TGF-β) were measured by ELISA. Results: The four patients had five full term pregnancies. Uveitis activity decreased after the first trimester but flared in the early postpartum period. Serum female hormones, highly elevated during pregnancy, drastically dropped post partum. Cytokine levels except TGF-β were mostly undetectable. Conclusion: Female hormones and TGF-β may contribute to the activity of uveitis during pregnancy and the postpartum period. PMID:15548800

  5. Selective phosphorylation of the Dlg1AB variant is critical for TCR-induced p38 activation and induction of proinflammatory cytokines in CD8+ T cells.

    PubMed

    Crocetti, Jillian; Silva, Oscar; Humphries, Lisa A; Tibbs, Michelle D; Miceli, M Carrie

    2014-09-15

    CD8(+) T cells respond to TCR stimulation by producing proinflammatory cytokines, and destroying infected or malignant cells through the production and release of cytotoxic granules. Scaffold protein Discs large homolog 1 (Dlg1) specifies TCR-dependent functions by channeling proximal signals toward the activation of p38-dependent proinflammatory cytokine gene expression and/or p38-independent cytotoxic granule release. Two Dlg1 variants are expressed in CD8(+) T cells via alternative splicing, Dlg1AB and Dlg1B, which have differing abilities coordinate TCR-dependent functions. Although both variants facilitate p38-independent cytotoxicity, only Dlg1AB coordinates p38-dependent proinflammatory cytokine expression. In this study, we identify TCR-induced Dlg1 tyrosine phosphorylation as a key regulatory step required for Dlg1AB-mediated p38-dependent functions, including proinflammatory cytokine expression. We find that Dlg1AB but not Dlg1B is tyrosine phosphorylated by proximal tyrosine kinase Lck in response to TCR stimulation. Furthermore, we identify Dlg1 tyrosine 222 (Y222) as a major site of Dlg1 phosphorylation required for TCR-triggered p38 activation and NFAT-dependent expression of proinflammatory cytokines, but not for p38-independent cytotoxicity. Taken together, our data support a model where TCR-induced phosphorylation of Dlg1 Y222 is a key point of control that endows Dlg1AB with the ability to coordinate p38 activation and proinflammatory cytokine production. We propose blocking Dlg1AB phosphorylation as a novel therapeutic target to specifically block proinflammatory cytokine production but not cytotoxicity.

  6. Violacein Treatment Modulates Acute and Chronic Inflammation through the Suppression of Cytokine Production and Induction of Regulatory T Cells.

    PubMed

    Verinaud, Liana; Lopes, Stefanie Costa Pinto; Prado, Isabel Cristina Naranjo; Zanucoli, Fábio; Alves da Costa, Thiago; Di Gangi, Rosária; Issayama, Luidy Kazuo; Carvalho, Ana Carolina; Bonfanti, Amanda Pires; Niederauer, Guilherme Francio; Duran, Nelson; Costa, Fábio Trindade Maranhão; Oliveira, Alexandre Leite Rodrigues; Höfling, Maria Alice da Cruz; Machado, Dagmar Ruth Stach; Thomé, Rodolfo

    2015-01-01

    Inflammation is a necessary process to control infection. However, exacerbated inflammation, acute or chronic, promotes deleterious effects in the organism. Violacein (viola), a quorum sensing metabolite from the Gram-negative bacterium Chromobacterium violaceum, has been shown to protect mice from malaria and to have beneficial effects on tumors. However, it is not known whether this drug possesses anti-inflammatory activity. In this study, we investigated whether viola administration is able to reduce acute and chronic autoimmune inflammation. For that purpose, C57BL/6 mice were intraperitoneally injected with 1 μg of LPS and were treated with viola (3.5mg/kg) via i.p. at the same time-point. Three hours later, the levels of inflammatory cytokines in the sera and phenotypical characterization of leukocytes were determined. Mice treated with viola presented a significant reduction in the production of inflammatory cytokines compared with untreated mice. Interestingly, although viola is a compound derived from bacteria, it did not induce inflammation upon administration to naïve mice. To test whether viola would protect mice from an autoimmune inflammation, Experimental Autoimmune Encephalomyelitis (EAE)-inflicted mice were given viola i.p. at disease onset, at the 10th day from immunization. Viola-treated mice developed mild EAE disease in contrast with placebo-treated mice. The frequencies of dendritic cells and macrophages were unaltered in EAE mice treated with viola. However, the sole administration of viola augmented the levels of splenic regulatory T cells (CD4+Foxp3+). We also found that adoptive transfer of viola-elicited regulatory T cells significantly reduced EAE. Our study shows, for the first time, that violacein is able to modulate acute and chronic inflammation. Amelioration relied in suppression of cytokine production (in acute inflammation) and stimulation of regulatory T cells (in chronic inflammation). New studies must be conducted in order to

  7. Violacein Treatment Modulates Acute and Chronic Inflammation through the Suppression of Cytokine Production and Induction of Regulatory T Cells

    PubMed Central

    Verinaud, Liana; Lopes, Stefanie Costa Pinto; Prado, Isabel Cristina Naranjo; Zanucoli, Fábio; Alves da Costa, Thiago; Di Gangi, Rosária; Issayama, Luidy Kazuo; Carvalho, Ana Carolina; Bonfanti, Amanda Pires; Niederauer, Guilherme Francio; Duran, Nelson; Costa, Fábio Trindade Maranhão; Oliveira, Alexandre Leite Rodrigues; Höfling, Maria Alice da Cruz; Machado, Dagmar Ruth Stach; Thomé, Rodolfo

    2015-01-01

    Inflammation is a necessary process to control infection. However, exacerbated inflammation, acute or chronic, promotes deleterious effects in the organism. Violacein (viola), a quorum sensing metabolite from the Gram-negative bacterium Chromobacterium violaceum, has been shown to protect mice from malaria and to have beneficial effects on tumors. However, it is not known whether this drug possesses anti-inflammatory activity. In this study, we investigated whether viola administration is able to reduce acute and chronic autoimmune inflammation. For that purpose, C57BL/6 mice were intraperitoneally injected with 1 μg of LPS and were treated with viola (3.5mg/kg) via i.p. at the same time-point. Three hours later, the levels of inflammatory cytokines in the sera and phenotypical characterization of leukocytes were determined. Mice treated with viola presented a significant reduction in the production of inflammatory cytokines compared with untreated mice. Interestingly, although viola is a compound derived from bacteria, it did not induce inflammation upon administration to naïve mice. To test whether viola would protect mice from an autoimmune inflammation, Experimental Autoimmune Encephalomyelitis (EAE)-inflicted mice were given viola i.p. at disease onset, at the 10th day from immunization. Viola-treated mice developed mild EAE disease in contrast with placebo-treated mice. The frequencies of dendritic cells and macrophages were unaltered in EAE mice treated with viola. However, the sole administration of viola augmented the levels of splenic regulatory T cells (CD4+Foxp3+). We also found that adoptive transfer of viola-elicited regulatory T cells significantly reduced EAE. Our study shows, for the first time, that violacein is able to modulate acute and chronic inflammation. Amelioration relied in suppression of cytokine production (in acute inflammation) and stimulation of regulatory T cells (in chronic inflammation). New studies must be conducted in order to

  8. Candida albicans stimulates cytokine production and leukocyte adhesion molecule expression by endothelial cells.

    PubMed Central

    Filler, S G; Pfunder, A S; Spellberg, B J; Spellberg, J P; Edwards, J E

    1996-01-01

    Endothelial cells have the potential to influence significantly the host immune response to blood-borne microbial pathogens, such as Candida albicans. We investigated the ability (of this organism to stimulate endothelial cell responses relevant to host defense in vitro. Infection with C. albicans induced endothelial cells to express mRNAs encoding E-selectin, intercellular adhesion molecule 1, vascular cell adhesion molecule 1, interleukin 6, interleukin 8, monocyte chemoattractant protein 1, and inducible cyclooxygenase (cox2). All three leukocyte adhesion molecule proteins were expressed on the surfaces of the endothelial cells after 8 h of exposure to C. albicans. An increase in secretion of all three cytokines was found after 12 h of infection. Cytochalasin D inhibited accumulation of the endothelial cell cytokine and leukocyte adhesion molecule mRNAs in response to C. albicans, suggesting that endothelial cell phagocytosis of the organism is required to induce this response. Live Candida tropicalis, Candida glabrata, a nongerminating strain of C. albicans, and killed C. albicans did not stimulate the expression of any of the cytokine or leukocyte adhesion molecule mRNAs. These findings indicate that a factor associated with live, germinating C. albicans is required for induction of endothelial cell mRNA expression. Furthermore, since endothelial cells phagocytize killed C. albicans, phagocytosis is likely necessary but not sufficient for this organism to stimulate mRNA accumulation. In conclusion, the secretion of proinflammatory cytokines and expression of leukocyte adhesion molecules by endothelial cells in response to C. albicans could enhance the host defense against this organism by contributing to the recruitment of activated leukocytes to sites of intravascular infection. PMID:8698486

  9. Cytokines and autoimmunity.

    PubMed Central

    Cavallo, M G; Pozzilli, P; Thorpe, R

    1994-01-01

    Although the immunopathology of most autoimmune diseases has been well defined, the mechanisms responsible for the breakdown of self-tolerance and which lead to the development of systemic and organ-specific autoaggression are still unclear. Evidence has accumulated which supports a role for a disregulated production of cytokines by leucocytes and possibly other cells in the pathogenesis of some autoimmune diseases. However, due to the complexity and heterogeneity of cytokine effects in the regulation of the immune response, it is difficult to determine whether abnormalities in the patterns of cytokine production are primary or secondary to the pathological process. Confusion is also caused by the fact that the biological activities of cytokines are multiple and often overlapping, and consequently it is difficult to focus on a unique effect of any one cytokine. Characterization of the potential and actual involvement of cytokines is important not only for a better understanding of the pathogenesis of autoimmune conditions, but particularly because of the implications for the development of immunotherapeutic strategies for the prevention and treatment of the diseases. PMID:8149655

  10. Inflammatory Cytokines in General and Central Obesity and Modulating Effects of Physical Activity

    PubMed Central

    Sander, Christian; Minkwitz, Juliane; Thormann, Julia; Chittka, Tobias; Mergl, Roland; Kirkby, Kenneth C.; Faßhauer, Mathias; Stumvoll, Michael; Holdt, Lesca M.; Teupser, Daniel; Hegerl, Ulrich; Himmerich, Hubertus

    2015-01-01

    Context Chronic systemic inflammation in obesity originates from local immune responses in visceral adipose tissue. However, assessment of a broad range of inflammation-mediating cytokines and their relationship to physical activity and adipometrics has scarcely been reported to date. Objective To characterize the profile of a broad range of pro- and anti-inflammatory cytokines and the impact of physical activity and energy expenditure in individuals with general obesity, central obesity, and non-obese subjects. Design, Setting, and Participants A cross-sectional study comprising 117 obese patients (body mass index (BMI) ≥ 30) and 83 non-obese community-based volunteers. Main Outcomes Measures Serum levels of interleukin (IL)-2, IL-4, IL-5, IL-10, IL-12, IL-13, granulocyte-macrophage colony-stimulating factor (GM-CSF), interferon (IFN)-γ and tumor necrosis factor (TNF)-α were measured. Physical activity and energy expenditure (MET) were assessed with actigraphy. Adipometrics comprised BMI, weight, abdominal-, waist- and hip-circumference, waist to hip ratio (WHR), and waist-to-height-ratio (WHtR). Results General obesity was associated with significantly elevated levels of IL-5, IL-10, IL-12, IL-13, IFN-γ and TNF-α, central obesity with significantly elevated IL-5, IL-10, IL-12, IL-13 and IFN-γ-levels. In participants with general obesity, levels of IL-4, IL-10 and IL-13 were significantly elevated in participants with low physical activity, even when controlled for BMI which was negatively associated with physical acitivity. Cytokines significantly correlated with adipometrics, particularly in obese participants. Conclusions Results confirm up-regulation of certain pro- and anti-inflammatory cytokines in obesity. In obese subjects, physical activity may lower levels and thus reduce pro-inflammatory effects of cytokines that may link obesity, insulin resistance and diabetes. PMID:25781614

  11. Inhibitory effects of devil's claw (secondary root of Harpagophytum procumbens) extract and harpagoside on cytokine production in mouse macrophages.

    PubMed

    Inaba, Kazunori; Murata, Kazuya; Naruto, Shunsuke; Matsuda, Hideaki

    2010-04-01

    Successive oral administration (50 mg/kg) of a 50% ethanolic extract (HP-ext) of devil's claw, the secondary root of Harpagophytum procumbens, showed a significant anti-inflammatory effect in the rat adjuvant-induced chronic arthritis model. HP-ext dose-dependently suppressed the lipopolysaccharide (LPS)-induced production of inflammatory cytokines [interleukin-1beta (IL-1beta), interleukin-6 (IL-6), and tumor necrosis factor-alpha (TNF-alpha)] in mouse macrophage cells (RAW 264.7). Harpagoside, a major iridoid glycoside present in devil's claw, was found to be one of the active agents in HP-ext and inhibited the production of IL-1beta, IL-6, and TNF-alpha by RAW 264.7.

  12. Production of lymphokine-like factors (cytokines) by simian virus 40-infected and simian virus 40-transformed cells.

    PubMed Central

    Bigazzi, P. E.; Yoshida, T.; Ward, P. A.; Cohen, S.

    1975-01-01

    Macrophage migration inhibitory (MIF-like) activity was demonstrated in the supernatant fluids from primary cultures of African green monkey kidney cells infected with simian virus 40 (SV 40) virus. Kidney cell cultures not infected by virus had no MIF activity. Supernatant fluids from continuous cultures of nontransformed and SV 40-transformed human fibroblasts contained MIF-like activity. Productive infection with SV 40 virus results in the production of a lymphokine-like factor, as previously observed in other virus-cell systems, involving mumps virus and Newcast,le disease virus. However, while infection with these paramyxoviruses causes the production of macrophage and neutrophil chemotactic agents as well as an MIF, SV 40 infection does not induce chemotactic factors. The results reported here, taken in conjunction with previous observations by ourselves and others, suggest that the production of lymphokine-like factors (cytokines) may represent a general biologic phenomenon, and that many, if not all, cell types, when appropriately stimulated, may be capable of such activity. PMID:168779

  13. Mycobacterium tuberculosis PPE32 promotes cytokines production and host cell apoptosis through caspase cascade accompanying with enhanced ER stress response

    PubMed Central

    Zeng, Jie; Abdalla, Abualgasim Elgaili; Xie, Jianping

    2016-01-01

    Tuberculosis, caused by Mycobacterium tuberculosis (MTB) infection, remains a grave global public health burden which claims the lives around two to three million annually. PE and PPE proteins, featured by the Pro-Glu (PE) or Pro-Pro-Glu (PPE) motifs at the conserved N-terminal domain, are abundant in the MTB genome. PPE32 can increase intracellular survival of mycobacteria through abnormally increase in cytokines production. PPE32 might subvert the macrophage immune response and thwart its bactericidal effect. THP-1 macrophages treated with PPE32 or infected with Mycobacterium smegmatis (MS) expression PPE32 showed increase of cytokines production and multiple hallmarks of apoptosis. We found that PPE32 significantly increases the expression of IL-12p40 and IL-32 through ERK1/2 signaling pathway. In addition, the cell viability of macrophage was inhibited after PPE32 stimulation. We noted that PPE32 induces cleavage of caspase-3 and caspase-9, while inhibition of caspase activity significantly abrogates the PPE32-induced cell apoptosis. Moreover, PPE32 treatment promotes endoplasmic reticulum stress related gene expression, suggesting ER stress might be responsible for PPE32-induced cell apoptosis. PMID:27634911

  14. Early induction of cytokines/cytokine receptors and Cox2, and activation of NF-κB in 4-nitroquinoline 1-oxide-induced murine oral cancer model.

    PubMed

    Liu, Yu-Ching; Ho, Heng-Chien; Lee, Miau-Rong; Lai, Kuang-Chi; Yeh, Chung-Min; Lin, Yueh-Min; Ho, Tin-Yun; Hsiang, Chien-Yun; Chung, Jing-Gung

    2012-07-15

    The purpose of this study was to identify the genes induced early in murine oral carcinogenesis. Murine tongue tumors induced by the carcinogen, 4-nitroquinoline 1-oxide (4-NQO), and paired non-tumor tissues were subjected to microarray analysis. Hierarchical clustering of upregulated genes in the tumor tissues revealed an association of induced genes with inflammation. Cytokines/cytokine receptors induced early were subsequently identified, clearly indicating their involvement in oral carcinogenesis. Hierarchical clustering also showed that cytokine-mediated inflammation was possibly linked with Mapk6. Cox2 exhibited the greatest extent (9-18 fold) of induction in the microarray data, and its early induction was observed in a 2h painting experiment by RT-PCR. MetaCore analysis showed that overexpressed Cox2 may interact with p53 and transcriptionally inhibit expression of several downstream genes. A painting experiment in transgenic mice also demonstrated that NF-κB activates early independently of Cox2 induction. MetaCore analysis revealed the most striking metabolic alterations in tumor tissues, especially in lipid metabolism resulting from the reduction of Pparα and Rxrg. Reduced expression of Mapk12 was noted, and MetaCore analysis established its relationship with decreased efficiency of Pparα phosphorylation. In conclusion, in addition to cytokines/cytokine receptors, the early induction of Cox2 and NF-κB activation is involved in murine oral carcinogenesis.

  15. Early induction of cytokines/cytokine receptors and Cox2, and activation of NF-κB in 4-nitroquinoline 1-oxide-induced murine oral cancer model

    SciTech Connect

    Liu, Yu-Ching; Ho, Heng-Chien; Lee, Miau-Rong; Lai, Kuang-Chi; Yeh, Chung-Min; Lin, Yueh-Min; Ho, Tin-Yun; Hsiang, Chien-Yun; Chung, Jing-Gung

    2012-07-15

    The purpose of this study was to identify the genes induced early in murine oral carcinogenesis. Murine tongue tumors induced by the carcinogen, 4-nitroquinoline 1-oxide (4-NQO), and paired non-tumor tissues were subjected to microarray analysis. Hierarchical clustering of upregulated genes in the tumor tissues revealed an association of induced genes with inflammation. Cytokines/cytokine receptors induced early were subsequently identified, clearly indicating their involvement in oral carcinogenesis. Hierarchical clustering also showed that cytokine-mediated inflammation was possibly linked with Mapk6. Cox2 exhibited the greatest extent (9–18 fold) of induction in the microarray data, and its early induction was observed in a 2 h painting experiment by RT-PCR. MetaCore analysis showed that overexpressed Cox2 may interact with p53 and transcriptionally inhibit expression of several downstream genes. A painting experiment in transgenic mice also demonstrated that NF-κB activates early independently of Cox2 induction. MetaCore analysis revealed the most striking metabolic alterations in tumor tissues, especially in lipid metabolism resulting from the reduction of Pparα and Rxrg. Reduced expression of Mapk12 was noted, and MetaCore analysis established its relationship with decreased efficiency of Pparα phosphorylation. In conclusion, in addition to cytokines/cytokine receptors, the early induction of Cox2 and NF-κB activation is involved in murine oral carcinogenesis.

  16. Effect of perceived stress on cytokine production in healthy college students.

    PubMed

    Sribanditmongkol, Vorachai; Neal, Jeremy L; Patrick, Thelma E; Szalacha, Laura A; McCarthy, Donna O

    2015-04-01

    Chronic psychological stress impairs antibody synthesis following influenza vaccination. Chronic stress also increases circulating levels of proinflammatory cytokines and glucocorticoids in elders and caregivers, which can impair antibody synthesis. The purpose of this study was to determine whether psychological stress increases ex vivo cytokine production or decreases glucocorticoid sensitivity (GCS) of peripheral blood leukocytes from healthy college students. A convenience sample of Reserve Officer Training Corps (ROTC) students completed the Perceived Stress Scale (PSS). Whole blood was incubated in the presence of influenza vaccine and dexamethasone to evaluate production of interleukin-6 (IL-6), interleukin-1-beta (IL-1β), tumor necrosis factor-alpha (TNF-α), and interferon-gamma (IFN-γ). Multiple regression models controlling for age, gender, and grade point average revealed a negative relationship between PSS and GCS for vaccine-stimulated production of IL-1β, IL-6, and TNF-α. These data increase our understanding of the complex relationship between chronic stress and immune function.

  17. Human Cytomegalovirus MicroRNAs miR-US5-1 and miR-UL112-3p Block Proinflammatory Cytokine Production in Response to NF-κB-Activating Factors through Direct Downregulation of IKKα and IKKβ

    PubMed Central

    Hancock, Meaghan H.; Hook, Lauren M.; Mitchell, Jennifer

    2017-01-01

    ABSTRACT Emerging evidence indicates that human cytomegalovirus (HCMV) manipulates host cell signaling pathways using both proteins and noncoding RNAs. Several studies have shown that HCMV induces NF-κB signaling early in infection, resulting in the induction of antiviral proinflammatory cytokines with a subsequent reduction of these cytokines late in infection. The mechanism for late cytokine reduction is unknown. In this study, we show that HCMV microRNAs (miRNAs) miR-US5-1 and miR-UL112-3p target the IκB kinase (IKK) complex components IKKα and IKKβ to limit production of proinflammatory cytokines in response to interleukin 1β (IL-1β) and tumor necrosis factor alpha (TNF-α). Transfection of miR-UL112-3p and miR-US5-1 mimics reduced endogenous IKKα and IKKβ protein levels, and site-directed mutagenesis of the 3′ untranslated regions (UTRs) identified the binding sites for each miRNA. Infection with mutant viruses lacking these miRNAs resulted in increased levels of IKKα and IKKβ proteins, an impaired ability to control NF-κB signaling at late times of lytic infection, and increased production of proinflammatory cytokines compared to wild-type virus in cell types relevant to HCMV infection in vivo. These phenotypes were rescued by preexpression of miR-US5-1 and miR-UL112-3p in infected cells or by a miR-US5-1/miR-UL112-3p double mutant virus that expresses short hairpin RNAs (shRNAs) targeting IKKα and IKKβ, demonstrating the gene specificity of the miRNAs. These observations describe a mechanism through which HCMV miRNAs expressed late in the infectious cycle downregulate proinflammatory cytokine production to create a cellular proviral environment. PMID:28270578

  18. EGCG attenuates pro-inflammatory cytokines and chemokines production in LPS-stimulated L02 hepatocyte.

    PubMed

    Liu, Qiaoli; Qian, Yun; Chen, Feng; Chen, Xiaoming; Chen, Zhi; Zheng, Min

    2014-01-01

    Endotoxin lipopolysaccharide (LPS) plays an important role in the acceleration of inflammatory reaction of hepatitis as the second attack. Compounds that can prevent inflammation by targeting LPS have potential therapeutic clinical application. Epigallocatechin-3-gallate (EGCG) has potent hepatocyte-protective effect and mild anti-hepatitis virus function. Here, we investigated whether EGCG attenuated the severity of inflammatory response in LPS-stimulated L02 hepatocytes. L02 hepatocytes were pretreated with EGCG for 2 h, then stimulated by LPS at 250 ng/ml. The expression levels of chemokine regulated upon activation normal T-cell expressed and secreted (Rantes) and monocyte chemotactic protein-1 (MCP-1), pro-inflammatory cytokines tumor necrosis factor-α (TNF-α) and interferon-γ, adhesion molecule intercellular adhesion molecule-1 (ICAM-1), oxidant stress molecules nitric oxide (NO), vascular endothelial growth factor (VEGF), and matrix metalloproteinase-2 (MMP-2) were tested by enzyme-linked immunosorbent assay. The expression of total extracellular signal-regulated kinase 1/2 (ERK1/2), phospho-ERK1/2 (p-ERK1/2), p-AKT, total p38, phospho-p38 (p-p38), total p65 and phospho-p65 (p-p65), IκBα, phospho-IκBα (p-IκBα) and TNF receptor associated factor 2 were tested by western blot analysis. Our results showed that pre-treatment with EGCG could significantly reduce the production of TNF-α, Rantes, MCP-1, ICAM-1, NO, VEGF, and MMP-2 in LPS-stimulated L02 hepatocytes in a dose-dependent manner. The effect of EGCG may be related to the inhibition of nuclear factor-κB (NF-κB) and mitogen-activated protein kinase (MAPK) signaling pathways by down-regulation of p-IκBα, p65, p-p65, p-p38, p-ERK1/2, and p-AKT. These results indicate that EGCG suppresses LPS-induced inflammatory response and oxidant stress and exerts its hepatocyte-protective activity partially by inhibiting NF-κB and MAPK pathways.

  19. Modulation in vitro of human natural cytotoxicity, lymphocyte proliferative response to mitogens and cytokine production by essential fatty acids.

    PubMed Central

    Purasiri, P; Mckechnie, A; Heys, S D; Eremin, O

    1997-01-01

    Essential fatty acids (EFA) have been shown in animal studies to have a differential effect on various aspects of immune reactivity. However, there have been few studies in humans. Therefore, we elected to investigate the effects of a variety of EFA [gamma-linolenic acid (GLA), eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA)] in vitro on human blood lymphocyte reactivity, cytokine secretion and natural cytotoxicity. The proliferative response to polyclonal mitogens (phytohaemagglutinin, pokeweed mitogen, concanavalin A), as measured by [3H]thymidine incorporation into newly synthesized lymphocytes, was inhibited (P < 0.05) by all EFAs tested, in a dose-dependent manner (3-15 micrograms/ml). The greatest inhibition of proliferation was caused by EPA and DHA. Similarly, EPA, DHA and GLA significantly reduced cytotoxic activity [expressed as lytic units, using 51 chromium-release assays natural killer (NK) (K562 cells) and lymphokine-activated (LAK) (Daudi cells) cells] (P < 0.05) in a concentration-dependent manner (5-50 micrograms/ml), without affecting cell viability. EPA and DHA exhibited greater suppression than GLA. Furthermore, the inhibition of cell proliferation and suppression of natural cytotoxicity was associated with marked decrease in cytokine [interleukin-1 (IL-1), IL-2, tumour necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma)] production in vitro. Our findings demonstrate that EFAs (GLA, EPA, DHA) have the potential to inhibit significantly various aspects of human lymphocyte cell-mediated and humoral immune reactivities. PMID:9415022

  20. CD73-derived adenosine and tenascin-C control cytokine production by epicardium-derived cells formed after myocardial infarction.

    PubMed

    Hesse, Julia; Leberling, Stella; Boden, Elisabeth; Friebe, Daniela; Schmidt, Timo; Ding, Zhaoping; Dieterich, Peter; Deussen, Andreas; Roderigo, Claudia; Rose, Christine R; Floss, Doreen M; Scheller, Jürgen; Schrader, Jürgen

    2017-03-31

    Epicardium-derived cells (EPDCs) play a fundamental role in embryonic cardiac development and are reactivated in the adult heart in response to myocardial infarction (MI). In this study, EPDCs from post-MI rat hearts highly expressed the ectoenzyme CD73 and secreted the profibrotic matricellular protein tenascin-C (TNC). CD73 on EPDCs extensively generated adenosine from both extracellular ATP and NAD. This in turn stimulated the release of additional nucleotides from a Brefeldin A-sensitive intracellular pool via adenosine-A2BR signaling, forming a positive-feedback loop. A2BR activation in addition strongly promoted the release of major regulatory cytokines, such as IL-6, IL-11, and VEGF. TNC was found to stimulate EPDC migration and, together with ATP-P2X7R signaling, to activate inflammasomes in EPDCs via TLR4. Our results demonstrate that EPDCs are an important source of various proinflammatory factors in the post-MI heart controlled by purinergic and TNC signaling.-Hesse, J., Leberling, S., Boden, E., Friebe, D., Schmidt, T., Ding, Z., Dieterich, P., Deussen, A., Roderigo, C., Rose, C. R., Floss, D. M., Scheller, J., Schrader, J. CD73-derived adenosine and tenascin-C control cytokine production by epicardium-derived cells formed after myocardial infarction.

  1. SATB1 packages densely-looped, transciptionally-active chromatinfor coordinated expression of cytokine genes

    SciTech Connect

    Cai, Shutao; Lee, Charles C.; Kohwi-Shigematsu, Terumi

    2006-05-23

    SATB1 is an important regulator of nuclear architecture that anchors specialized DNA sequences onto its cage-like network and recruits chromatin remodeling/modifying factors to control gene transcription. We studied the role of SATB1 in regulating the coordinated expression of Il5, Il4, and Il13 from the 200kb cytokine gene cluster region of mouse chromosome 11 during T-helper 2 (Th2)-cell activation. We show that upon cell activation, SATB1 is rapidly induced to form a unique transcriptionally-active chromatin structure that includes the cytokine gene region. Chromatin is folded into numerous small loops all anchored by SATB1, is histone H3 acetylated at lysine 9/14, and associated with Th2-specific factors, GATA3, STAT6, c-Maf, the chromatin-remodeling enzyme Brg-1, and RNA polymerase II across the 200kb region. Before activation, the chromatin displays some of these features, such as association with GATA3 and STAT6, but these were insufficient for cytokine gene expression. Using RNA interference (RNAi), we show that upon cell activation, SATB1 is not only required for chromatin folding into dense loops, but also for c-Maf induction and subsequently for Il4, Il5, and Il13 transcription. Our results show that SATB1 is an important determinant for chromatin architecture that constitutes a novel higher-order, transcriptionally-active chromatin structure upon Th2-cell activation.

  2. Periploca forrestii Saponin Ameliorates Murine CFA-Induced Arthritis by Suppressing Cytokine Production

    PubMed Central

    Liu, Yingqin; Li, Minghui; He, Qiuhong; Yang, Xinping; Ruan, Fang

    2016-01-01

    Periploca forrestii Schltr. has been used as a Chinese folk medicine due to its versatile pharmacological effects such as promoting wounds and rheumatoid arthritis. However, the antiarthritic activity of Periploca forrestii saponin (PFS) and its active compound Periplocin has still not been demonstrated. Here, we evaluated the antiarthritic effects of PFS in adjuvant-induced arthritis (AIA) rats by intragastric administration at a dose of 50 mg/kg. The anti-inflammatory activities of Periplocin were also examined in LPS-induced AIA splenocytes and synoviocytes. PFS significantly ameliorated joint swelling; inhibited bone erosion in joints; lowered levels of IL-6 and TGF-β1 in AIA rat splenocyte; and reduced joint protein expression levels of phospho-STAT3 and IKKα. Using LPS-induced AIA splenocytes, we demonstrate that Periplocin suppressed the key proinflammatory cytokines levels of IL-6, IFN-γ, TGF-β1, and IL-13 and IL-22 and transcription factor levels of T-bet, GATA3, and C-Jun genes. Periplocin also suppressed LPS-induced cytokine secretion from synoviocytes. Our study highlights the antiarthritic activity of PFS and its derived Periplocin and the underlying mechanisms. These results provide a strong rationale for further testing and validation of the use of Periploca forrestii Schltr. as an alternative modality for the treatment of RA. PMID:28057980

  3. The Epidermal Growth Factor Receptor Increases Cytokine Production and Cutaneous Inflammation in Response to Ultraviolet Irradiation

    PubMed Central

    El-Abaseri, Taghrid Bahig; Repertinger, Susan K.; Hansen, Laura A.

    2013-01-01

    The epidermal growth factor receptor (EGFR) is activated in cutaneous keratinocytes upon ultraviolet (UV) exposure and has been implicated in ultraviolet-(UV-)induced inflammation and skin tumorigenesis. Egfr mutant mice and EGFR inhibitors were used to investigate the hypothesis that EGFR activation augments inflammation following UV irradiation. Topical treatment of mouse skin with the EGFR inhibitor AG1478 before UV exposure suppressed UV-induced erythema, edema, mast cell infiltration, and neutrophil infiltration. Genetic ablation of Egfr and EGFR inhibition by AG1478 also suppressed the increase in the proinflammatory cytokines tumor necrosis factor α (TNF-α), interleukin-1α, KC (murine IL-8), and cyclooxygenase-2 (COX-2) after UV exposure of cultured keratinocytes. Finally, genetic ablation of inhibition of EGFR in cultured keratinocytes decreased p38 activation after UV, while inhibition of p38 kinase reduced COX-2 expression after UV. These data demonstrate that EGFR regulates multiple aspects of UV-induced inflammation and suggest activation of p38 kinase leading to increased COX-2 and cytokine expression as one mechanism through which it acts. PMID:23878744

  4. Inflammatory cytokines presented from polymer matrices differentially generate and activate DCs in situ

    PubMed Central

    Ali, Omar A.; Tayalia, Prakriti; Shvartsman, Dmitry; Lewin, Sarah; Mooney, David J.

    2014-01-01

    During infection, inflammatory cytokines mobilize and activate dendritic cells (DCs), which are essential for efficacious T cell priming and immune responses that clear the infection. Here we designed macroporous poly(lactide-co-glycolide) (PLG) matrices to release the inflammatory cytokines GM-CSF, Flt3L and CCL20, in order to mimic infection-induced DC recruitment. We then tested the ability of these infection mimics to function as cancer vaccines via induction of specific, anti-tumor T cell responses. All vaccine systems tested were able to confer specific anti-tumor T cell responses and longterm survival in a therapeutic, B16-F10 melanoma model. However, GM-CSF and Flt3L vaccines resulted in similar survival rates, and outperformed CCL20 loaded scaffolds, even though they had differential effects on DC recruitment and generation. GM-CSF signaling was identified as the most potent chemotactic factor for conventional DCs and significantly enhanced surface expression of MHC(II) and CD86(+), which are utilized for priming T cell immunity. In contrast, Flt3L vaccines led to greater numbers of plasmacytoid DCs (pDCs), correlating with increased levels of T cell priming cytokines that amplify T cell responses. These results demonstrate that 3D polymer matrices modified to present inflammatory cytokines may be utilized to effectively mobilize and activate different DC subsets in vivo for immunotherapy. PMID:24688455

  5. PPARγ activation following apoptotic cell instillation promotes resolution of lung inflammation and fibrosis via regulation of efferocytosis and proresolving cytokines

    PubMed Central

    Yoon, Y-S; Kim, S-Y; Kim, M-J; Lim, J-H; Cho, M-S; Kang, J L

    2015-01-01

    Changes in macrophage phenotype have been implicated in apoptotic cell-mediated immune modulation via induction of peroxisome proliferator-activated receptor-γ (PPARγ). In this study, we characterized PPARγ induction by apoptotic cell instillation over the course of bleomycin-induced lung injury in C57BL/6 mice. Next, the role of PPARγ activation in resolving lung inflammation and fibrosis was investigated. Our data demonstrate that apoptotic cell instillation after bleomycin results in immediate and prolonged enhancement of PPARγ mRNA and protein in alveolar macrophages and lung. Moreover, PPARγ activity and expression of its target molecules, including CD36, macrophage mannose receptor, and arginase 1, were persistently enhanced following apoptotic cell instillation. Coadministration of the PPARγ antagonist, GW9662, reversed the enhanced efferocytosis, and the reduced proinflammatory cytokine expression, neutrophil recruitment, myeloperoxidase activity, hydroxyproline contents, and fibrosis markers, including type 1 collagen α2, fibronectin and α-smooth muscle actin (α-SMA), in the lung by apoptotic cell instillation. In addition, inhibition of PPARγ activity reversed the expression of transforming growth factor-β (TGF-β), interleukin (IL)-10, and hepatocyte growth factor (HGF). These findings indicate that one-time apoptotic cell instillation contributes to anti-inflammatory and antifibrotic responses via upregulation of PPARγ expression and subsequent activation, leading to regulation of efferocytosis and production of proresolving cytokines. PMID:25586556

  6. Pancreatic stellate cells are activated by proinflammatory cytokines: implications for pancreatic fibrogenesis

    PubMed Central

    Apte, M; Haber, P; Darby, S; Rodgers, S; McCaughan, G; Korsten, M; Pirola, R; Wilson, J

    1999-01-01

    BACKGROUND—The pathogenesis of pancreatic fibrosis is unknown. In the liver, stellate cells play a major role in fibrogenesis by synthesising increased amounts of collagen and other extracellular matrix (ECM) proteins when activated by profibrogenic mediators such as cytokines and oxidant stress. 
AIMS—To determine whether cultured rat pancreatic stellate cells produce collagen and other ECM proteins, and exhibit signs of activation when exposed to the cytokines platelet derived growth factor (PDGF) or transforming growth factor β (TGF-β). 
METHODS—Cultured pancreatic stellate cells were immunostained for the ECM proteins procollagen III, collagen I, laminin, and fibronectin using specific polyclonal antibodies. For cytokine studies, triplicate wells of cells were incubated with increasing concentrations of PDGF or TGF-β. 
RESULTS—Cultured pancreatic stellate cells stained strongly positive for all ECM proteins tested. Incubation of cells with 1, 5, and 10 ng/ml PDGF led to a significant dose related increase in cell counts as well as in the incorporation of 3H-thymidine into DNA. Stellate cells exposed to 0.25, 0.5, and 1 ng/ml TGF-β showed a dose dependent increase in α smooth muscle actin expression and increased collagen synthesis. In addition, TGF-β increased the expression of PDGF receptors on stellate cells. 
CONCLUSIONS—Pancreatic stellate cells produce collagen and other extracellular matrix proteins, and respond to the cytokines PDGF and TGF-β by increased proliferation and increased collagen synthesis. These results suggest an important role for stellate cells in pancreatic fibrogenesis. 

 Keywords: pancreatic fibrosis; stellate cell activation; cytokines PMID:10075961

  7. Submerged cultivation of Ganoderma lucidum and the effects of its polysaccharides on the production of human cytokines TNF-α, IL-12, IFN-γ, IL-2, IL-4, IL-10 and IL-17.

    PubMed

    Habijanic, Jožica; Berovic, Marin; Boh, Bojana; Plankl, Mojca; Wraber, Branka

    2015-01-25

    An original strain of Ganoderma lucidum (W.Curt.:Fr.) Lloyd, MZKI G97 isolated from Slovenian habitats was grown by a submerged liquid substrate cultivation in a laboratory stirred tank reactor. Five fractions of extracellular and cell-wall polysaccharides were obtained by extraction, ethanol precipitation, and purification by ion-exchange, gel and affinity chromatography. The capacity of isolated polysaccharide fractions to induce innate inflammatory cytokines, and to modulate cytokine responses of activated lymphocytes was investigated. Human peripheral blood mononuclear cells (PBMC) were activated in vitro with polysaccharide fractions, in order to induce innate inflammatory cytokines: tumor necrosis factor alpha (TNF-α), interleukin (IL) 12 and interferon gamma (IFN-γ). For the immunomodulation capacity, polysaccharide fractions were cultured with ionomycine and phorbol myristate acetate (IONO+PMA) activated PBMC, and the concentrations of induced IL-2, IL-4, IFN-γ, IL-10 and IL-17 were measured. The results showed that polysaccharides from G. lucidum induced moderate to high amounts of innate inflammatory cytokines. Fungal cell-wall polysaccharides were stronger innate inflammatory cytokines inducers, while extracellular polysaccharides demonstrated a higher capacity to modulate cytokine responses of IONO+PMA induced production of IL-17. The results indicate that G. lucidum polysaccharides enhance Th1 response with high levels of IFN-γ and IL-2, and display low to no impact on IL-4 production. A similar pattern was observed at regulatory cytokine IL-10. All of the polysaccharide fractions tested induced IL-17 production at different concentration levels.

  8. Treatment With Lenalidomide Modulates T-Cell Immunophenotype and Cytokine Production in Patients With Chronic Lymphocytic Leukemia

    PubMed Central

    Lee, Bang-Ning; Gao, Hui; Cohen, Evan N.; Badoux, Xavier; Wierda, William G.; Estrov, Zeev; Faderl, Stefan H.; Keating, Michael J.; Ferrajoli, Alessandra; Reuben, James M.

    2015-01-01

    BACKGROUND Lenalidomide, an immunomodulatory agent, has activity in lymphoproliferative disorders. The authors, therefore, evaluated its effects on T-cell immunophenotype and cytokine production in patients with chronic lymphocytic leukemia (CLL). METHODS To study the immunomodulatory effects of lenalidomide in CLL, the authors recruited 24 patients with untreated CLL enrolled in a phase 2 clinical trial of lenalidomide and obtained peripheral blood specimens for immunologic studies consisting of enumeration of T cells and assessing their ability to synthesize cytokines after activation through T-cell receptor (TCR). RESULTS After 3 cycles of therapy, patients had a significant reduction in percentage (%) and absolute lymphocyte count (ALC) and an increase in percentage of T cells, percentage of activated CD8+ T cells producing IFN-γ, and percentage of regulatory T (TR) cells when compared with their respective levels before treatment. After 15 cycles of treatment, responder patients had significant reduction in percentage of lymphocytes and ALC, percentage of activated CD4+ T cells producing IL-2, IFN-γ, or TNF-α, and percentage of TR cells when compared with their perspective levels after 3 cycles of treatment. Furthermore, the numbers of activated CD4+ T cells producing IL-2, IFN-γ, or TNF-α, activated CD8+ T cells producing IFN-γ, and TR cells normalized to the range of healthy subjects. CONCLUSIONS Treatment with lenalidomide resulted in the normalization of functional T-cell subsets in responders, suggesting that lenalidomide may modulate cell-mediated immunity in patients with CLL. PMID:21858802

  9. Molecular interactions between T cells and fibroblast-like synoviocytes: role of membrane tumor necrosis factor-alpha on cytokine-activated T cells.

    PubMed

    Tran, Chinh N; Lundy, Steven K; White, Peter T; Endres, Judith L; Motyl, Christopher D; Gupta, Raj; Wilke, Cailin M; Shelden, Eric A; Chung, Kevin C; Urquhart, Andrew G; Fox, David A

    2007-11-01

    The mechanism of fibroblast-like synoviocyte (FLS) transformation into an inflammatory phenotype in rheumatoid arthritis (RA) is not fully understood. FLS interactions with invading leukocytes, particularly T cells, are thought to be a critical component of this pathological process. Resting T cells and T cells activated through the T-cell receptor have previously been shown to induce inflammatory cytokine production by FLS. More recently, a distinct population of T cells has been identified in RA synovium that phenotypically resembles cytokine-activated T (Tck) cells. Using time lapse microscopy, the interactions of resting, superantigen-activated, and cytokine-activated T cells with FLS were visualized. Rapid and robust adhesion of Tck and superantigen-activated T cells to FLS was observed that resulted in flattening of the T cells and a crawling movement on the FLS surface. Tck also readily activated FLS to produce interleukin IL-6 and IL-8 in a cell contact-dependent manner that was enhanced by exogenous IL-17. Although LFA-1 and ICAM-1 co-localized at the Tck-FLS synapse, blocking the LFA-1/ICAM-1 interaction did not substantially inhibit Tck effector function. However, antibody blocking of membrane tumor necrosis factor (TNF)-alpha on the Tck surface did inhibit FLS cytokine production, thus illustrating a novel mechanism for involvement of TNF-alpha in cell-cell interactions in RA synovium and for the effectiveness of TNF-alpha blockade in the treatment of RA.

  10. Immunomodulatory effects of inosine pranobex on cytokine production by human lymphocytes.

    PubMed

    Lasek, Witold; Janyst, Michał; Wolny, Rafał; Zapała, Łukasz; Bocian, Katarzyna; Drela, Nadzieja

    2015-06-01

    Inosine pranobex (inosine dimepranol acedoben, isoprinosine) (Inos) is an immunomodulatory and antiviral drug used in some viral infections, especially in patients with weakened immunity. In the present study, effects of Inos on the production of cytokines attributable to Th1 (IL-2, IFN-g, and TNF-a) or Th2 cells (IL-4, IL-5, and IL-10) were tested in human peripheral blood lymphocyte cultures stimulated with phytohemagglutinin (PHA). Inos enhanced TNF-a secretion significantly (in short-term--24-hour, and prolonged term--72-hour cultures) and IFN-g (in 72-hour cultures). Surprisingly, production of IL-10 by PHA-stimulated lymphocytes was suppressed by Inos in a dose-dependent manner in both 24-hour and 72-hour cultures. These results shed some light on immunomodulatory properties of Inos and suggest applicability of this agent in patients with a depressed function of the immune system.

  11. Alteration of serum inflammatory cytokines in active pulmonary tuberculosis following anti-tuberculosis drug therapy.

    PubMed

    Chowdhury, Imran Hussain; Ahmed, Albin Mostaque; Choudhuri, Subhadip; Sen, Aditi; Hazra, Avijit; Pal, Nishith Kumar; Bhattacharya, Basudev; Bahar, Bojlul

    2014-11-01

    Active pulmonary tuberculosis (APTB) is associated with a failure of the host immune system to control the invading Mycobacterium tuberculosis (Mtb). The objective of this study was to quantify and assess the role of serum inflammatory cytokines in active pulmonary tuberculosis patients following anti-tuberculosis drug (ATD) therapy. Blood samples were collected from APTB patients and normal healthy subjects (NHS) (total n=204) at baseline and 2, 4 and 6 months post-therapy and the abundance of serum inflammatory cytokines were measured by cytokine specific ELISA. Compared to NHS, APTB patients at baseline had higher levels of serum pro-inflammatory cytokines IL-12p40 (P<0.001), IFN-γ (P<0.001), TNF-α (P<0.01), IL-1β (P<0.001) and IL-6 (P<0.001) and anti-inflammatory cytokines IL-10 (P<0.001) and TGF-β1 (P<0.001) while there was no change in the level of IL-4. In APTB patients, the serum levels of IFN-γ, TNF-α, IL-6 and TGF-β1 directly relate to the bacterial load while the TNF-α, IL-1β, IL-6 and TGF-β1 relate to radiological severity. At baseline, the IL-6 level in NHS and APTB patients differed most and following ATD therapy, this level rapidly decreased and stabilized by 4-month in APTB patients. It is concluded that a subtle reduction in the serum level of IL-6 of the APTB patients following ATD therapy might play a vital role in immune-protection of the host against Mtb infection and hence the serum IL-6 level can be a useful marker to diagnose the effectiveness of therapy in the patients.

  12. Diet-induced obesity attenuates cytokine production following an immune challenge.

    PubMed

    Baumgarner, Katherine M; Setti, Sharay; Diaz, Carolyn; Littlefield, Alyssa; Jones, Amanda; Kohman, Rachel A

    2014-07-01

    Obesity increases susceptibility for numerous diseases and neurological disorders including cardiovascular disease, metabolic syndrome, and dementia. One factor that may contribute to the increased risk for these conditions is the development of chronic inflammation. The current study evaluated whether diet-induced obesity (DIO) affects cognitive performance by increasing neuroinflammation and prolonging the behavioral and inflammatory response to an immune challenge. Adult male C57BL/6J mice were fed a high-fat (60% fat) or control diet (10% fat) for 2 or 5 months. After consuming their respective diets for two months, sickness associated behaviors were assessed 4 and 24h after a lipopolysaccharide (LPS) or saline injection. In a separate experiment, DIO and control mice were tested for spatial learning in the water maze and challenged with LPS one month later. Peripheral cytokine production was assessed in adipose and spleen samples and the neuroinflammatory response was assessed in hippocampal, cortical, and brain samples. DIO impaired acquisition of a spatial learning task relative to control mice. However, these deficits are unlikely to be related to inflammation as DIO showed no changes in basal cytokine levels within the periphery or brain. Further, in response to LPS DIO mice showed comparable or attenuated levels of the proinflammatory cytokines interleukin-1β and interleukin-6 relative to control mice. DIO also reduced hippocampal expression of brain-derived neurotrophic factor and the pre-synaptic marker synaptophysin. Presently, the data indicate that DIO suppresses aspects of the immune response and that cognitive deficits associated with DIO may be related to reduced neurotrophic support rather than inflammation.

  13. Multiparameter flow cytometric approach for simultaneous evaluation of proliferation and cytokine-secreting activity in T cells responding to allo-stimulation.

    PubMed

    Tanaka, Yuka; Ohdan, Hideki; Onoe, Takashi; Asahara, Toshimasa

    2004-08-01

    We report a method combining mixed lymphocyte reaction (MLR) using a carboxyfluorescein diacetate succinimidyl ester (CFSE)-labeling technique, intracellular cytokine immunofluorescence staining (ICIS), and multiparameter flow cytometry for simultaneous determination of proliferation and cytokine-secreting activity in T cells responding to allo-stimulation. C57BL/6 (B6) mice and Balb/c mice were used in the experiments. CFSE-labeled responder splenocytes were cultured with irradiated stimulator splenocytes, followed by ICIS. In both the Balb/c stimulator-versus-B6 responder (Balb/c-vs.-B6) and the B6-vs.-Balb/c allogeneic combinations, interleukin (IL)-2 secreting cells and interferon (IFN)-gamma secreting cells were identified predominantly in proliferating CD4+ and CD8+ T cell fractions, respectively. The suitability of this method was proven by demonstrating a close relationship between the values of cytokines in culture supernatants (that were determined by Cytometric Bead Array assay) and indexes for cytokine-production (that were obtained by multiplying the percentage of cytokine-producing cells in T cells and mean fluorescence intensity of cytokine-staining determined by the combined MLR and ICIS).

  14. Pharmacokinetics and cytokine production in heroin and morphine-treated mice.

    PubMed

    Pacifici, R; di Carlo, S; Bacosi, A; Pichini, S; Zuccaro, P

    2000-08-01

    The parallelism between serum levels of heroin and morphine (M) metabolites and the production of interleukin-1beta (IL-1beta), interleukin-2 (IL-2), interleukin-10 (IL-10), tumor necrosis factor alpha (TNF-alpha), transforming growth factor-beta1 (TGF-beta1), and interferon-gamma (IFN-gamma) from murine splenocyte cultures following s.c. injection with 20 mg/kg heroin or M in C57/BL mice is described. The pharmacokinetic profiles of M and inactive morphine-3-glucuronide (M3G) in morphine-treated mice nearly overlapped those in heroin-treated mice, with the only difference being the presence of 6-monoacetylmorphine (AM) in profiles of the latter group. Heroin and M significantly increased production of IL-1beta, IL-2, TNF-alpha and IFN-gamma at 3, 20 and 40 min from treatment, peaking at 20 min, though the effect was very brief. At 24 h production was greatly inhibited, and this depressive effect lasted longer than the stimulatory effect. At 48 h only a partial recovery was observed. Heroin and M also had a highly stimulatory effect on the release of anti-inflammatory cytokines such as TGF-beta1 and IL-10, though this effect was observed after 120 min, peaking at 24 h and then somewhat decreasing at 48 h. This study demonstrates that the more rapid and pronounced immune response to heroin treatment was due to the presence of AM. Both heroin and M produced a biphasic effect on cytokine production: the central opioid or non-opioid receptors are involved in exogenous opiod-induced stimulatory effects, whereas peripheral opioid or non-opioid receptors are involved in depressive effects. Deficient or excess expression of these key mediators may predispose the host to aberrant defence mechanisms.

  15. Profile of cytokine production within the periparasitic granuloma in human alveolar echinococcosis.

    PubMed

    Harraga, Saïd; Godot, Véronique; Bresson-Hadni, Solange; Mantion, Georges; Vuitton, Dominique Angèle

    2003-02-01

    Th2 responses, especially IL-10 secretion by circulating mononuclear cells are associated with the progressive form of AE and Th1 responses with resistance. The HLA B8, DR3, DQ2 haplotype is associated with the severity of AE in humans through immune-mediated mechanisms including an elevated production of Interleukin-10 (IL-10). Granulomatous infiltration of mononuclear cells around the parasitic vesicles is a hallmark of this disease; however, cytokine production by granuloma cells has never been studied. Tissue samples were taken in the periparasitic area and in the central area of the periparasitic granulomatous lesions from a patient with a progressive AE at surgery. Six pieces for each zone were incubated in culture medium with antibiotics and IL-2, together with irradiated autologous peripheral blood mononuclear cells as feeder cells. After four days the dead feeder cells were removed by density gradient centrifugation. Lymphocytes were stimulated with Echinococcus multilocularis vesicular fluid antigen (Emf) or PHA to study IL-10, IFN-, and IL-4 production in the supernatant. Emf-stimulated mononuclear cells from the central part of the lesions secreted more IL-10 and less IFN-gamma than cells from the periphery of the granuloma. At the basal level, IL-10 secretion by the locally infiltrating cells was also high and this confirms at the local granuloma level our previous results obtained from cultures of circulating mononuclear cells. The present study confirms that IL-10 secretion is a key feature of the immune response against E. multilocularis in humans. The location of the cells which produce the highest amount of IL-10, those in contact with parasitic structures, suggests that the parasite itself is able to modulate the immune response of the host so that the infiltrating cells cannot participate in the effector phase of the cellular immune response. The nature of the parasitic structures involved and the mechanisms which lead to an imbalanced cytokine

  16. Adsorption properties of an activated carbon for 18 cytokines and HMGB1 from inflammatory model plasma.

    PubMed

    Inoue, Satoru; Kiriyama, Kentaro; Hatanaka, Yoshihiro; Kanoh, Hirofumi

    2015-02-01

    The ability of an activated carbon (AC) to adsorb 18 different cytokines with molecular weights ranging from 8 kDa to 70 kDa and high mobility group box-1 (HMGB1) from inflammatory model plasma at 310 K and the mechanisms of adsorption were examined. Porosity analysis using N2 gas adsorption at 77K showed that the AC had micropores with diameters of 1-2 nm and mesopores with diameters of 5-20 nm. All 18 cytokines and HMGB1 were adsorbed on the AC; however, the shapes of the adsorption isotherms changed depending on the molecular weight. The adsorption isotherms for molecules of 8-10 kDa, 10-20 kDa, 20-30 kDa, and higher molecular weights were classified as H-2, L-3, S-3, and S-1 types, respectively. These results suggested that the adsorption mechanism for the cytokines and HMGB1 in the mesopores and on the surface of the AC differed as a function of the molecular weight. On the basis of these results, it can be concluded that AC should be efficient for cytokine adsorption.

  17. Soluble TNF-like cytokine (TL1A) production by immune complexes stimulated monocytes in rheumatoid arthritis.

    PubMed

    Cassatella, Marco A; Pereira-da-Silva, Gabriela; da Silva, Gabriela Pereira; Tinazzi, Ilaria; Facchetti, Fabio; Scapini, Patrizia; Calzetti, Federica; Tamassia, Nicola; Wei, Ping; Nardelli, Bernardetta; Roschke, Viktor; Vecchi, Annunciata; Mantovani, Alberto; Bambara, Lisa M; Edwards, Steven W; Carletto, Antonio

    2007-06-01

    TNF-like cytokine (TL1A) is a newly identified member of the TNF superfamily of ligands that is important for T cell costimulation and Th1 polarization. However, despite increasing information about its functions, very little is known about expression of TL1A in normal or pathological states. In this study, we report that mononuclear phagocytes appear to be a major source of TL1A in rheumatoid arthritis (RA), as revealed by their strong TL1A expression in either synovial fluids or synovial tissue of rheumatoid factor (RF)-seropositive RA patients, but not RF-/RA patients. Accordingly, in vitro experiments revealed that human monocytes express and release significant amounts of soluble TL1A when stimulated with insoluble immune complexes (IC), polyethylene glycol precipitates from the serum of RF+/RA patients, or with insoluble ICs purified from RA synovial fluids. Monocyte-derived soluble TL1A was biologically active as determined by its capacity to induce apoptosis of the human erythroleukemic cell line TF-1, as well as to cooperate with IL-12 and IL-18 in inducing the production of IFN-gamma by CD4(+) T cells. Because RA is a chronic inflammatory disease with autoimmune etiology, in which ICs, autoantibodies (including RF), and various cytokines contribute to its pathology, our data suggest that TL1A could be involved in its pathogenesis and contribute to the severity of RA disease that is typical of RF+/RA patients.

  18. Treatment of mice with fenbendazole attenuates allergic airways inflammation and Th2 cytokine production in a model of asthma.

    PubMed

    Cai, Yeping; Zhou, Jiansheng; Webb, Dianne C

    2009-01-01

    Mouse models have provided a significant insight into the role of T-helper (Th) 2 cytokines such as IL-5 and IL-13 in regulating eosinophilia and other key features of asthma. However, the validity of these models can be compromised by inadvertent infection of experimental mouse colonies with pathogens such as oxyurid parasites (pinworms). While the benzimidazole derivative, fenbendazole (FBZ), is commonly used to treat such outbreaks, the effects of FBZ on mouse models of Th2 disease are largely unknown. In this investigation, we show that mice fed FBZ-supplemented food during the in utero and post-weaning period developed attenuated lung eosinophilia, antigen-specific IgG1 and Th2 cytokine responses in a model of asthma. Treatment of the mediastinal lymph node cells from allergic mice with FBZ in vitro attenuated cell proliferation, IL-5 and IL-13 production and expression of the early lymphocyte activation marker, CD69 on CD4(+) T cells and CD19(+) B cells. In addition, eosinophilia and Th2 responses remained attenuated after a 4-week withholding period in allergic mice treated preweaning with FBZ. Thus, FBZ modulates the amplitude of Th2 responses both in vivo and in vitro.

  19. Distinct TCR signaling pathways drive proliferation and cytokine production in T cells.

    PubMed

    Guy, Clifford S; Vignali, Kate M; Temirov, Jamshid; Bettini, Matthew L; Overacre, Abigail E; Smeltzer, Matthew; Zhang, Hui; Huppa, Johannes B; Tsai, Yu-Hwai; Lobry, Camille; Xie, Jianming; Dempsey, Peter J; Crawford, Howard C; Aifantis, Iannis; Davis, Mark M; Vignali, Dario A A

    2013-03-01

    The physiological basis and mechanistic requirements for a large number of functional immunoreceptor tyrosine-based activation motifs (ITAMs; high ITAM multiplicity) in the complex of the T cell antigen receptor (TCR) and the invariant signaling protein CD3 remain obscure. Here we found that whereas a low multiplicity of TCR-CD3 ITAMs was sufficient to engage canonical TCR-induced signaling events that led to cytokine secretion, a high multiplicity of TCR-CD3 ITAMs was required for TCR-driven proliferation. This was dependent on the formation of compact immunological synapses, interaction of the adaptor Vav1 with phosphorylated CD3 ITAMs to mediate the recruitment and activation of the oncogenic transcription factor Notch1 and, ultimately, proliferation induced by the cell-cycle regulator c-Myc. Analogous mechanistic events were also needed to drive proliferation in response to weak peptide agonists. Thus, the TCR-driven pathways that initiate cytokine secretion and proliferation are separable and are coordinated by the multiplicity of phosphorylated ITAMs in TCR-CD3.

  20. Serum Cytokines Th1, Th2, and Th17 Expression Profiling in Active Lupus Nephritis-IV: From a Southern Chinese Han Population

    PubMed Central

    Wang, Yin; Hu, Weiping; Wang, Ning; Sun, Qingyi; Liu, Qingyan; Liu, Xiaocong; Hou, Xianghua; Cheng, Ao

    2016-01-01

    Systemic lupus erythematosus (SLE) is a chronic autoimmune disease characterized by aberrant T cell immune response. Diffuse proliferative lupus nephritis (LN-IV) is the most common, severe, and active form of lupus nephritis. In this study, we investigated the production of Th1, Th2, and Th17 cytokines in prediction of active form of LN-IV. ProcartaPlex multiplex immunoassays panels were used for detection of serum Th1, Th2, and Th17 cytokines profiling. Th1 and Th17 cytokines (IL-18, IFN-γ, IL-12p70, IL-6, and IL-17A) were considerably expressed in the serum of lupus nephritis IV patients in comparison to the healthy control. However, only IL18 and IL6 were higher in class IV versus class III lupus nephritis. Importantly, the ratios of Th1/Th2 (IL-18/IL-4) and Th17/Th2 (IL-17A/IL-4) were significantly elevated in LN-IV when compared with LN-III, LN-V, and healthy controls. Consistently, the serum cytokines IL-18, IL-17A, and IFN-γ were markedly expressed in LN-IV patient glomeruli and interstitial tissue compared to other classes of LN by IHC. ROC further suggests that IL-18 was a potential marker for LN-IV. The data from our study suggests that the early detection and quantification of these cytokines may help in prediction of active form of LN-IV. PMID:27738386

  1. METEORIN-LIKE is a cytokine associated with barrier tissues and alternatively activated macrophages

    PubMed Central

    Ushach, Irina; Burkhardt, Amanda M.; Martinez, Cynthia; Hevezi, Peter A.; Gerber, Peter Arne; Buhren, Bettina Alexandra; Schrumpf, Holger; Valle-Rios, Ricardo; Vazquez, Monica I.; Homey, Bernhard; Zlotnik, Albert

    2014-01-01

    Cytokines are involved in many functions of the immune system including initiating, amplifying and resolving immune responses. Through bioinformatics analyses of a comprehensive database of gene expression (BIGE: Body Index of Gene Expression) we observed that a small secreted protein encoded by a poorly characterized gene called meteorin-like (METRNL), is highly expressed in mucosal tissues, skin and activated macrophages. Further studies indicate that Metrnl is produced by Alternatively Activated Macrophages (AAM) and M-CSF cultured bone marrow macrophages (M2-like macrophages). In the skin, METRNL is expressed by resting fibroblasts and IFNγ-treated keratinocytes. A screen of human skin-associated diseases showed significant over-expression of METRNL in psoriasis, prurigo nodularis, actinic keratosis and atopic dermatitis. METRNL is also up-regulated in synovial membranes of human rheumatoid arthritis. Taken together, these results indicate that Metrnl represents a novel cytokine, which is likely involved in both innate and acquired immune responses. PMID:25486603

  2. A poxvirus-encoded semaphorin induces cytokine production from monocytes and binds to a novel cellular semaphorin receptor, VESPR.

    PubMed

    Comeau, M R; Johnson, R; DuBose, R F; Petersen, M; Gearing, P; VandenBos, T; Park, L; Farrah, T; Buller, R M; Cohen, J I; Strockbine, L D; Rauch, C; Spriggs, M K

    1998-04-01

    The vaccinia virus A39R protein is a member of the semaphorin family. A39R.Fc protein was used to affinity purify an A39R receptor from a human B cell line. Tandem mass spectrometry of receptor peptides yielded partial amino acid sequences that allowed the identification of corresponding cDNA clones. Sequence analysis of this receptor indicated that it is a novel member of the plexin family and identified a semaphorin-like domain within this family, thus suggesting an evolutionary relationship between receptor and ligand. A39R up-regulated ICAM-1 on, and induced cytokine production from, human monocytes. These data, then, describe a receptor for an immunologically active semaphorin and suggest that it may serve as a prototype for other plexin-semaphorin binding pairs.

  3. Epstein-Barr virus (EBV) can immortalize B-cll cells activated by cytokines.

    PubMed

    Wendel-Hansen, V; Sällström, J; De Campos-Lima, P O; Kjellström, G; Sandlund, A; Siegbahn, A; Carlsson, M; Nilsson, K; Rosén, A

    1994-03-01

    B-type of chronic lymphocytic leukemia (B-CLL) cells are inert to the potent transforming action of Epstein-Barr virus (EBV). The mitogenic action of Staphylococcus aureus Cowan I (SAC), MP6-thioredoxin, and interleukin 2 (IL-2), agents previously shown to induce proliferation in normal as well as in B-CLL cells, lifted this block, and EBV-positive cell lines could be established. It was not possible to establish cell lines of leukemic origin from cultures that were incubated with EBV alone or cytokine mix alone. CLL-cells infected with EBV only, expressed the viral nuclear antigen complex (EBNA), but not the viral latent membrane protein (LMP). They were not activated as measured by cell size and 3H-thymidine incorporation. In contrast, cells incubated with EBV and cytokine mix expressed both EBNA and LMP in parallel with enlargement and increased 3H-thymidine incorporation. These results emphasize that LMP expression is a prerequisite for growth transformation and immortalization and that cytokine activation signals are required for its expression in B-CLLs. Cells incubated with SAC/MP6-thioredoxin/IL-2 did not express any of the viral antigens, but were activated with regard to the mentioned parameters. Nine cell lines were established from six patients. From each of the three patients, we obtained 'twin'-pair lines: one corresponding to the malignant cell and the other to a normal B-lymphoblastoid cell. Thus, malignant and normal B-cell counterparts, from the very same donor, are at hand for comparative studies. The cell lines have been carried out for more than 12 months in culture. We conclude that B-CLL that are refractory to EBV-transformation can be rendered susceptible through in vitro cytokine activation.

  4. STAT1-activating cytokines limit Th17 responses through both T-bet-dependent and independent mechanisms1

    PubMed Central

    Villarino, Alejandro V.; Gallo, Eugenio; Abbas, Abul K.

    2010-01-01

    Given the association with autoimmune disease, there is great interest in defining cellular factors that limit overactive or misdirected Th17-type inflammation. Using in vivo and in vitro models, we investigated the molecular mechanisms for cytokine-mediated inhibition of Th17 responses, focusing on the role of STAT1 and T-bet in this process. These studies demonstrate that, during systemic inflammation, STAT1- and T-bet-deficient T cells each exhibit a hyper-Th17 phenotype relative to WT controls. However, IL-17 production was higher in the absence of T-bet and, when both STAT1 and T-bet were deleted, there was no further increase, with the double-deficient cells instead behaving more like STAT1-deficient counterparts. Similar trends were observed during in vitro priming, with production of Th17-type cytokines higher in T-bet−/− T cells than in either STAT1−/− or STAT1−/− T-bet−/− counterparts. The ability of IFN-γ and IL-27 to suppress Th17 responses was reduced in T-bet-deficient cells and, most importantly, ectopic T-bet could suppress signature Th17 gene products, including IL-17A, IL-17F, IL-22 and RORγT, even in STAT1-deficient T cells. Taken together, these studies formally establish that, downstream of IFN-γ, IL-27 and likely all STAT1-activating cytokines, there are both STAT1 and T-bet-dependent pathways capable of suppressing Th17 responses. PMID:20974984

  5. The use of a spaceflight-compatible device to perform WBC surface marker staining and whole-blood mitogenic activation for cytokine detection by flow cytometry

    NASA Technical Reports Server (NTRS)

    Crucian, B. E.; Sams, C. F.

    1999-01-01

    Significant changes have recently been described regarding circulating peripheral immune cells immediately following spaceflight. Existing methods for immunophenotype staining of peripheral blood in terrestrial labs do not meet the constraints for flight on the Space Shuttle. We have recently described the development and use of the Whole Blood Staining Device (WBSD), a simple device for staining flow cytometry specimens during spaceflight. When preparing samples with the WBSD, all liquids are safely contained as the cells are moved through staining, lysis and fixation steps. Here we briefly review the use of the WBSD, and then describe another versatile adaptation, a modification to perform intracellular staining of cytokines for detection by flow cytometry. Alterations in cytokine production have been reported both in ground-based simulated microgravity culture and in astronaut samples returning from spaceflight. Data regarding microgravity effects on cytokine production for specific subpopulations of cells is lacking. Flow cytometric cytokine analysis offers the unique ability to perform simultaneous surface marker analysis and positively identity cytokine producing subsets of cells. The utilization of the WBSD provides the ability to perform rapid and routine mitogenic activation during spaceflight coupled with the ability to perform simultaneous surface marker analysis. The only external requirements for this procedure are an in-flight 37-degree incubator and the capacity for 4-degree storage.

  6. CDCP1 Identifies a CD146 Negative Subset of Marrow Fibroblasts Involved with Cytokine Production

    PubMed Central

    Iwata, Mineo; Torok-Storb, Beverly; Wayner, Elizabeth A.; Carter, William G.

    2014-01-01

    In vitro expanded bone marrow stromal cells contain at least two populations of fibroblasts, a CD146/MCAM positive population, previously reported to be critical for establishing the stem cell niche and a CD146-negative population that expresses CUB domain-containing protein 1 (CDCP1)/CD318. Immunohistochemistry of marrow biopsies shows that clusters of CDCP1+ cells are present in discrete areas distinct from areas of fibroblasts expressing CD146. Using a stromal cell line, HS5, which approximates primary CDCP1+ stromal cells, we show that binding of an activating antibody against CDCP1 results in tyrosine-phosphorylation of CDCP1, paralleled by phosphorylation of Src Family Kinases (SFKs) Protein Kinase C delta (PKC-δ). When CDCP1 expression is knocked-down by siRNA, the expression and secretion of myelopoietic cytokines is increased. These data suggest CDCP1 expression can be used to identify a subset of marrow fibroblasts functionally distinct from CD146+ fibroblasts. Furthermore the CDCP1 protein may contribute to the defining function of these cells by regulating cytokine expression. PMID:25275584

  7. CD80 and CD86 knockdown in dendritic cells regulates Th1/Th2 cytokine production in asthmatic mice

    PubMed Central

    LI, JIAN-GUO; DU, YU-MO; YAN, ZHI-DONG; YAN, JIA; ZHUANSUN, YONG-XUN; CHEN, RUI; ZHANG, WEI; FENG, SU-LING; RAN, PI-XIN

    2016-01-01

    Dendritic cells (DCs) are associated with the activation and differentiation of T helper (Th) cells. Cluster of differentiation (CD)80 and CD86, the co-stimulatory molecules highly expressed in DCs, have are prominent in promoting the differentiation of Th cells toward Th2 cells. However, little is known about the effect of CD80 and CD86 knockdown on Th1/Th2 cytokine production in mature DCs (mDCs). The aim of the present study was to investigate whether small-interfering RNA (siRNA) could suppress the surface expression of CD80 and CD86 in mDCs. The effects of CD80 and CD86 knockdown in mDCs on Th1/Th2 cytokine expression were examined using an asthmatic murine model. DCs were isolated, separated and cultured in vitro. Flow cytometry was used to examine the expression of CD11c, CD80 and CD86 on the DCs. The DCs were transfected with CD80- and CD86-specific siRNA, while non-siRNA and negative siRNA controls were also designed. Then, the mRNA and protein expression levels of CD80 and CD86 were determined by reverse transcription-quantitative polymerase chain reaction and flow cytometry, respectively. The levels of interferon (IFN)-γ and interleukin (IL)-4 produced by T cells co-cultured with mDCs were measured by enzyme-linked immunosorbent assay. Substantial downregulation of CD80 and CD86 mRNA and protein levels were observed in the mDCs following transfection with siRNA. The level of IFN-γ produced by T cells co-cultured with mDCs was significantly increased in the siRNA group, while IL-4 production was significantly decreased. These results show that specific targeting of CD80 and CD86 with siRNA is able to suppress CD80/CD86 expression and consequently regulate Th1/Th2 cytokine levels by increasing IFN-γ production and decreasing IL-4 levels in an asthmatic murine model. PMID:26998006

  8. Methamphetamine administration modifies leukocyte proliferation and cytokine production in murine tissues.

    PubMed

    Peerzada, Habibullah; Gandhi, Jay A; Guimaraes, Allan J; Nosanchuk, Joshua D; Martinez, Luis R

    2013-08-01

    Methamphetamine (METH) is a potent and highly addictive central nervous system (CNS) stimulant. Additionally, METH adversely impacts immunological responses, which might contribute to the higher rate and more rapid progression of certain infections in drug abusers. However no studies have shown the impact of METH on inflammation within specific organs, cellular participation and cytokine production. Using a murine model of METH administration, we demonstrated that METH modifies, with variable degrees, leukocyte recruitment and alters cellular mediators in the lungs, liver, spleen and kidneys of mice. Our findings demonstrate the pleotropic effects of METH on the immune response within diverse tissues. These alterations have profound implications on tissue homeostasis and the capacity of the host to respond to diverse insults, including invading pathogens.

  9. Intracellular cytokine production by dengue virus-specific T cells correlates with subclinical secondary infection.

    PubMed

    Hatch, Steven; Endy, Tim P; Thomas, Stephen; Mathew, Anuja; Potts, James; Pazoles, Pamela; Libraty, Daniel H; Gibbons, Robert; Rothman, Alan L

    2011-05-01

    The pathophysiology of dengue virus infection remains poorly understood, although secondary infection is strongly associated with more severe disease. In the present study, we performed a nested, case-control study comparing the responses of pre-illness peripheral blood mononuclear cells between children who would subsequently develop either subclinical or symptomatic secondary infection 6-11 months after the baseline blood samples were obtained and frozen. We analyzed intracellular cytokine production by CD4(+) and CD8(+) cells in response to stimulation with dengue antigen. We found higher frequencies of dengue virus-specific TNFα, IFNγ-, and IL-2-producing T cells among schoolchildren who subsequently developed subclinical infection, compared with those who developed symptomatic secondary dengue virus infection. Although other studies have correlated immune responses during secondary infection with severity of disease, to our knowledge this is the first study to demonstrate a pre-infection dengue-specific immune response that correlates specifically with a subclinical secondary infection.

  10. Production Systems. Laboratory Activities.

    ERIC Educational Resources Information Center

    Gallaway, Ann, Ed.

    This production systems guide provides teachers with learning activities for secondary students. Introductory materials include an instructional planning outline and worksheet, an outline of essential elements, domains and objectives, a course description, and a content outline. The guide contains 30 modules on the following topics: production…

  11. Cytokines and immune surveillance in humans

    NASA Technical Reports Server (NTRS)

    Sonnenfeld, Gerald

    1993-01-01

    Evidence from both human and rodent studies has indicated that alterations in immunological parameters occur after space flight. Among the parameters shown, by us and others, to be affected is the production of interferons. Interferons are a family of cytokines that are antiviral and play a major role in regulating immune responses that control resistance to infection. Alterations in interferon and other cytokine production and activity could result in changes in immunity and a possible compromise of host defenses against both opportunistic and external infections. The purpose of the present study is to further explore the effects of space flight on cytokines and cytokine-directed immunological function.

  12. Inhibitory effects of geranium essential oil and its major component, citronellol, on degranulation and cytokine production by mast cells.

    PubMed

    Kobayashi, Yuko; Sato, Harumi; Yorita, Mika; Nakayama, Hiroto; Miyazato, Hironari; Sugimoto, Keiichiro; Jippo, Tomoko

    2016-06-01

    We investigated the effects of geranium essential oil (GEO) on anaphylaxis. GEO can exert antioxidant and anti-inflammatory effects, but its roles in allergic reactions are incompletely understood. Here, we used mouse cells to show that GEO inhibited the degranulation of cultured mast cells (CMCs). Citronellol is the major component of GEO and inhibited CMC degranulation. The l-enantiomer of citronellol more effectively suppressed CMC degranulation than did d-citronellol. We also examined whether citronellol could inhibit the immunoglobulin (Ig) E-induced production of tumor necrosis factor (TNF)-α. Treatment with various concentrations of citronellol before CMC activation with IgE significantly inhibited the induction of TNF-α in a dose-dependent manner. Mechanistically, citronellol suppressed the phosphorylation of mitogen-activated protein kinase (ERK), which is critical for ERK activation and the production of inflammatory cytokines in mast cells. These findings suggest that citronellol may represent a candidate compound for the effective treatment of allergic diseases.

  13. Use of Synthetic Derivatives To Determine the Minimal Active Structure of Cytokine-Inducing Lipoteichoic Acid▿

    PubMed Central

    Deininger, Susanne; Figueroa-Perez, Ignacio; Sigel, Stefanie; Stadelmaier, Andreas; Schmidt, Richard R.; Hartung, Thomas; von Aulock, Sonja

    2007-01-01

    Lipoteichoic acid (LTA) from gram-positive bacteria is the counterpart to lipopolysaccharide from gram-negative bacteria. LTA, which activates Toll-like receptor 2 (TLR2), induces a unique cytokine and chemokine pattern. The chemical synthesis of LTA proved its immunostimulatory properties. To determine the minimal active structure of LTA, we reduced synthetic LTA in a number of steps down to the synthetic anchor and employed these molecules to stimulate interleukin-8 (IL-8) release in human whole blood. Ten times more of the synthetic structures with four to six d-alanine-substituted polyglycerophosphate units (50 nM) than of the native LTA preparation was required to induce IL-8 release. A further reduction to three backbone units with two or no d-alanine residues resulted in cytokine induction only from 500 nM. The synthetic anchor was not able to induce IL-8 release even at 5 μM. When the LTA derivatives were used at 500 nM, they induced increasing levels of IL-8 and tumor necrosis factor alpha with increasing elongation of the backbone. Peritoneal macrophages were less responsive than human blood to the synthetic structures. Therefore, TLR2 dependency could be shown only with cells from TLR2-deficient mice for the two largest synthetic structures. This was confirmed by using TLR2-transfected HEK 293 cells. Taken together, these data indicate that although the synthetic anchor (which, unlike the native anchor, contains only myristic acid) cannot induce cytokine release, the addition of three backbone units, even without d-alanine substituents, confers this ability. Lengthening of the chain with d-alanine-substituted backbone units results in increased cytokine-inducing potency and a more sensitive response. PMID:17928431

  14. Moraxella catarrhalis induces mast cell activation and nuclear factor kappa B-dependent cytokine synthesis.

    PubMed

    Krishnaswamy, G; Martin, R; Walker, E; Li, C; Hossler, F; Hall, K; Chi, D S

    2003-01-01

    Human mast cells are often found perivascularly and at mucosal sites and may play crucial roles in the inflammatory response. Recent studies have suggested a prominent role for mast cells in host defense. In this study, we analyzed the effects of a common airway pathogen, Moraxella catarrhalis and a commensal bacterium, Neiserria cinerea, on activation of human mast cells. Human mast cell leukemia cells (HMC-1) were activated with either phorbol myristate acetate (PMA) and calcium ionophore or with varying concentrations of heat-killed suspensions of bacteria. Supernatants were assayed for the cytokines interleukin-4 (IL-4), granulocyte macrophage colony stimulating factor (GM-CSF), IL-6, IL-8, IL-13 and monocyte chemotactic protein-1 (MCP-1). Nuclear proteins were isolated and assayed by electrophoretic mobility shift assay (EMSA) for nuclear factor kappaB (NF-kappaB) nuclear binding activity. In some experiments, NF-kappaB inhibitor, Bay-11 was added to determine functional significance. Both M. catarrhalis and N. cinerea induced mast cell activation and selective secretion of two key inflammatory cytokines, IL-6 and MCP-1. This was accompanied by NF-kappaB activation. Neither spun bacterial supernatants nor bacterial lipopolysaccharide induced cytokine secretion, suggesting need for direct bacterial contact with mast cells. Scanning electron microscopy revealed active aggregation of bacteria over mast cell surfaces. The NF-kappaB inhibitor, Bay-11, inhibited expression of MCP-1. These findings suggest the possibility of direct interactions between human mast cells and common bacteria and provide evidence for a novel role for human mast cells in innate immunity.

  15. Innate production of T(H)2 cytokines by adipose tissue-associated c-Kit(+)Sca-1(+) lymphoid cells.

    PubMed

    Moro, Kazuyo; Yamada, Taketo; Tanabe, Masanobu; Takeuchi, Tsutomu; Ikawa, Tomokatsu; Kawamoto, Hiroshi; Furusawa, Jun-Ichi; Ohtani, Masashi; Fujii, Hideki; Koyasu, Shigeo

    2010-01-28

    Innate immune responses are important in combating various microbes during the early phases of infection. Natural killer (NK) cells are innate lymphocytes that, unlike T and B lymphocytes, do not express antigen receptors but rapidly exhibit cytotoxic activities against virus-infected cells and produce various cytokines. Here we report a new type of innate lymphocyte present in a novel lymphoid structure associated with adipose tissues in the peritoneal cavity. These cells do not express lineage (Lin) markers but do express c-Kit, Sca-1 (also known as Ly6a), IL7R and IL33R. Similar lymphoid clusters were found in both human and mouse mesentery and we term this tissue 'FALC' (fat-associated lymphoid cluster). FALC Lin(-)c-Kit(+)Sca-1(+) cells are distinct from lymphoid progenitors and lymphoid tissue inducer cells. These cells proliferate in response to IL2 and produce large amounts of T(H)2 cytokines such as IL5, IL6 and IL13. IL5 and IL6 regulate B-cell antibody production and self-renewal of B1 cells. Indeed, FALC Lin(-)c-Kit(+)Sca-1(+) cells support the self-renewal of B1 cells and enhance IgA production. IL5 and IL13 mediate allergic inflammation and protection against helminth infection. After helminth infection and in response to IL33, FALC Lin(-)c-Kit(+)Sca-1(+) cells produce large amounts of IL13, which leads to goblet cell hyperplasia-a critical step for helminth expulsion. In mice devoid of FALC Lin(-)c-Kit(+)Sca-1(+) cells, such goblet cell hyperplasia was not induced. Thus, FALC Lin(-)c-Kit(+)Sca-1(+) cells are T(H)2-type innate lymphocytes, and we propose that these cells be called 'natural helper cells'.

  16. Microscale to manufacturing scale-up of cell-free cytokine production--a new approach for shortening protein production development timelines.

    PubMed

    Zawada, James F; Yin, Gang; Steiner, Alexander R; Yang, Junhao; Naresh, Alpana; Roy, Sushmita M; Gold, Daniel S; Heinsohn, Henry G; Murray, Christopher J

    2011-07-01

    Engineering robust protein production and purification of correctly folded biotherapeutic proteins in cell-based systems is often challenging due to the requirements for maintaining complex cellular networks for cell viability and the need to develop associated downstream processes that reproducibly yield biopharmaceutical products with high product quality. Here, we present an alternative Escherichia coli-based open cell-free synthesis (OCFS) system that is optimized for predictable high-yield protein synthesis and folding at any scale with straightforward downstream purification processes. We describe how the linear scalability of OCFS allows rapid process optimization of parameters affecting extract activation, gene sequence optimization, and redox folding conditions for disulfide bond formation at microliter scales. Efficient and predictable high-level protein production can then be achieved using batch processes in standard bioreactors. We show how a fully bioactive protein produced by OCFS from optimized frozen extract can be purified directly using a streamlined purification process that yields a biologically active cytokine, human granulocyte-macrophage colony-stimulating factor, produced at titers of 700 mg/L in 10 h. These results represent a milestone for in vitro protein synthesis, with potential for the cGMP production of disulfide-bonded biotherapeutic proteins.

  17. Differential S1P Receptor Profiles on M1- and M2-Polarized Macrophages Affect Macrophage Cytokine Production and Migration

    PubMed Central

    Müller, Jan; von Bernstorff, Wolfram; Heidecke, Claus-Dieter

    2017-01-01

    Introduction. Macrophages are key players in complex biological processes. In response to environmental signals, macrophages undergo polarization towards a proinflammatory (M1) or anti-inflammatory (M2) phenotype. Sphingosine 1-phosphate (S1P) is a bioactive lysophospholipid that acts via 5 G-protein coupled receptors (S1P1–5) in order to influence a broad spectrum of biological processes. This study assesses S1P receptor expression on macrophages before and after M1 and M2 polarization and performs a comparative analysis of S1P signalling in the two activational states of macrophages. Methods. Bone marrow derived macrophages (BMDM) from C57 BL/6 mice were cultured under either M1- or M2-polarizing conditions. S1P-receptor expression was determined by quantitative RT-PCR. Influence of S1P on macrophage activation, migration, phagocytosis, and cytokine secretion was assessed in vitro. Results. All 5 S1P receptor subclasses were expressed in macrophages. Culture under both M1- and M2-polarizing conditions led to significant downregulation of S1P1. In contrast, M1-polarized macrophages significantly downregulated S1P4. The expression of the remaining three S1P receptors did not change. S1P increased expression of iNOS under M2-polarizing conditions. Furthermore, S1P induced chemotaxis in M1 macrophages and changed cytokine production in M2 macrophages. Phagocytosis was not affected by S1P-signalling. Discussion. The expression of different specific S1P receptor profiles may provide a possibility to selectively influence M1- or M2-polarized macrophages. PMID:28367448

  18. Relationship of cytokines and AGE products in diabetic and non-diabetic patients with cataract

    PubMed Central

    Hamid, Sadaf; Gul, Anjuman; Hamid, Qamar

    2016-01-01

    Objectives Cytokines are important mediators of inflammatory and immune responses. The aim of this study was to investigate the changes in cytokines concentration (IL-6, IL-8 and TNF-α) and serum advanced glycation end products (sAGEs) in senile diabetics with or without cataract and non-diabetic patients with cataract. Methodology The study included 124 subjects (sixty or over sixty years age), distributed as four groups thirty senile diabetic patients with cataract (Group I) (16 female and 14 male), thirty senile non-diabetic patients with cataract (Group II) (15 female and 15 male), thirty three senile diabetic patients without any complication (Group III) (16 female and 17 male), thirty one apparently normal healthy individuals (Group IV) (16 female and 15 male), age, sex and weight matched with senile control subjects were investigated. Patients were selected on clinical grounds from Eye Ward Jinnah Postgraduate Medical Centre. Results Interleukin-6 (IL-6), interleukin-8 (IL-8) and tumor necrosis factor-α (TNF-α) levels were significantly increased (P < 0.001) in Group I and III as compared to Group II and IV. Fasting blood glucose, glycosylated hemoglobin, serum fructosamine, malondialdehyde (MDA), sAGEs, IL-6, IL-8 and TNF-α levels were significantly increased (P < 0.001) in Group I as compared to Group II and the levels were almost same in Group II and IV. There was a significant decrease in serum vitamin E and total antioxidant status (p< 0.001) in Group I and Group III as compared to Group II and Group IV. Conclusion The results of the present study thus demonstrated that levels increased in both condition but are more severe in diabetic patients with cataract that may be a predictor for cataractogenesis and the levels were almost same in Group II and IV. PMID:27833515

  19. Activation of cytokine genes in T cells during primary and secondary murine influenza pneumonia.

    PubMed

    Carding, S R; Allan, W; McMickle, A; Doherty, P C

    1993-02-01

    The patterns of cytokine mRNA expression in mice with primary or secondary influenza pneumonia have been assessed by in situ hybridization analysis of cells from both the mediastinal lymph node (MLN) and the virus-infected lung. Evidence of substantial transcriptional activity was found in all lymphocyte subsets recovered from both anatomical sites. The kinetics of cytokine mRNA expression after primary infection with an H3N2 virus were in accord with the idea that the initial response occurs in regional lymphoid tissue, with the effector T cells later moving to the lung. This temporal separation was much less apparent for the more rapid secondary response resulting from challenge of H3N2-primed mice with an H1N1 virus. Among the T cell receptor alpha/beta+ subsets, transcripts for interferon (IFN) gamma and tumor necrosis factor beta were most commonly found in the CD8+ population whereas mRNA for interleukin (IL) 4 and IL-10 was much more prevalent in CD4+ T cells. The gamma/delta T cells expressed mRNA for all cytokines tested, with IL-2, IL-4, and IFN-gamma predominating among those recovered from the inflammatory exudate. At particular time points, especially early in the MLN and late in the infected lung, the frequency of mRNA+ lymphocytes was much higher than would be expected from current understanding of the prevalence of virus-specific precursors and effectors. If this response is typical, induction of cytokine gene expression for T cells that are not responding directly to the invading pathogen may be a prominent feature of acute virus infections.

  20. Tacrolimus does not alter the production of several cytokines and antimicrobial peptide in Malassezia furfur-infected-keratinocytes.

    PubMed

    Balato, Anna; Paoletti, Iole; De Gregorio, Vincenza; Cantelli, Mariateresa; Ayala, Fabio; Donnarumma, Giovanna

    2014-03-01

    Topical immunosuppressant therapy is widely used in the treatment of inflammatory skin diseases, such as atopic dermatitis and psoriasis. Besides its beneficial therapeutic effects, application of topical anti-inflammatory drugs may render the epidermis more vulnerable to invading pathogens by suppressing innate immune responses in keratinocytes (KCs). Cytokines, chemokines and antimicrobial peptides (AMPs) produced by epithelial cells enable them to participate in innate and acquired immune responses. The aim of the present work was to study the influence of tacrolimus (FK506) on KCs infected with Malassezia furfur (M. furfur), evaluating the expression of pro-inflammatory cytokines IL-1α and IL-6, chemokine IL-8, anti-inflammatory cytokines transforming growth factor beta1 (TGF-β1) and IL-10 and AMP β-defensin-2. Human KCs were obtained from surgical specimens of normal adult skin. The expression of mRNAs in KCs: FK506-treated, FK506-treated and M. furfur-infected as well as only M. furfur-infected was quantified by real-time quantitative polymerase chain reaction. Next, the production of the AMP β-defensin-2 and of the above-mentioned pro-inflammatory and anti-inflammatory cytokines was evaluated using enzyme-linked immunosorbent assay. In this study, FK506 did not alter cytokine and AMP production by KCs; this led us to hypothesise that it may not enhance the risk of mycotic skin infections.

  1. Hedgehog Signaling Non-Canonical Activated by Pro-Inflammatory Cytokines in Pancreatic Ductal Adenocarcinoma

    PubMed Central

    Wang, Yuqiong; Jin, Gang; Li, Quanjiang; Wang, Zhiping; Hu, Weimin; Li, Ping; Li, Shude; Wu, Hongyu; Kong, Xiangyu; Gao, Jun; Li, Zhaoshen

    2016-01-01

    Hedgehog(HH) pathway is found to be activated through a manner of canonical, or the non-canonical HH pathways. Distinct hyperplasia stroma around tumor cells is supposed to express pro-inflammatory cytokines abundantly, such as tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β), etc. in pancreatic ductal adenocarcinoma (PDAC) tissues. In this study we observed the effects of TNF-α and IL-1β on HH pathway activation in PDAC cells, and explored their activation manners. Our results showed that pro-inflammatory cytokines, TNF-α and IL-1β, could up-regulate the expression of GLI1 gene, increase its nuclear protein expression and promote malignant cell behaviors including migration, invasion, epithelial-mesenchymal transition (EMT) and drug resistance as well. Moreover, GLI1 promoter-reporter assay in combination with blocking either NF-κB or Smoothened (SMO) suggested that TNF-α and IL-1β could transcriptionally up-regulate expression of GLI1 completely via NF-κB, whereas ablation of SMO could not completely attenuate the regulation effects of TNF-α and IL-1β on GLI1 expression. Collectively, our results indicated that TNF-α and IL-1β in hyperplasia stroma can promote the PDAC cell development by activating HH pathway, through both the canonical and non-canonical HH activation ways. PMID:27877222

  2. Poly IC triggers a cathepsin D- and IPS-1-dependent pathway to enhance cytokine production and mediate dendritic cell necroptosis.

    PubMed

    Zou, Jian; Kawai, Taro; Tsuchida, Tetsuo; Kozaki, Tatsuya; Tanaka, Hiroki; Shin, Kyung-Sue; Kumar, Himanshu; Akira, Shizuo

    2013-04-18

    RIG-I-like receptors (RLRs) sense virus-derived RNA or polyinosinic-polycytidylic acid (poly IC) to exert antiviral immune responses. Here, we examine the mechanisms underlying the adjuvant effects of poly IC. Poly IC was taken up by dendritic cells (DCs), and it induced lysosomal destabilization, which, in turn, activated an RLR-dependent signaling pathway. Upon poly IC stimulation, cathepsin D was released into the cytoplasm from the lysosome to interact with IPS-1, an adaptor molecule for RLRs. This interaction facilitated cathepsin D cleavage of caspase 8 and the activation of the transcription factor NF-κB, resulting in enhanced cytokine production. Further recruitment of the kinase RIP-1 to this complex initiated the necroptosis of a small number of DCs. HMGB1 released by dying cells enhanced IFN-β production in concert with poly IC. Collectively, these findings suggest that cathepsin D-triggered, IPS-1-dependent necroptosis is a mechanism that propagates the adjuvant efficacy of poly IC.

  3. Sertoli cells have a functional NALP3 inflammasome that can modulate autophagy and cytokine production

    PubMed Central

    Hayrabedyan, Soren; Todorova, Krassimira; Jabeen, Asma; Metodieva, Gergana; Toshkov, Stavri; Metodiev, Metodi V.; Mincheff, Milcho; Fernández, Nelson

    2016-01-01

    Sertoli cells, can function as non-professional tolerogenic antigen-presenting cells, and sustain the blood-testis barrier formed by their tight junctions. The NOD-like receptor family members and the NALP3 inflammasome play a key role in pro-inflammatory innate immunity signalling pathways. Limited data exist on NOD1 and NOD2 expression in human and mouse Sertoli cells. Currently, there is no data on inflammasome expression or function in Sertoli cells. We found that in primary pre-pubertal Sertoli cells and in adult Sertoli line, TLR4\\NOD1 and NOD2 crosstalk converged in NFκB activation and elicited a NALP3 activation, leading to de novo synthesis and inflammasome priming. This led to caspase-1 activation and IL-1β secretion. We demonstrated this process was controlled by mechanisms linked to autophagy. NOD1 promoted pro-IL-1β restriction and autophagosome maturation arrest, while NOD2 promoted caspase-1 activation, IL-1β secretion and autophagy maturation. NALP3 modulated NOD1 and pro-IL-1β expression, while NOD2 inversely promoted IL-1β. This study is proof of concept that Sertoli cells, upon specific stimulation, could participate in male infertility pathogenesis via inflammatory cytokine induction. PMID:26744177

  4. MiR-150 impairs inflammatory cytokine production by targeting ARRB-2 after blocking CD28/B7 costimulatory pathway

    PubMed Central

    Sang, Wei; Wang, Ying; Zhang, Cong; Zhang, Dianzheng; Sun, Cai; Niu, Mingshan; Zhang, Zhe; Wei, Xiangyu; Pan, Bin; Chen, Wei; Yan, Dongmei; Zeng, Lingyu; Loughran, Thomas P.; Xu, Kailin

    2016-01-01

    MiR-150, a major modulator negatively regulating the development and differentiation of various immune cells, is widely involved in orchestrating inflammation. In transplantation immunity, miR-150 can effectively induce immune tolerance, although the underlying mechanisms have not been fully elucidated. In the current study, we found that miR-150 is elevated after blocking CD28/B7 co-stimulatory signaling pathway and impaired IL-2 production by targeting ARRB2. Further investigation suggested that miR-150 not only repressed the level of ARRB2/PDE4 directly but also prevented AKT/ARRB2/PDE4 trimer recruitment into the lipid raft by inhibiting the activities of PI3K and AKT through the cAMP-PKA-Csk signaling pathway. This leads to the interruption of cAMP degradation and subsequently results in inhibition of the NF-kB pathway and reduced production of both IL-2 and TNF. In conclusion, our study demonstrated that miR-150 can effectively prevent CD28/B7 co-stimulatory signaling transduction, decrease production of inflammatory cytokines, such as IL-2 and TNF, and elicit the induction of immune tolerance. Therefore, miR-150 could become a novel potential therapeutic target in transplantation immunology. PMID:26549736

  5. JANEX-1, a JAK3 inhibitor, protects pancreatic islets from cytokine toxicity through downregulation of NF-{kappa}B activation and the JAK/STAT pathway

    SciTech Connect

    Lv, Na; Kim, Eun-Kyung; Song, Mi-Young; Choi, Ha-Na; Moon, Woo Sung; Park, Sung-Joo; Park, Jin-Woo; Kwon, Kang-Beom; Park, Byung-Hyun

    2009-07-15

    JANEX-1/WHI-P131, a selective Janus kinase 3 (JAK3) inhibitor, has been shown to delay the onset of diabetes in the NOD mouse model. However, the molecular mechanism by which JANEX-1 protects pancreatic {beta}-cells is unknown. In the current study, we investigated the role of JANEX-1 on interleukin (IL)-1{beta} and interferon (IFN)-{gamma}-induced {beta}-cell damage using isolated islets. JANEX-1-pretreated islets showed resistance to cytokine toxicity, namely suppressed nitric oxide (NO) production, reduced inducible form of NO synthase (iNOS) expression, and decreased islet destruction. The molecular mechanism by which JANEX-1 inhibits iNOS expression was mediated through suppression of the nuclear factor {kappa}B (NF-{kappa}B) and JAK/signal transducer and activator of transcription (STAT) pathways. Islets treated with the cytokines downregulated the protein levels of suppressor of cytokine signaling (SOCS)-1 and SOCS-3, but pretreatment with JANEX-1 attenuated these decreases. Additionally, islets from JAK3{sup -/-} mice were more resistant to cytokine toxicity than islets from control mice. These results demonstrate that JANEX-1 protects {beta}-cells from cytokine toxicity through suppression of the NF-{kappa}B and JAK/STAT pathways and upregulation of SOCS proteins, suggesting that JANEX-1 may be used to preserve functional {beta}-cell mass.

  6. Pro-inflammatory cytokines enhance ERAD and ATF6α pathway activity in salivary glands of Sjögren's syndrome patients.

    PubMed

    Barrera, María-José; Aguilera, Sergio; Castro, Isabel; Cortés, Juan; Bahamondes, Verónica; Quest, Andrew F G; Molina, Claudio; González, Sergio; Hermoso, Marcela; Urzúa, Ulises; Leyton, Cecilia; González, María-Julieta

    2016-12-01

    Salivary gland (SG) acinar-cells are susceptible to endoplasmic reticulum (ER) stress related to their secretory activity and the complexity of synthesized secretory products. SGs of Sjögren's syndrome patients (SS)-patients show signs of inflammation and altered proteostasis, associated with low IRE1α/XBP-1 pathway activity without avert increases in apoptosis. Acinar-cells may avoid apoptosis by activation of the ATF6α pathway and ER-associated protein degradation (ERAD). The aim of this study was to evaluate the role of pro-inflammatory cytokines in ATF6α pathway/ERAD activation and cell viability in labial salivary glands (LSG) of SS-patients. In biopsies from SS-patients increased ATF6α signaling pathway activity, as evidenced by generation of the ATF6f cleavage fragment, and increased expression of ERAD machinery components, such as EDEM1, p97, SEL1L, gp78, UBE2J1, UBE2G2, HERP and DERLIN1, were observed compared to controls. Alternatively, for pro- (active-caspase-3) and anti-apoptotic (cIAP2) markers no significant difference between the two experimental groups was detected. Increased presence of ATF6f and ERAD molecules correlated significantly with increased expression of pro-inflammatory cytokines. These observations were corroborated in vitro in 3D-acini treated with TNF-α and/or IFN-γ, where an increase in the expression and activation of the ATF6α sensor and ERAD machinery components was detected under ER stress conditions, while changes in cell viability and caspase-3 activation were not observed. Cytokine stimulation protected cells from death when co-incubated with an ERAD machinery inhibitor. Alternatively, when cytokines were eliminated from the medium prior to ERAD inhibition, cell death increased, suggesting that the presence of pro-inflammatory cytokines in the medium is essential to maintain cell viability. In conclusion, the ATF6α pathway and the ERAD machinery are active in LSG of SS-patients. Both were also activated by TNF

  7. Altered production of immunoregulatory cytokines by invariant Valpha19 TCR-bearing cells dependent on the duration and intensity of TCR engagement.

    PubMed

    Shimamura, Michio; Huang, Yi-Ying; Kobayashi, Masumi; Goji, Hiroshi

    2009-02-01

    Cells bearing invariant Valpha19-Jalpha33 TCR alpha chains are believed to participate in the regulation of inflammatory autoimmune diseases. In this study, the potential to produce immunoregulatory cytokines by these cells was characterized in order to find the mechanism underlying their immunoregulatory functions. Serum levels of IL-4, IL-10, transforming growth factor-beta, IFN-gamma and IL-17 increased in mice over-expressing an invariant Valpha19-Jalpha33 TCR alpha transgene (Valpha19 Tg) in response to anti-CD3 antibody injection. NK1.1(+) Valpha19 Tg(+), but not NK1.1(-) Valpha19 Tg(+) cells, promptly produced immunoregulatory IL-4, IFN-gamma and IL-17 upon invariant TCR engagement with immobilized anti-CD3 antibody in culture. The activation of Valpha19 Tg(+) cells then triggered the production of pro-inflammatory cytokines by bystander cells. Interestingly, the ratio of T(h)2 cytokines such as IL-4, IL-5 and IL-10, but not pro-inflammatory IL-17, to IFN-gamma was increased when the intensity of the stimulation to invariant TCR was attenuated. Collectively, these findings suggest that invariant Valpha19 TCR(+) cells have the potential to participate in the regulation of inflammatory autoimmunity by producing T(h)2-biased cytokines in certain circumstances.

  8. Effect of sesamin against cytokine production from influenza type A H1N1-induced peripheral blood mononuclear cells: computational and experimental studies.

    PubMed

    Fanhchaksai, Kanda; Kodchakorn, Kanchanok; Pothacharoen, Peraphan; Kongtawelert, Prachya

    2016-01-01

    In 2009, swine flu (H1N1) had spread significantly to levels that threatened pandemic influenza. There have been many treatments that have arisen for patients since the WHO first reported the disease. Although some progress in controlling influenza has taken place during the last few years, the disease is not yet under control. The development of new and less expensive anti-influenza drugs is still needed. Here, we show that sesamin from the seeds of the Thai medicinal plant Sesamum indicum has anti-inflammatory cytokines in human peripheral blood mononuclear cells (PBMCs) induced by 2009 influenza virus type A H1N1. In this study, the combinatorial screening method combined with the computational approach was applied to investigate the new molecular binding structures of sesamin against the 2009 influenza virus type A H1N1 (p09N1) crystallized structure. Experimental methods were applied to propose the mechanisms of sesamin against cytokine production from H1N1-induced human PBMC model. The molecular dynamics simulation of sesamin binding with the p09N1 crystallized structure showed new molecular binding structures at ARG118, ILE222, ARG224, and TYR406, and it has been proposed that sesamin could potentially be used to produce anti-H1N1 compounds. Furthermore, the mechanisms of sesamin against cytokine production from influenza type A H1N1-induced PBMCs by ELISA and signaling transduction showed that sesamin exhibits the ability to inhibit proinflammatory cytokines, IL-1β and TNF-α, and to enhance the activity of the immune cell cytokine IL-2 via downregulating the phosphorylated JNK, p38, and ERK1/2 MAPK signaling pathways. This information might very well be useful in the prevention and treatment of immune-induced inflammatory disorders.

  9. Mnk Kinases in Cytokine Signaling and Regulation of Cytokine Responses

    PubMed Central

    Joshi, Sonali; Platanias, Leonidas C.

    2013-01-01

    The kinases Mnk1 and Mnk2 are activated downstream of the p38 MAPK and MEK/ERK signaling pathways. Extensive work over the years has shown that these kinases control phosphorylation of the eukaryotic initiation factor 4E (eIF4E) and regulate engagement of other effector elements, including hnRNPA1 and PSF. Mnk kinases are ubiquitously expressed and play critical roles in signaling for various cytokine receptors, while there is emerging evidence that they have important functions as mediators of pro-inflammatory cytokine production. In this review the mechanisms of activation of MNK pathways by cytokine receptors are addressed and their roles in diverse cytokine-dependent biological processes are reviewed. The clinical-translational implications of such work and the relevance of future development of specific MNK inhibitors for the treatment of malignancies and auto-immune disorders are discussed. PMID:23710261

  10. KLF2 deficiency in T cells results in unrestrained cytokine production and bystander chemokine receptor upregulation

    PubMed Central

    Weinreich, Michael A.; Takada, Kensuke; Skon, Cara; Reiner, Steven L.; Jameson, Stephen C.; Hogquist, Kristin A.

    2009-01-01

    SUMMARY The transcription factor KLF2 regulates T cell trafficking by promoting expression of the lipid binding receptor, S1P1, and the selectin, CD62L. Recently, it was proposed that KLF2 also represses the expression of chemokine receptors. We confirm the upregulation of the chemokine receptor CXCR3 on KLF2 deficient T cells. However, we show that this is a cell nonautonomous effect, as revealed by CXCR3 upregulation on WT bystander cells in mixed bone marrow chimeras with KLF2 deficient cells. Furthermore, we show that KLF2 deficient T cells overproduce IL-4, leading to the upregulation of CXCR3 through an IL-4 receptor and eomesodermin dependent pathway. Consistent with the increased IL-4 production, we find high levels of serum IgE in mice with T cell specific KLF2 deficiency. Our findings support a model where KLF2 regulates T cell trafficking by direct regulation of S1P1 and CD62L, and restrains spontaneous cytokine production in naive T cells. PMID:19592277

  11. Importance of hydroxyapatite particles characteristics on cytokines production by human monocytes in vitro.

    PubMed

    Laquerriere, Patrice; Grandjean-Laquerriere, Alexia; Jallot, Edouard; Balossier, Gerard; Frayssinet, Patrick; Guenounou, Moncef

    2003-07-01

    Calcium phosphate bioceramics have been applied as bone substitutes for several decades. Aseptic loosening after total joint arthroplasty is a major problem in orthopaedic surgery. Hydroxyapatite particles from materials wear have been reported as the main cause of implant failure. For this reason, an investigation into possible wear particles from materials used in the implant may lead to longevity after arthroplasty. Monocytes are among the first cells to colonize the inflammatory site. In the present study, we have evaluated the inflammatory response after exposition to particles with different characteristics (size, sintering temperature and shape). Our data demonstrate that the most important characteristic was the shape and the size of the particles. The needle shaped particles induced the larger production of TNF-alpha, IL-6 and IL-10 by cells. To a less manner, the smallest particles induced an increase of the expression and production of the cytokines studied (TNF-alpha, IL-6 and IL-10). The sintering temperature appeared to be a less important characteristic even though it was involved in the dissolution/precipitation process.

  12. PRODUCTS OF ACTIVATED LYMPHOCYTES

    PubMed Central

    Sorg, Clemens; Bloom, Barry R.

    1973-01-01

    General methods were developed and applied to the biosynthesis and purification of products of activated lymphocytes available in minute quantities. The activity studied here was the migration inhibitory factor (MIF) produced by purified protein derivative (PPD)- or concanavalin A (Con A)-stimulated lymphocytes obtained from one guinea pig or less. The methods selected yielded results in terms of two chemical parameters characteristic of the molecules involved, namely Kd on Sephadex G-75 and isoionic point, pI, on isoelectric focusing. When supernatants were fractionated on G-75 columns, there were several areas even in control supernatants which produced migration inhibition relative to medium controls. However, in PPD- and Con A-stimulated supernatants, at least one peak of MIF activity was found solely in the stimulated cultures, with a Kd of 0.15. A double-labeling technique was used to characterize the proteins of this peak. Control, unstimulated cultures were labeled with [14C]leucine and stimulated cultures were labeled with [3H]leucine. After mixing the supernatants and G-75 filtration, a major "ratiolabeled" broad peak. i.e. one with increased 3H/14C ratio, was found. When a narrow portion of this peak about Kd 0.15, containing most of the MIF activity, was subjected to analytical isoelectric focusing, all of the label was associated with proteins of lower net charge than albumin. A unique ratiolabeled peak was found in PPD- and Con A-stimulated fractions with a pI of approx. 5.3. A micropreparative isoelectric focusing technique was developed and yielded MIF activity in the same region as the major ratiolabeled peak. Further study will be required to ascertain whether the ratiolabeled protein is MIF. By following the Kd, pI, and 3H/14C labeling ratio, at least 14 products of activated lymphocytes, synthesized either de novo or in increased amounts, could be distinguished. PMID:4688317

  13. In vitro lymphoproliferative response and cytokine production in mice with experimental disseminated candidiasis

    PubMed Central

    Khosravi, Ali Reza; Shokri, Hojjatollah; Eshghi, Shahin

    2017-01-01

    Objective(s): Systemic candidiasis is an infection of Candida albicans (C. albicans) causing disseminated disease and sepsis, invariably when host defenses are compromised. We investigated the histopathological changes as well as the lymphoproliferative responses and cytokine production of splenic cells after stimulation with Concanavalin A (Con A) and Pokeweed mitogen (PWM) in mice with disseminated candidiasis. Materials and Methods: Lymphoproliferative responses were stimulated in vitro with Con A (1 µg/ml) and PWM (1 µg/ml) mitogens in Roswell Park Memorial Institute (RPMI) 1640 media, and the production of interferon (IFN)-γ and interleukin-4 (IL-4) in the supernatants was measured by enzyme-linked immunosorbent assay (ELISA). Results: The results revealed that C. albicans organisms multiplied to a greater extent in the kidneys than in the liver and spleen of infected mice. The most predominant forms of C. albicans in different parts of the kidneys were yeast mixed with hyphal forms. Infected mice had a significantly increased proliferative response when splenocytes were stimulated with PWM (2.0±0.16) and Con A (1.9±0.19) (P<0.05). PWM and Con A-stimulated production of IFN-γ significantly tended to be higher in infected mice (PWM: 68.4±14.0 pg/ml; Con A: 53.7±17.3 pg/ml) when compared to controls (P<0.05). Stimulation with PWM and Con A showed no differences in IL-4 production between infected mice and controls. Conclusion: These findings demonstrated a significant increase in both cell proliferation and IFN-γ secretion in supernatants of PWM and Con A- stimulated splenocyte cultures obtained from mice with disseminated candidiasis. PMID:28293397

  14. Nuclear Factor of Activated T Cells and Cytokines Gene Expression of the T Cells in AIDS Patients with Immune Reconstitution Inflammatory Syndrome during Highly Active Antiretroviral Therapy

    PubMed Central

    Chen, Heling; Xie, Yirui; Su, Junwei; Huang, Ying; Xu, Lijun; Yin, Michael; Zhou, Qihui

    2017-01-01

    Background. The etiology of immune reconstitution inflammatory syndrome (IRIS) in AIDS patients after the initiation of HAART remains unknown. Several researches indicated that the development of IRIS is associated with the production and variation of cytokines, whose gene expression are closely related to the Ca2+/CN-nuclear factor of activated T cells (NFAT) pathway. Methods. We studied the expression of NFAT isoforms and their major target cytokines genes in peripheral blood CD3+ T cells of subjects through fluorescence quantitative PCR and explored the expression changes of these genes before and after HAART. Results. After the initiation of HARRT, NFAT1, IL-6, and IL-8 gene expression showed a reversal trend in the CD3+ T cells of the IRIS group and changed from low expression before HARRT to high expression after HARRT. In particular, the relative gene expression of NFAT1 was markedly higher compared with the other three isoforms. The IRIS group also showed higher NFAT4, NFAT2, NFAT1, IL-1β, IL-10, IL-2, IL-18, and TNF-α gene expression than the non-IRIS group. Conclusion. This study suggested that high expression levels of IL-2, IL-6, IL-8, TNF-α, IL-1β, IL-10, IL-12, and IL-18 can predict the risk of IRIS. The increased expression of NFAT1 and NFAT4 may promote the expression of cytokines, such as IL-6, IL-8, and TNF-α, which may promote the occurrence of IRIS. PMID:28316373

  15. Bromelain inhibits lipopolysaccharide-induced cytokine production in human THP-1 monocytes via the removal of CD14.

    PubMed

    Huang, Jing-Rong; Wu, Chia-Chuan; Hou, Rolis Chien-Wei; Jeng, Kee-Ching

    2008-01-01

    Bromelain has been reported to have anti-inflammatory and immunomodulatory effects. However, the anti-inflammatory mechanism of bromelain is unclear. Therefore, we investigated the effect of bromelain on cytokine production from lipopolysaccharide (LPS)-stimulated human peripheral blood mononuclear cells (PBMC) and monocytic leukemia THP-1 cells. The result showed that bromelain (50-100 microg/ml) significantly and reversibly reduced tumor necrosis factor (TNF)-alpha interleukin- (IL)-1beta and IL-6 from LPS-induced PBMC and THP-1 cells. This effect was correlated with reduced LPS-induced TNF-alpha mRNA and NF-kappaB activity in THP-1 cells. In addition, bromelain dose-dependently inhibited LPS-induced prostaglandin E(2), thromboxane B(2) and COX-2 mRNA but not COX-1 mRNA. Importantly, bromelain degraded TNF-alpha and IL-1beta molecules, reduced the expression of surface marker CD14 but not Toll-like receptor 4 from THP-1 cells. Taken together, the results suggest that the suppression of signaling pathways by bromelain's proteolytic activity may contribute to the anti-inflammatory activity of bromelain.

  16. Technical Advance: Autofluorescence-based sorting: rapid and nonperturbing isolation of ultrapure neutrophils to determine cytokine production

    PubMed Central

    Dorward, David A.; Lucas, Christopher D.; Alessandri, Ana L.; Marwick, John A.; Rossi, Fiona; Dransfield, Ian; Haslett, Christopher; Dhaliwal, Kevin; Rossi, Adriano G.

    2013-01-01

    The technical limitations of isolating neutrophils without contaminating leukocytes, while concurrently minimizing neutrophil activation, is a barrier to determining specific neutrophil functions. We aimed to assess the use of FACS for generating highly pure quiescent neutrophil populations in an antibody-free environment. Peripheral blood human granulocytes and murine bone marrow-derived neutrophils were isolated by discontinuous Percoll gradient and flow-sorted using FSC/SSC profiles and differences in autofluorescence. Postsort purity was assessed by morphological analysis and flow cytometry. Neutrophil activation was measured in unstimulated-unsorted and sorted cells and in response to fMLF, LTB4, and PAF by measuring shape change, CD62L, and CD11b expression; intracellular calcium flux; and chemotaxis. Cytokine production by human neutrophils was also determined. Postsort human neutrophil purity was 99.95% (sem=0.03; n=11; morphological analysis), and 99.68% were CD16+ve (sem=0.06; n=11), with similar results achieved for murine neutrophils. Flow sorting did not alter neutrophil activation or chemotaxis, relative to presorted cells, and no differences in response to agonists were observed. Stimulated neutrophils produced IL-1β, although to a lesser degree than CXCL8/IL-8. The exploitation of the difference in autofluorescence between neutrophils and eosinophils by FACS is a quick and effective method for generating highly purified populations for subsequent in vitro study. PMID:23667167

  17. Modulation of pro- and anti-inflammatory cytokines in active and latent tuberculosis by coexistent Strongyloides stercoralis infection.

    PubMed

    George, Parakkal Jovvian; Pavan Kumar, Nathella; Jaganathan, Jeeva; Dolla, Chandrakumar; Kumaran, Paul; Nair, Dina; Banurekha, Vaithilingam V; Shen, Kui; Nutman, Thomas B; Babu, Subash

    2015-12-01

    Helminth infections are known to induce modulation of both innate and adaptive immune responses in active and latent tuberculosis (TB). However, the role of helminth infections in modulating systemic cytokine responses in active and latent tuberculosis (LTB) is not known. To define the systemic cytokine levels in helminth-TB coinfection, we measured the circulating plasma levels of Type 1, Type 2, Type 17, other pro-inflammatory and regulatory cytokines in individuals with active TB (ATB) with or without coexistent Strongyloides stercoralis (Ss) infection by multiplex ELISA. Similarly, we also measured the same cytokine levels in individuals with LTB with or without concomitant Ss infection in a cross-sectional study. Our data reveal that individuals with ATB or LTB and coexistent Ss infection have significantly lower levels of Type 1 (IFNγ, TNFα and IL-2) and Type 17 (IL-17A and IL-17F) cytokines compared to those without Ss infection. In contrast, those with ATB and LTB with Ss infection have significantly higher levels of the regulatory cytokines (IL-10 and TGFβ), and those with LTB and Ss infection also have significantly higher levels of Type 2 cytokines (IL-4, IL-5 and IL-13) as well. Finally, those with LTB (but not ATB) exhibit significantly lower levels of other pro-inflammatory cytokines (IFNα, IFNβ, IL-6, IL-12 and GM-CSF). Our data therefore reveal a profound effect of Ss infection on the systemic cytokine responses in ATB and LTB and indicate that coincident helminth infections might influence pathogenesis of TB infection and disease.

  18. Th1 cytokines promote T-cell binding to antigen-presenting cells via enhanced hyaluronan production and accumulation at the immune synapse

    PubMed Central

    Bollyky, Paul L; Evanko, Stephen P; Wu, Rebecca P; Potter-Perigo, Susan; Long, S Alice; Kinsella, Brian; Reijonen, Helena; Guebtner, Kelly; Teng, Brandon; Chan, Christina K; Braun, Kathy R; Gebe, John A; Nepom, Gerald T; Wight, Thomas N

    2010-01-01

    Hyaluronan (HA) production by dendritic cells (DCs) is known to promote antigen presentation and to augment T-cell activation and proliferation. We hypothesized that pericellular HA can function as intercellular ‘glue' directly mediating T cell–DC binding. Using primary human cells, we observed HA-dependent binding between T cells and DCs, which was abrogated upon pre-treatment of the DCs with 4-methylumbelliferone (4-MU), an agent which blocks HA synthesis. Furthermore, T cells regulate HA production by DCs via T cell-derived cytokines in a T helper (Th) subset-specific manner, as demonstrated by the observation that cell-culture supernatants from Th1 but not Th2 clones promote HA production. Similar effects were seen upon the addition of exogenous Th1 cytokines, IL-2, interferon γ (IFN-γ) and tumor necrosis factor α (TNF-α). The critical factors which determined the extent of DC–T cell binding in this system were the nature of the pre-treatment the DCs received and their capacity to synthesize HA, as T-cell clones which were pre-treated with monensin, added to block cytokine secretion, bound equivalently irrespective of their Th subset. These data support the existence of a feedforward loop wherein T-cell cytokines influence DC production of HA, which in turn affects the extent of DC–T cell binding. We also document the presence of focal deposits of HA at the immune synapse between T-cells and APC and on dendritic processes thought to be important in antigen presentation. These data point to a pivotal role for HA in DC–T cell interactions at the IS. PMID:20228832

  19. Th1 cytokines promote T-cell binding to antigen-presenting cells via enhanced hyaluronan production and accumulation at the immune synapse.

    PubMed

    Bollyky, Paul L; Evanko, Stephen P; Wu, Rebecca P; Potter-Perigo, Susan; Long, S Alice; Kinsella, Brian; Reijonen, Helena; Guebtner, Kelly; Teng, Brandon; Chan, Christina K; Braun, Kathy R; Gebe, John A; Nepom, Gerald T; Wight, Thomas N

    2010-05-01

    Hyaluronan (HA) production by dendritic cells (DCs) is known to promote antigen presentation and to augment T-cell activation and proliferation. We hypothesized that pericellular HA can function as intercellular 'glue' directly mediating T cell-DC binding. Using primary human cells, we observed HA-dependent binding between T cells and DCs, which was abrogated upon pre-treatment of the DCs with 4-methylumbelliferone (4-MU), an agent which blocks HA synthesis. Furthermore, T cells regulate HA production by DCs via T cell-derived cytokines in a T helper (Th) subset-specific manner, as demonstrated by the observation that cell-culture supernatants from Th1 but not Th2 clones promote HA production. Similar effects were seen upon the addition of exogenous Th1 cytokines, IL-2, interferon gamma (IFN-gamma) and tumor necrosis factor alpha (TNF-alpha). The critical factors which determined the extent of DC-T cell binding in this system were the nature of the pre-treatment the DCs received and their capacity to synthesize HA, as T-cell clones which were pre-treated with monensin, added to block cytokine secretion, bound equivalently irrespective of their Th subset. These data support the existence of a feedforward loop wherein T-cell cytokines influence DC production of HA, which in turn affects the extent of DC-T cell binding. We also document the presence of focal deposits of HA at the immune synapse between T-cells and APC and on dendritic processes thought to be important in antigen presentation. These data point to a pivotal role for HA in DC-T cell interactions at the IS.

  20. Cytokine profile and natural killer cell activity in Listeria monocytogenes infected mice treated orally with Petiveria alliacea extract.

    PubMed

    Queiroz, M L; Quadros, M R; Santos, L M

    2000-08-01

    In this work, we investigated the effects of Petiveria alliacea extract on the production of Th1-type and Th2-type cytokines and on NK cells activity in normal and Listeria monocytogenes infected mice. Our results demonstrated that in normal/non-infected mice P. alliacea administration led to increased levels of Interleukin-2 (IL-2). The infection alone enhanced INF-gamma levels and NK cell activity at 48 and 72 hours of infection. The treatment with five consecutive doses of 1000 mg/kg/day of P. alliacea extract, given previously to infection, led to further increases in IL-2 levels, in relation to normal/non-infected/P. alliacea treated controls, and in INF-gamma levels at 72 h of infection, compared to infected mice. On the other hand, the production of IL-4 and IL-10 were not altered either by the infection or by the treatment with P. alliacea extract. NK cells activity increased at 48 h and 72 h following the inoculation of the bacteria. When mice were treated with P. alliacea previously to infection, NK activity was higher than that observed at 48 h, 72 h and 120 h of infection in the infected animal. Based on these findings we suggest that P. alliacea up-regulates anti-bacterial immune response by enhancing both Th1 function and the activity of NK cells.

  1. Surface protein Esp enhances pro-inflammatory cytokine expression through NF-κB activation during enterococcal infection.

    PubMed

    Zou, Jun; Shankar, Nathan

    2016-01-01

    Enterococcal surface protein (Esp) is encoded on a pathogenicity island in Enterococcus faecalis and E. faecium and is involved in biofilm formation and binding to epithelial cells. In this study, using Esp-expressing E. faecalis MMH594 and its isogenic Esp-deficient strain, as well as purified Esp, we show that Esp is sufficient for activation of NF-κB and the subsequent production of pro-inflammatory cytokines IL-1β and TNF-α in macrophages in vitro. In a mouse peritonitis model, we also show that mice infected with Esp-expressing E. faecalis showed comparatively higher levels of cytokines TNF-α, IL-1β and IL-6 in peritoneal fluid, and IL-6 in serum. Moreover, neutrophil infiltration and tissue damage in the liver was higher in the mice infected with the Esp-expressing strain compared with mice infected with the Esp-deficient mutant. These results add Esp to the growing list of enterococcal virulence factors that can modulate inflammation during infection and has implications for enterococcal pathogenesis.

  2. Borrelia-induced cytokine production is mediated by spleen tyrosine kinase (Syk) but is Dectin-1 and Dectin-2 independent.

    PubMed

    Oosting, Marije; Buffen, Kathrin; Cheng, Shih-Chin; Verschueren, Ineke C; Koentgen, Frank; van de Veerdonk, Frank L; Netea, Mihai G; Joosten, Leo A B

    2015-12-01

    Although it is known that Borrelia species express sugar-like structures on their outer surface, not much is known about the role of these structures in immune recognition by host cells. Fungi, like Candida albicans, are mainly recognized by C-type lectin receptors, in specific Dectin-1 and Dectin-2. In this study we assessed the role of Dectin-1 and Dectin-2 in the recognition process of Borrelia spirochetes. Using specific inhibitors against these receptors on human cells did not influenced cytokine production. Individuals carrying a SNP leading to an early stop codon in the DECTIN-1 gene also did not lead to differential induction of Borrelia-dependent cytokines. After injection of live Borrelia into knee joints of Dectin-2 deficient mice a trend towards lower inflammation was observed. Inhibition of Syk in human cells resulted in lower cytokine production after Borrelia stimulation. In conclusion, Dectin-1 and Dectin-2 seem not to play a major role in Borrelia recognition or Borrelia-induced inflammation. However, Syk seems to be involved in Borrelia-induced cytokine production.

  3. The Effect of IL-4 Gene Polymorphisms on Cytokine Production in Patients with Chronic Periodontitis and in Healthy Controls

    PubMed Central

    Bartova, Jirina; Janatova, Tatjana; Svobodova, Kazi; Fassmann, Antonin; Belacek, Jaromir

    2014-01-01

    Chronic periodontitis (CP) is an inflammatory disease of the teeth-supporting tissues in which genetic predisposition, dental plaque bacteria, and immune mechanisms all play important roles. The aim of this study was to evaluate the occurrence of IL-4 gene polymorphisms in chronic periodontitis and to investigate the association between polymorphisms and cytokines production after bacterial stimulation. Sixty-two subjects (47 CP patients and 15 healthy controls) with detected two polymorphisms in the IL-4 gene (-590C/T and intron 3 VNTR) were examined. Production of cytokines (IL-1α, IL-1β, IL-4, IL-5, IL-6, IL-10, IL-17, TNFα, INFγ, and VEGF) was studied after in vitro stimulation of isolated peripheral blood by mitogens (Pokeweed mitogen, Concanavalin A), dental plaque bacteria (Aggregatibacter actinomycetemcomitans, Tannerella forsythia, Porphyromonas gingivalis, and Prevotella intermedia), and Heat Shock Protein (HSP) 70 by the Luminex multiplex cytokine analysis system. The results were correlated with IL-4 genotypes in patients with CP and healthy controls. The mononuclear cells isolated from peripheral blood of CP patients with selected IL-4 polymorphisms significantly altered the production of IFNγ, IL-10, IL-1β, IL-1α, TNFα, and IL-6 after stimulation by HSP 70 or selected bacteria (from P < 0.001 to P < 0.05). IL-4 gene polymorphisms may influence the function of mononuclear cells to produce not only interleukin-4 but also other cytokines, especially in patients with CP. PMID:25530681

  4. Modulation of the pro-inflammatory cytokines and matrix metalloproteinases production in co-cultivated human keratinocytes and melanocytes.

    PubMed

    Decean, H; Perde-Schrepler, M; Tatomir, C; Fischer-Fodor, E; Brie, I; Virag, P

    2013-10-01

    The human epidermis exerts immunoregulatory functions through the variety of cytokines and other molecules elaborated by keratinocytes and melanocytes. Their constitutive production is very low; however, considerably increased upon stimulation. In vivo, keratinocytes and melanocytes have a typical exposure in the skin, referred as melanocyte epidermal unit. In the present study we co-cultivated these cells in vitro proposing to elucidate some communication links in close cell-to-cell association. We assessed the amounts of IL-6, IL-8, and matrix metalloproteinases (MMP-2 and MMP-9) in individually and co-cultured cells, exposed or not to UVB radiation. Normal human epidermal keratinocytes and melanocytes were grown in specific media and supplements. Cells were exposed to UVB radiation (100 mJ/cm(2)) to create comparable stress to the environmental one. Cytokines were determined with ELISA and confirmed with Western blot and metalloproteinases with gel zimography. Pure cultures of keratinocytes and melanocytes released low amounts of cytokines and metalloproteinases, these secretions being enhanced by UVB irradiation. In co-cultures, the cell-to-cell proximity triggered signals which markedly augmented the cytokines' secretions, whereas metalloproteinases were down-regulated. UVB irradiation did not influence either of these secretions in co-cultures. Concurrently with the highest levels of the pro-inflammatory cytokines, MMP-9 was up-regulated creating pro-inflammatory conditions and premises for changes in cellular survival, differentiation and phenotype. A complex network of interactions occurred between keratinocytes and melanocytes in co-cultures, resulting in modulated pro-inflammatory cytokines and metalloproteinases productions. Therefore, any disturbances in the microenvironmental signaling system and its molecular constituents may result in inflammation or even tumorigenesis in the epidermis.

  5. Sulfur Mustard Induced Cytokine Production and Cell Death: Investigating the Potential Roles of the p38, p53, and NF-kappaB Signaling Pathways with RNA Interference

    DTIC Science & Technology

    2010-01-01

    vation pathway in cytokine production in SM-exposed NHEK. Although there are some exceptions, the clas- sical NF-κB activation pathway with the p50–p65...military campaigns since the First World War. Skin is a major target tissue, and clin- ical presentations of SM cutaneous injuries are char- acterized by...3,5]. Both p38 and NF-κB are activated in SM-exposed cells [7,9–11], but it is NF-κB that has long been im- plicated as playing a primary role in SM

  6. Neutrophil migration across monolayers of cytokine-prestimulated endothelial cells: a role for platelet-activating factor and IL-8

    PubMed Central

    1992-01-01

    In a previous study we observed that neutrophils respond with a rapid rise in [Ca2+]i during adherence to cytokine-activated endothelial cells (EC), caused by EC membrane-associated platelet-activating factor (PAF). In the present study, we investigated whether this form of PAF was important in neutrophil adherence and migration across monolayers of rIL-1 beta- or rTNF alpha-prestimulated EC. PAF receptor antagonists prevented neutrophil migration across cytokine-pretreated EC by approximately 60% (P less than 0.005) without interfering with the process of adherence. The antagonists WEB 2086 and L-652,731 had no effect on neutrophil migration across resting EC induced by formylmethionyl-leucyl-phenylalanine (FMLP). A murine anti-IL-8 antiserum was found to also partially inhibit the neutrophil transmigration across cytokine-activated EC. When the anti-IL-8 antiserum was used in combination with a PAF receptor antagonist, neutrophil migration across cytokine-pretreated monolayers of EC was completely prevented. During transmigration, LAM-1 and CD44 on the neutrophils were down-modulated; both WEB 2086 and anti-IL-8 antiserum partially prevented this down-modulation caused by cytokine- prestimulated EC. Our results indicate that human neutrophils are activated and guided by EC-associated PAF and EC-derived IL-8 during the in vitro diapedesis in between cytokine-stimulated EC. PMID:1315317

  7. Suppressed cytokine production in whole blood cultures is related to iron status and is partially corrected following weight reduction in morbidly obese pre-menopausal women

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Assess ex vivo whole-blood cytokine production and its association with iron status in obese versus non-obese women. Determine the change in ex vivo whole-blood cytokine production six months after restrictive bariatric surgery in the obese group. Subjects were 17 obese (BMI: 46.6 ±7.9 kg/m2) and 1...

  8. Porcine reproductive and respiratory syndrome virus infection triggers HMGB1 release to promote inflammatory cytokine production

    SciTech Connect

    Duan, Erzhen; Wang, Dang; Luo, Rui; Luo, Jingyi; Gao, Li; Chen, Huanchun; Fang, Liurong Xiao, Shaobo

    2014-11-15

    The high mobility group box 1 (HMGB1) protein is an endogenous damage-associated molecular pattern (DAMP) molecule involved in the pathogenesis of various infectious agents. Based on meta-analysis of all publicly available microarray datasets, HMGB1 has recently been proposed as the most significant immune modulator during the porcine response to porcine reproductive and respiratory syndrome virus (PRRSV) infection. However, the function of HMGB1 in PRRSV pathogenesis is unclear. In this study, we found that PRRSV infection triggers the translocation of HMGB1 from the nucleus to the extracellular milieu in MARC-145 cells and porcine alveolar macrophages. Although HMGB1 has no effect on PRRSV replication, HMGB1 promotes PRRSV-induced NF-κB activation and subsequent expression of inflammatory cytokines through receptors RAGE, TLR2 and TLR4. Our findings show that HMGB1 release, triggered by PRRSV infection, enhances the efficiency of virus-induced inflammatory responses, thereby providing new insights into the pathogenesis of PRRSV infection. - Highlights: • PRRSV infection triggers HMGB1 release from MARC-145 cells and PAMs. • HMGB1 does not significantly affect PRRSV proliferation. • HMGB1 is involved in PRRSV-induced NF-κB activation and inflammatory responses. • HMGB1 promotes PRRSV-induced inflammatory responses through TLR2/4 and RAGE.

  9. Estrogen alters baseline and inflammatory-induced cytokine levels independent from hypothalamic-pituitary-adrenal axis activity.

    PubMed

    Shivers, Kai-Yvonne; Amador, Nicole; Abrams, Lisa; Hunter, Deirtra; Jenab, Shirzad; Quiñones-Jenab, Vanya

    2015-04-01

    Although estrogen reduces inflammatory-mediated pain responses, the mechanisms behind its effects are unclear. This study investigated if estrogen modulates inflammatory signaling by reducing baseline or inflammation-induced cytokine levels in the injury-site, serum, dorsal root ganglia (DRG) and/or spinal cord. We further tested whether estrogen effects on cytokine levels are in part mediated through hypothalamic-pituitary-adrenal (HPA) axis activation. Lumbar DRG, spinal cord, serum, and hind paw tissue were analyzed for cytokine levels in 17β-estradiol-(20%) or vehicle-(100% cholesterol) treated female rats following ovariectomy/sham adrenalectomy (OVX), adrenalectomy/sham ovariectomy (ADX) or ADX+OVX operation at baseline and post formalin injection. Formalin significantly increased pro-inflammatory interleukin (IL)-6 levels in the paw, as well as pro- and anti-inflammatory cytokine levels in the DRG, spinal cord and serum in comparison to naïve conditions. Estrogen replacement significantly increased anti-inflammatory IL-10 levels in the DRG. Centrally, estradiol significantly decreased pro-inflammatory tumor necrosis factor (TNF)-α and IL-1β levels, as well as IL-10 levels, in the spinal cord in comparison to cholesterol treatment. At both sites, most estradiol modulatory effects occurred irrespective of pain or surgical condition. Estradiol alone had no influence on cytokine release in the paw or serum, indicating that estrogen effects were site-specific. Although cytokine levels were altered between surgical conditions at baseline and following formalin administration, ADX operation did not significantly reverse estradiol's modulation of cytokine levels. These results suggest that estrogen directly regulates cytokines independent of HPA axis activity in vivo, in part by reducing cytokine levels in the spinal cord.

  10. Impact of ingestion of rice bran and shitake mushroom extract on lymphocyte function and cytokine production in healthy rats.

    PubMed

    Giese, Scott; Sabell, George Richard; Coussons-Read, Mary

    2008-01-01

    This article provides a controlled evaluation of the ability of dietary supplementation with a commercially available rice bran extract modified with shitake mushroom extract (MGN-3) to support the immune function by assessing the ability of immunocytes to proliferate and produce cytokines in response to a mitogenic challenge. Twenty-four male Lewis rats were fed a control diet (Maypo sweetened oatmeal) or Maypo containing the recommended daily dose of MGN-3 for 2 weeks. This treatment modestly enhanced mitogen enhanced proliferation of splenocytes and interferon-gamma (IFN-g) production, and significantly increased proliferation of splenocytes to the superantigen toxic shock syndrome toxin-1 (TSST-1) as well as natural killer (NK) cell activity and production of interleukin-2 (IL-2) by stimulated lymphocytes. These data support the contention that ingestion of MGN-3 can support immune cell function. These data add to a growing body of data showing that ingestion of MGN-3 improves the ability of immune cells to proliferate the lyse tumor cells, suggesting that it may have utility as a dietary aid to support the immune system.

  11. Abdominal visceral adiposity influences CD4+ T cell cytokine production in pregnancy.

    PubMed

    Ozias, Marlies K; Li, Shengqi; Hull, Holly R; Brooks, William M; Petroff, Margaret G; Carlson, Susan E

    2015-02-01

    Women with pre-gravid obesity are at risk for pregnancy complications. While the macrophage response of obese pregnant women categorized by body mass index (BMI) has been documented, the relationship between the peripheral CD4(+) T cell cytokine profile and body fat compartments during pregnancy is unknown. In this study, third trimester peripheral CD4(+) T cell cytokine profiles were measured in healthy pregnant women [n=35; pre-pregnancy BMI: 18.5-40]. CD4(+) T cells were isolated from peripheral blood mononuclear cells and stimulated to examine their capacity to generate cytokines. Between 1 and 3weeks postpartum, total body fat was determined by dual-energy X-ray absorptiometry and abdominal subcutaneous and visceral fat masses were determined by magnetic resonance imaging. Pearson's correlation was performed to assess relationships between cytokines and fat mass. Results showed that greater abdominal visceral fat mass was associated with a decrease in stimulated CD4(+) T cell cytokine expression. IFN-gamma, TNF-alpha, IL-12p70, IL-10 and IL-17A were inversely related to visceral fat mass. Chemokines CCL3 and IL-8 and growth factors G-CSF and FLT-3L were also inversely correlated. Additionally, total body fat mass was inversely correlated with FGF-2 while abdominal subcutaneous fat mass and BMI were unrelated to any CD4(+) T cell cytokine. In conclusion, lower responsiveness of CD4(+) T cell cytokines associated with abdominal visceral fat mass is a novel finding late in gestation.

  12. Effect of recombinant α1-antitrypsin Fc-fused (AAT-Fc)protein on the inhibition of inflammatory cytokine production and streptozotocin-induced diabetes.

    PubMed

    Lee, Siyoung; Lee, Youngmin; Hong, Kwangwon; Hong, Jaewoo; Bae, Suyoung; Choi, Jida; Jhun, Hyunjhung; Kwak, Areum; Kim, Eunsom; Jo, Seunghyun; Dinarello, Charles A; Kim, Soohyun

    2013-05-20

    α1-Antitrypsin (AAT) is a member of the serine proteinase inhibitor family that impedes the enzymatic activity of serine proteinases, including human neutrophil elastase, cathepsin G and neutrophil proteinase 3. Here, we expressed recombinant AAT by fusing the intact AAT gene to the constant region of IgG1 to generate soluble recombinant AAT-Fc protein. The recombinant AAT-Fc protein was produced in Chinese hamster ovary (CHO) cells and purified using mini-protein A affinity chromatography. Recombinant AAT-Fc protein was tested for antiinflammatory function and AAT-Fc sufficiently suppressed tumor necrosis factor (TNF)-α-induced interleukin (IL)-6 in human peripheral blood mononuclear cells (PBMCs) and inhibited cytokine-induced TNFα by different cytokines in mouse macrophage Raw 264.7 cells. However, AAT-Fc failed to suppress lipopolysaccharide-induced cytokine production in both PBMCs and macrophages. In addition, our data showed that AAT-Fc blocks the development of hyperglycemia in a streptozotocin-induced mouse model of diabetes. Interestingly, we also found that plasma-derived AAT specifically inhibited the enzymatic activity of elastase but that AAT-Fc had no inhibitory effect on elastase activity.

  13. Role of antigen presentation in the production of pro-inflammatory cytokines in obese adipose tissue.

    PubMed

    Majdoubi, Abdelilah; Kishta, Osama A; Thibodeau, Jacques

    2016-06-01

    Type II diabetes regroups different physiological anomalies that ultimately lead to low-grade chronic inflammation, insulin resistance and loss of pancreatic β-cells. Obesity is one of the best examples of such a condition that can develop into Metabolic Syndrome, causing serious health problems of great socio-economic consequences. The pathological outcome of obesity has a genetic basis and depends on the delicate balance between pro- and anti-inflammatory effectors of the immune system. The causal link between obesity and inflammation is well established. While innate immunity plays a key role in the development of a pro-inflammatory state in obese adipose tissues, it has now become clear that adaptive immune cells are also involved and participate in the cascade of events that lead to metabolic perturbations. The efficacy of some immunotherapeutic protocols in reducing the symptoms of obesity-driven metabolic syndrome in mice implicated all arms of the immune response. Recently, the production of pathogenic immunoglobulins and pro-inflammatory cytokines by B and T lymphocytes suggested an auto-immune basis for the establishment of a non-healthy obese state. Understanding the cellular landscape of obese adipose tissues and how immune cells sustain chronic inflammation holds the key to the development of targeted therapies. In this review, we emphasize the role of antigen-presenting cells and MHC molecules in obese adipose tissue and the general contribution of the adaptive arm of the immune system in inflammation-induced insulin resistance.

  14. Coculturing human endometrial epithelial cells and stromal fibroblasts alters cell-specific gene expression and cytokine production

    PubMed Central

    Chen, Joseph C.; Erikson, David W.; Piltonen, Terhi T.; Meyer, Michelle R.; Barragan, Fatima; McIntire, Ramsey H.; Tamaresis, John S.; Vo, Kim Chi; Giudice, Linda C.; Irwin, Juan C.

    2013-01-01

    Objective To determine the effects of coculturing endometrial epithelial cells (eEC) with paired endometrial stromal fibroblasts (eSF) on cell-specific gene expression and cytokine secretion patterns. Design In vitro study. Setting University research laboratory. Patient(s) Endometrial biopsies were obtained from premenopausal women. Intervention(s) Polarized eEC and subject-paired eSF were cultured for 12.5 hours alone (monoculture) or combined in a two-chamber coculture system without cell-cell contact. Cells and conditioned media were analyzed for global gene expression and cytokine secretion, respectively. Purified, endometrial tissue-derived eEC and eSF isolated by fluorescent activated cell sorting (FACS) were used as noncultured controls. Main Outcome Measure(s) Cell-specific global gene expression profiling and analysis of secreted cytokines in eEC/eSF cocultures and respective monocultures. Result(s) Transepithelial resistance, diffusible tracer exclusion, expression of tight junction proteins, and apical/basolateral vectorial secretion confirmed eEC structural and functional polarization. Distinct transcriptomes of eEC and eSF were consistent with their respective lineages and their endometrial origin. Coculture of eEC with eSF resulted in altered cell-specific gene expression and cytokine secretion. Conclusion(s) This coculture model provides evidence that interactions between endometrial functionally polarized epithelium and stromal fibroblasts affect cell-specific gene expression and cytokine secretion underscoring their relevance when modeling endometrium in vitro. PMID:23849844

  15. Neonatal Plasma Polarizes TLR4-Mediated Cytokine Responses towards Low IL-12p70 and High IL-10 Production via Distinct Factors

    PubMed Central

    Belderbos, Mirjam E.; Levy, Ofer; Stalpers, Femke; Kimpen, Jan L.; Meyaard, Linde; Bont, Louis

    2012-01-01

    Human neonates are highly susceptible to infection, which may be due in part to impaired innate immune function. Neonatal Toll-like receptor (TLR) responses are biased against the generation of pro-inflammatory/Th1-polarizing cytokines, yet the underlying mechanisms are incompletely defined. Here, we demonstrate that neonatal plasma polarizes TLR4-mediated cytokine production. When exposed to cord blood plasma, mononuclear cells (MCs) produced significantly lower TLR4-mediated IL-12p70 and higher IL-10 compared to MC exposed to adult plasma. Suppression by neonatal plasma of TLR4-mediated IL-12p70 production, but not induction of TLR4-mediated IL-10 production, was maintained up to the age of 1 month. Cord blood plasma conferred a similar pattern of MC cytokine responses to TLR3 and TLR8 agonists, demonstrating activity towards both MyD88-dependent and MyD88-independent agonists. The factor causing increased TLR4-mediated IL-10 production by cord blood plasma was heat-labile, lost after protein depletion and independent of lipoprotein binding protein (LBP) or soluble CD14 (sCD14). The factor causing inhibition of TLR4-mediated IL-12p70 production by cord blood plasma was resistant to heat inactivation or protein depletion and was independent of IL-10, vitamin D and prostaglandin E2. In conclusion, human neonatal plasma contains at least two distinct factors that suppress TLR4-mediated IL-12p70 production or induce IL-10 or production. Further identification of these factors will provide insight into the ontogeny of innate immune development and might identify novel targets for the prevention and treatment of neonatal infection. PMID:22442690

  16. Suppressor of Cytokine Signaling 2 Negatively Regulates NK Cell Differentiation by Inhibiting JAK2 Activity

    PubMed Central

    Kim, Won Sam; Kim, Mi Jeong; Kim, Dong Oh; Byun, Jae-Eun; Huy, Hangsak; Song, Hae Young; Park, Young-Jun; Kim, Tae-Don; Yoon, Suk Ran; Choi, Eun-Ji; Jung, Haiyoung; Choi, Inpyo

    2017-01-01

    Suppressor of cytokine signaling (SOCS) proteins are negative regulators of cytokine responses. Although recent reports have shown regulatory roles for SOCS proteins in innate and adaptive immunity, their roles in natural killer (NK) cell development are largely unknown. Here, we show that SOCS2 is involved in NK cell development. SOCS2−/− mice showed a high frequency of NK cells in the bone marrow and spleen. Knockdown of SOCS2 was associated with enhanced differentiation of NK cells in vitro, and the transplantation of hematopoietic stem cells (HSCs) into congenic mice resulted in enhanced differentiation in SOCS2−/− HSCs. We found that SOCS2 could inhibit Janus kinase 2 (JAK2) activity and JAK2-STAT5 signaling pathways via direct interaction with JAK2. Furthermore, SOCS2−/− mice showed a reduction in lung metastases and an increase in survival following melanoma challenge. Overall, our findings suggest that SOCS2 negatively regulates the development of NK cells by inhibiting JAK2 activity via direct interaction. PMID:28383049

  17. Over-expression of GTP-cyclohydrolase 1 feedback regulatory protein attenuates LPS and cytokine-stimulated nitric oxide production.

    PubMed

    Nandi, Manasi; Kelly, Peter; Vallance, Patrick; Leiper, James

    2008-02-01

    GTP-cyclohydrolase 1 (GTP-CH1) catalyses the first and rate-limiting step for the de novo production of tetrahydrobiopterin (BH(4)), an essential cofactor for nitric oxide synthase (NOS). The GTP-CH1-BH(4) pathway is emerging as an important regulator in a number of pathologies associated with over-production of nitric oxide (NO) and hence a more detailed understanding of this pathway may lead to novel therapeutic targets for the treatment of certain vascular diseases. GTP-CH1 activity can be inhibited by BH(4) through its protein-protein interactions with GTP-CH1 regulatory protein (GFRP), and transcriptional and post-translational modification of both GTP-CH1 and GFRP have been reported in response to proinflammatory stimuli. However, the functional significance of GFRP/GTP-CH1 interactions on NO pathways has not yet been demonstrated. We aimed to investigate whether over-expression of GFRP could affect NO production in living cells. Over-expression of N-terminally Myc-tagged recombinant human GFRP in the murine endothelial cell line sEnd 1 resulted in no significant effect on basal BH(4) nor NO levels but significantly attenuated the rise in BH(4) and NO observed following lipopolysaccharide and cytokine stimulation of cells. This study demonstrates that GFRP can play a direct regulatory role in iNOS-mediated NO synthesis and suggests that the allosteric regulation of GTP-CH1 activity by GFRP may be an important mechanism regulating BH(4) and NO levels in vivo.

  18. Pimaradienoic Acid Inhibits Carrageenan-Induced Inflammatory Leukocyte Recruitment and Edema in Mice: Inhibition of Oxidative Stress, Nitric Oxide and Cytokine Production

    PubMed Central

    Casagrande, Rubia; Verri, Waldiceu A.

    2016-01-01

    Pimaradienoic acid (PA; ent-pimara-8(14),15-dien-19-oic acid) is a pimarane diterpene found in plants such as Vigueira arenaria Baker (Asteraceae) in the Brazilian savannas. Although there is evidence on the analgesic and in vitro inhibition of inflammatory signaling pathways, and paw edema by PA, its anti-inflammatory effect deserves further investigation. Thus, the objective of present study was to investigate the anti-inflammatory effect of PA in carrageenan-induced peritoneal and paw inflammation in mice. Firstly, we assessed the effect of PA in carrageenan-induced leukocyte recruitment in the peritoneal cavity and paw edema and myeloperoxidase activity. Next, we investigated the mechanisms involved in the anti-inflammatory effect of PA. The effect of PA on carrageenan-induced oxidative stress in the paw skin and peritoneal cavity was assessed. We also tested the effect of PA on nitric oxide, superoxide anion, and inflammatory cytokine production in the peritoneal cavity. PA inhibited carrageenan-induced recruitment of total leukocytes and neutrophils to the peritoneal cavity in a dose-dependent manner. PA also inhibited carrageenan-induced paw edema and myeloperoxidase activity in the paw skin. The anti-inflammatory mechanism of PA depended on maintaining paw skin antioxidant activity as observed by the levels of reduced glutathione, ability to scavenge the ABTS cation and reduce iron as well as by the inhibition of superoxide anion and nitric oxide production in the peritoneal cavity. Furthermore, PA inhibited carrageenan-induced peritoneal production of inflammatory cytokines TNF-α and IL-1β. PA presents prominent anti-inflammatory effect in carrageenan-induced inflammation by reducing oxidative stress, nitric oxide, and cytokine production. Therefore, it seems to be a promising anti-inflammatory molecule that merits further investigation. PMID:26895409

  19. Beneficial metabolic activities of inflammatory cytokine interleukin 15 in obesity and type 2 diabetes.

    PubMed

    Ye, Jianping

    2015-06-01

    In obesity, chronic inflammation is believed to induce insulin resistance and impairs adipose tissue function. Although this view is supported by a large body of literature, it has been challenged by growing evidence that pro-inflammatory cytokines may favor insulin sensitivity through induction of energy expenditure. In this review article, interleukin 15 (IL-15) is used as a new example to explain the beneficial effects of the proinflammatory cytokines. IL-15 is secreted by multiple types of cells including macrophages, neutrophils and skeletal muscle cells. IL-15 expression is induced in immune cells by endotoxin and in muscle cells by physical exercise. Its transcription is induced by transcription factor NF-κB. IL-15 binds to its receptor that contains three different subunits (α, β and γ) to activate JAK/STAT, PI3K/Akt, IKK/NF-κB and JNK/AP1 pathways in cells. In the regulation of metabolism, IL-15 reduces weight gain without inhibiting food intake in rodents. IL-15 suppresses lipogenesis, stimulates brown fat function, improves insulin sensitivity through weight loss and energy expenditure. In human, circulating IL-15 is negatively associated with body weight. In the immune system, IL-15 stimulates proliferation and differentiation of T cells, NK cells, monocytes and neutrophils. In the anti-obesity effects of IL-15, T cells and NK cells are not required, but leptin receptor is required. In summary, evidence from human and rodents supports that the pro-inflammatory cytokine IL-15 may enhance energy expenditure to protect the body from obesity and type 2 diabetes. The mechanism of IL-15 action remains to be fully uncovered in the regulation of energy expenditure.

  20. Activation of the human. beta. sub 2 -interferon/hepatocyte-stimulating factor/interleukin 6 promoter by cytokines, viruses, and second messenger agonists

    SciTech Connect

    Ray, A.; Tatter, S.B.; May, L.T.; Sehgal, P.B. )

    1988-09-01

    The hallmark of {beta}{sub 2}-interferon (IFN-{beta}{sub 2})/hepatocyte-stimulating factor/interleukin 6 gene expression is its inducibility in different types of human cells (fibroblasts, monocytes, epithelial cells, and endothelial cells) by different stimuli, which include cytokines such as tumor necrosis factor, interleukin 1 (IL-1) and platelet-derived growth factor, different viruses, and bacterial products such as endotoxin. The activation by cytokines, viruses, and second messenger agonists of the IFN-{beta}{sub 2} promoter linked to the bacterial chloramphenicol acetyltransferase (CAT) gene was studied after transfection into HeLa cells. A chimeric gene containing IFN-{beta}{sub 2} DNA from -1180 to +13 linked to the CAT gene was inducible {approx}10-fold by phorbol 12-myristate 13-acetate (PMA), followed, in decreasing order, by pseudorabies and Sendai viruses; serum; the cytokines tumor necrosis factor, IL-1, and epidermal growth factor; the cAMP agonists BrcAMP and forskolin and the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine; poly(I){center dot}poly(C); 1,2-diacylglycerol and the calcium ionophore A23187. The region between -225 and -113 in IFN-{beta}{sub 2}, which contains DNA motifs similar to the regulatory elements in the human c-fos gene, appears to contain the major cis-acting regulatory elements responsible for the activation of the IFN-{beta}{sub 2} promoter by several different cytokines, viruses, and second messenger agonists.

  1. T cells translate individual, quantal activation into collective, analog cytokine responses via time-integrated feedbacks

    PubMed Central

    Tkach, Karen E; Barik, Debashis; Voisinne, Guillaume; Malandro, Nicole; Hathorn, Matthew M; Cotari, Jesse W; Vogel, Robert; Merghoub, Taha; Wolchok, Jedd; Krichevsky, Oleg; Altan-Bonnet, Grégoire

    2014-01-01

    Variability within isogenic T cell populations yields heterogeneous ‘local’ signaling responses to shared antigenic stimuli, but responding clones may communicate ‘global’ antigen load through paracrine messengers, such as cytokines. Such coordination of individual cell responses within multicellular populations is critical for accurate collective reactions to shared environmental cues. However, cytokine production may saturate as a function of antigen input, or be dominated by the precursor frequency of antigen-specific T cells. Surprisingly, we found that T cells scale their collective output of IL-2 to total antigen input over a large dynamic range, independently of population size. Through experimental quantitation and computational modeling, we demonstrate that this scaling is enforced by an inhibitory cross-talk between antigen and IL-2 signaling, and a nonlinear acceleration of IL-2 secretion per cell. Our study reveals how time-integration of these regulatory loops within individual cell signaling generates scaled collective responses and can be leveraged for immune monitoring. DOI: http://dx.doi.org/10.7554/eLife.01944.001 PMID:24719192

  2. Synergy between Common γ Chain Family Cytokines and IL-18 Potentiates Innate and Adaptive Pathways of NK Cell Activation.

    PubMed

    Nielsen, Carolyn M; Wolf, Asia-Sophia; Goodier, Martin R; Riley, Eleanor M

    2016-01-01

    Studies to develop cell-based therapies for cancer and other diseases have consistently shown that purified human natural killer (NK) cells secrete cytokines and kill target cells after in vitro culture with high concentrations of cytokines. However, these assays poorly reflect the conditions that are likely to prevail in vivo in the early stages of an infection and have been carried out in a wide variety of experimental systems, which has led to contradictions within the literature. We have conducted a detailed kinetic and dose-response analysis of human NK cell responses to low concentrations of IL-12, IL-15, IL-18, IL-21, and IFN-α, alone and in combination, and their potential to synergize with IL-2. We find that very low concentrations of both innate and adaptive common γ chain cytokines synergize with equally low concentrations of IL-18 to drive rapid and potent NK cell CD25 and IFN-γ expression; IL-18 and IL-2 reciprocally sustain CD25 and IL-18Rα expression in a positive feedback loop; and IL-18 synergizes with FcγRIII (CD16) signaling to augment antibody-dependent cellular cytotoxicity. These data indicate that NK cells can be rapidly activated by very low doses of innate cytokines and that the common γ chain cytokines have overlapping but distinct functions in combination with IL-18. Importantly, synergy between multiple signaling pathways leading to rapid NK cell activation at very low cytokine concentrations has been overlooked in prior studies focusing on single cytokines or simple combinations. Moreover, although the precise common γ chain cytokines available during primary and secondary infections may differ, their synergy with both IL-18 and antigen-antibody immune complexes underscores their contribution to NK cell activation during innate and adaptive responses. IL-18 signaling potentiates NK cell effector function during innate and adaptive immune responses by synergy with IL-2, IL-15, and IL-21 and immune complexes.

  3. Salivary histatin 3 inhibits heat shock cognate protein 70-mediated inflammatory cytokine production through toll-like receptors in human gingival fibroblasts

    PubMed Central

    2014-01-01

    Background Salivary histatins are bioactive peptides related to the innate immune system associated with antimicrobial activities. However, very little is known about the physiological and biological functions of histatins against host cells or their role in oral cell inflammation. Histatin 3 binds to heat shock cognate protein 70 (HSC70, a constitutively expressed heat shock protein (HSP)). It is unclear whether HSC70 is involved in the inflammatory response in oral cells. Injured oral cells release some intracellular proteins including HSC70. It is possible that released HSC70 induces toll-like receptor (TLR) activation, just as extracellular HSP70 (a stress inducible HSP) does, and that histatin 3 affects this process. Therefore, we tested the hypothesis that HSC70 activates TLR signaling and histatin 3 inhibits this activation and inflammatory cytokine production. Methods A nuclear factor (NF)-κB-dependent luciferase reporter plasmid was transfected into HEK293 cells stably expressing TLR2 with coreceptor CD14 (293-TLR2/CD14 cells) or stably expressing TLR4 with CD14 and the accessory molecule MD2 (293-TLR4/MD2-CD14 cells). The cells were stimulated with HSC70 in the presence or absence of histatin 3, and examined using luciferase assays. We also stimulated human gingival fibroblasts (HGFs) with HSC70 with or without histatin 3. Then, we analyzed the levels of inflammatory cytokines (interleukin (IL)-6 and IL-8) in the culture media. Cell proteins were analyzed using enzyme-linked immunosorbent assay and Western blotting with antibodies of mitogen-activated protein kinases and NF-κB inhibitor IκB-α, respectively. Histatin 3-bound form of HSC70 was analyzed using limited V8 protease proteolysis. Results HSC70 induced NF-κB activation in a dose-dependent manner in 293-TLR2/CD14 and 293-TLR4/MD2-CD14 cells, and histatin 3 inhibited this process and when histatin 3 binding to HSC70 was precluded by 15-deoxyspergualin, which augmented NF

  4. Melatonin and zinc treatment: distinctive modulation of cytokine production in chronic experimental Trypanosoma cruzi infection.

    PubMed

    Brazão, Vânia; Del Vecchio Filipin, Marina; Santello, Fabricia Helena; Caetano, Leony Cristina; Abrahão, Ana Amélia Carraro; Toldo, Míriam Paula Alonso; do Prado, José Clóvis

    2011-12-01

    Melatonin by exhibiting antioxidant, anti-aging, and immunomodulatory properties favorably modulate the immune function, protecting the hosts from several infectious diseases. Zinc is an essential trace element important for the efficiency of the immune system in reason of its widespread role in the activity of enzymes, transcription factors and cytokines. The etiology of Chagas' disease, caused by a protozoan parasite Trypanosoma cruzi, has been the focus of considerable discussion, although chronic phase still remains not fully understood. This study showed that zinc and melatonin treatment did not affect the percentage of both CD4+ and CD8+ T lymphocytes subsets in chronically infected animals. Increased levels of IL-2 and IL-10, as well as, enhanced thymocyte proliferation in T. cruzi infected groups under zinc and melatonin therapy was observed as compared to untreated group. Conversely, during the chronic phase of infection, macrophages counts were reduced in melatonin and zinc-melatonin treated animals. The combined actions of zinc and melatonin have beneficial effects in counteracting parasite-induced immune dysregulation, protecting animals against the harmful actions of chronic T. cruzi infection. Furthermore, our results provide an experimental basis for further studies on the role of immunomodulatory therapies.

  5. Thrombin potently stimulates cytokine production in human vascular smooth muscle cells but not in mononuclear phagocytes.

    PubMed

    Kranzhöfer, R; Clinton, S K; Ishii, K; Coughlin, S R; Fenton, J W; Libby, P

    1996-08-01

    Thrombosis frequently occurs during atherogenesis and in response to vascular injury. Accumulating evidence supports a role for inflammation in the same situation. The present study therefore sought links between thrombosis and inflammation by determining whether thrombin, which is present in active form at sites of thrombosis, can elicit inflammatory functions of human monocytes and vascular smooth muscle cells (SMCs), two major constituents of advanced atheroma. Human alpha-thrombin (EC50, approximately equal to 500 pmol/L) potently induced interleukin (IL)-6 release from SMCs. The tethered-ligand thrombin receptor appeared to mediate this effect. Furthermore, alpha-thrombin also rapidly increased levels of mRNA encoding IL-6 and monocyte chemotactic protein-1 (MCP-1) in SMCs. In contrast, only alpha-thrombin concentrations of > or = 100 nmol/L could stimulate release of IL-6 or tumor necrosis factor-alpha (TNF alpha) in peripheral blood monocytes or monocyte-derived macrophages. Lipid loading of macrophages did not augment thrombin responsiveness. Likewise, only alpha-thrombin concentrations of > or = 100 nmol/L increased levels of IL-6, IL-1 beta, MCP-1, or TNF alpha mRNA in monocytes. Differential responses of SMCs and monocytes to thrombin extended to early agonist-mediated increases in [Ca2+]i. SMCs and endothelial cells, but not monocytes, contained abundant mRNA encoding the thrombin receptor and displayed cell surface thrombin receptor expression detected with a novel monoclonal antibody. Thus, the level of thrombin receptors appeared to account for the differential thrombin susceptibility of SMCs and monocytes. These data suggest that SMCs may be more sensitive than monocytes/macrophages to thrombin activation in human atheroma. Cytokines produced by thrombin-activated SMCs may contribute to ongoing inflammation in atheroma complicated by thrombosis or subjected to angioplasty.

  6. T helper 2 and regulatory T-cell cytokine production by mast cells: a key factor in the pathogenesis of IgG4-related disease.

    PubMed

    Takeuchi, Mai; Sato, Yasuharu; Ohno, Kyotaro; Tanaka, Satoshi; Takata, Katsuyoshi; Gion, Yuka; Orita, Yorihisa; Ito, Toshihiro; Tachibana, Tomoyasu; Yoshino, Tadashi

    2014-08-01

    IgG4-related disease is a systemic disorder with unique clinicopathological features and uncertain etiological features and is frequently related to allergic disease. T helper 2 and regulatory T-cell cytokines have been reported to be upregulated in the affected tissues; thus, the production of these cytokines by T helper 2 and regulatory T cells has been suggested as an important factor in the pathogenesis of IgG4-related disease. However, it is not yet clear which cells produce these cytokines in IgG4-related disease, and some aspects of the disorder cannot be completely explained by T-cell-related processes. To address this, we analyzed paraffin-embedded sections of tissues from nine cases of IgG4-related submandibular gland disease, five cases of submandibular sialolithiasis, and six cases of normal submandibular gland in order to identify potential key players in the pathogenesis of IgG4-related disease. Real-time polymerase chain reaction analysis confirmed the significant upregulation of interleukin (IL)4, IL10, and transforming growth factor beta 1 (TGFβ1) in IgG4-related disease. Interestingly, immunohistochemical studies indicated the presence of mast cells expressing these cytokines in diseased tissues. In addition, dual immunofluorescence assays identified cells that were double-positive for each cytokine and for KIT, which is expressed by mast cells. In contrast, the distribution of T cells did not correlate with cytokine distribution in affected tissues. We also found that the mast cells were strongly positive for IgE. This observation supports the hypothesis that mast cells are involved in IgG4-related disease, as mast cells are known to be closely related to allergic reactions and are activated in the presence of elevated non-specific IgE levels. In conclusion, our results indicate that mast cells produce T helper 2 and regulatory T-cell cytokines in tissues affected by IgG4-related disease and possibly have an important role in disease

  7. Do mechanical strain and TNF-α interact to amplify pro-inflammatory cytokine production in human annulus fibrosus cells?

    PubMed

    Likhitpanichkul, Morakot; Torre, Olivia M; Gruen, Jadry; Walter, Benjamin A; Hecht, Andrew C; Iatridis, James C

    2016-05-03

    During intervertebral disc (IVD) injury and degeneration, annulus fibrosus (AF) cells experience large mechanical strains in a pro-inflammatory milieu. We hypothesized that TNF-α, an initiator of IVD inflammation, modifies AF cell mechanobiology via cytoskeletal changes, and interacts with mechanical strain to enhance pro-inflammatory cytokine production. Human AF cells (N=5, Thompson grades 2-4) were stretched uniaxially on collagen-I coated chambers to 0%, 5% (physiological) or 15% (pathologic) strains at 0.5Hz for 24h under hypoxic conditions with or without TNF-α (10ng/mL). AF cells were treated with anti-TNF-α and anti-IL-6. ELISA assessed IL-1β, IL-6, and IL-8 production and immunocytochemistry measured F-actin, vinculin and α-tubulin in AF cells. TNF-α significantly increased AF cell pro-inflammatory cytokine production compared to basal conditions (IL-1β:2.0±1.4-84.0±77.3, IL-6:10.6±9.9-280.9±214.1, IL-8:23.9±26.0-5125.1±4170.8pg/ml for basal and TNF-α treatment, respectively) as expected, but mechanical strain did not. Pathologic strain in combination with TNF-α increased IL-1β, and IL-8 but not IL-6 production of AF cells. TNF-α treatment altered F-actin and α-tubulin in AF cells, suggestive of altered cytoskeletal stiffness. Anti-TNF-α (infliximab) significantly inhibited pro-inflammatory cytokine production while anti-IL-6 (atlizumab) did not. In conclusion, TNF-α altered AF cell mechanobiology with cytoskeletal remodeling that potentially sensitized AF cells to mechanical strain and increased TNF-α-induced pro-inflammatory cytokine production. Results suggest an interaction between TNF-α and mechanical strain and future mechanistic studies are required to validate these observations.

  8. Medical-grade silicone induces release of proinflammatory cytokines in peripheral blood mononuclear cells without activating T cells.

    PubMed

    Miro-Mur, Francesc; Hindié, Mathilde; Kandhaya-Pillai, Renuka; Tobajas, Vanessa; Schwartz, Simo; Alijotas-Reig, Jaume

    2009-08-01

    For more than 40 years, silicone implants had been employed in aesthetic, cosmetic medicine, and plastic surgery. Although adverse reactions produced by these products are rare, cases of immuno-mediated reactions have been reported. To evaluate the aspects of immuno-reactivity to medical-grade silicone dermal filler, peripheral blood mononuclear cells (PBMC) of 39 individuals were studied. PBMC used include individuals with silicone injection-related delayed adverse reactions, with silicone injections, and healthy control. Silicone induced production of TNF-alpha and IL-6 in all three groups. Notably, elevated production of IL-6 was observed in nonstimulated PBMC and also the percentage of CD4(+)CD69(+) T cells was higher in PHA-stimulated PBMC from individuals with silicone injection-related adverse reactions when compared with other two groups. However, IFN-gamma was not released in silicone-stimulated or silicone+LPS-stimulated PBMC from any group and no production of IL-2 was measured indicating no proliferative response of PBMC. Subsequently, no CD4(+)CD69(+) T cells were observed in these conditions. Finally, the inflammatory response in silicone-stimulated cultures of monocyte-derived macrophages with autologous lymphocytes is lesser than that observed in PBMC. In conclusion, silicone induces a release of proinflammatory cytokines but does not act as a polyclonal activator of CD4(+) T cells. Thus, silicone is mounting an immune response in individuals with silicone-related adverse effects but is not silicone antigen-dependent.

  9. Decreasing SMPD1 activity in BEAS-2B bronchial airway epithelial cells results in increased NRF2 activity, cytokine synthesis and neutrophil recruitment.

    PubMed

    MacFadden-Murphy, Elyse; Roussel, Lucie; Martel, Guy; Bérubé, Julie; Rousseau, Simon

    2017-01-22

    Niemann-Pick disease (NPD) type B is a rare autosomal recessive disease characterized by variable levels of impairment in sphingomyelin phosphodiesterase 1 (SMPD1) activity. Lung involvement is the most important prognostic factor in NPD-B, with recurrent respiratory infections starting in infancy being the major cause of morbidity and mortality. We hypothesized that decreased SMPD1 activity impaired airway epithelium host defense response. SMPD1 activity was reduced using inducible shRNA. Surprisingly, decreasing SMPD1 activity by 50%, resulted in increased neutrophil recruitment, both at baseline and in response to bacterial stimulation. This correlated with elevated levels of cytokine mRNA shown to contribute to neutrophil recruitment in unstimulated (e.g. IL-8 and GRO-α) and infected cells (e.g. IL-8, GRO-α, GM-CSF and CCL20). Instead of preventing the host defence responses, decreased SMPD1 activity results in an inflammatory response even in the absence of infection. Moreover, decreasing SMPD1 activity resulted in a pro-oxidative shift. Accordingly, expression of an inactive mutant, SMPD1[L225P] but not the WT enzyme increased activation of the antioxidant transcription factor NRF2. Therefore, decreasing SMPD1 activity by 50% in airway epithelial cells, the equivalent of the loss of one allele, results in the accumulation of oxidants that activates NRF2 and a concomitant increased cytokine production as well as neutrophil recruitment. This can result in a chronic inflammatory state that impairs host defence similar to scenarios observe in other chronic inflammatory lung disease such as Chronic Obstructive Pulmonary Disease or Cystic Fibrosis.

  10. Determination of the Absolute Number of Cytokine mRNA Molecules within Individual Activated Human T Cells

    NASA Technical Reports Server (NTRS)

    Karr, Laurel J.; Marshall, Gwen; Hockett, Richard D.; Bucy, R. Pat; Curreri, Peter A. (Technical Monitor)

    2002-01-01

    A primary function of activated T cells is the expression and subsequent secretion of cytokines, which orchestrate the differentiation of other lymphocytes, modulate antigen presenting cell activity, and alter vascular endothelium to mediate an immune response. Since many features of immune regulation probably result from modest alterations of endogenous rates of multiple interacting processes, quantitative analysis of the frequency and specific activity of individual T cells is critically important. Using a coordinated set of quantitative methods, the absolute number of molecules of several key cytokine mRNA species in individual T cells has been determined. The frequency of human blood T cells activated in vitro by mitogens and recall protein antigens was determined by intracellular cytokine protein staining, in situ hybridization for cytokine mRNA, and by limiting dilution analysis for cytokine mRNA+ cells. The absolute number of mRNA molecules was simultaneously determined in both homogenates of the entire population of cells and in individual cells obtained by limiting dilution, using a quantitative, competitive RT-PCR assay. The absolute numbers of mRNA molecules in a population of cells divided by the frequency of individual positive cells, yielded essentially the same number of mRNA molecules per cell as direct analysis of individual cells by limiting dilution analysis. Mean numbers of mRNA per positive cell from both mitogen and antigen activated T cells, using these stimulation conditions, were 6000 for IL-2, 6300 for IFN-gamma, and 1600 for IL-4.

  11. PGRP negatively regulates NOD-mediated cytokine production in rainbow trout liver cells

    PubMed Central

    Jang, Ju Hye; Kim, Hyun; Jang, Mi Jung; Cho, Ju Hyun

    2016-01-01

    Pattern-recognition receptors (PRRs) initiate innate immunity via pathogen recognition. Recent studies suggest that signalling pathways downstream of different PRRs and their crosstalk effectively control immune responses. However, the cross-regulation among PRRs and its effects have yet to be fully described in fish. Here, we examined the crosstalk between OmPGRP-L1, a long form of PGRP in rainbow trout, and other PRRs during pathogenic infections. OmPGRP-L1 expression was increased in RTH-149 cells by iE-DAP and MDP, which are agonists of NOD1 and NOD2, respectively. The silencing of NOD1 and NOD2 specifically inhibited the upregulation of OmPGRP-L1 expression induced by their cognate ligands. Suppression of RIP2 and NF-κB activation prevented the induction of OmPGRP-L1 expression. An in silico analysis and electrophoretic mobility shift assay revealed that the promoter of OmPGRP-L1 has NF-κB binding sites, suggesting that OmPGRP-L1 is produced through the NOD-RIP2-NF-κB signalling pathway. Loss-of-function and gain-of-function experiments indicated that OmPGRP-L1 downregulates the induction of NOD-mediated pro-inflammatory cytokine expression. Mechanistically, secreted OmPGRP-L1 inhibited the activation of the NOD-induced NF-κB pathway via downregulation of TAK1 and IκBα phosphorylation through A20 expression. Our data demonstrate that OmPGRP-L1 and NODs might play interdependent roles in the inflammatory response to bacterial infections in rainbow trout. PMID:27991595

  12. Downregulating galectin-3 inhibits proinflammatory cytokine production by human monocyte-derived dendritic cells via RNA interference.

    PubMed

    Chen, Swey-Shen; Sun, Liang-Wu; Brickner, Howard; Sun, Pei-Qing

    2015-03-01

    Galectin-3 (Gal-3), a β-galactoside-binding lectin, serves as a pattern-recognition receptor (PRR) of dendritic cells (DCs) in regulating proinflammatory cytokine production. Galectin-3 (Gal-3) siRNA downregulates expression of IL-6, IL-1β and IL-23 p19, while upregulates IL-10 and IL-12 p35 in TLR/NLR stimulated human MoDCs. Furthermore, Gal-3 siRNA-treated MoDCs enhanced IFN-γ production in SEB-stimulated CD45RO CD4 T-cells, but attenuated IL-17A and IL-5 production by CD4 T-cells. Addition of neutralizing antibodies against Gal-3, or recombinant Gal-3 did not differentially modulate IL-23 p19 versus IL-12 p35. The data indicate that intracellular Gal-3 acts as cytokine hub of human DCs in responding to innate immunity signals. Gal-3 downregulation reprograms proinflammatory cytokine production by MoDCs that inhibit Th2/Th17 development.

  13. Phospholipase Cgamma2 is critical for Dectin-1-mediated Ca2+ flux and cytokine production in dendritic cells.

    PubMed

    Xu, Shengli; Huo, Jianxin; Lee, Koon-Guan; Kurosaki, Tomohiro; Lam, Kong-Peng

    2009-03-13

    Dectin-1 is a C-type lectin that recognizes beta-glucan in the cell walls of fungi and plays an important role in anti-fungal immunity. It signals via tyrosine kinase Syk and adaptor protein Card9 to activate NF-kappaB leading to proinflammatory cytokine production in dendritic cells (DCs). Other than this, not much else is known of the mechanism of Dectin-1 signaling. We demonstrate here that stimulation of DCs with zymosan triggers an intracellular Ca2+ flux that can be attenuated by a blocking anti-Dectin-1 antibody or by pre-treatment of cells with the phospholipase C (PLC) gamma-inhibitor U73122, suggesting that Dectin-1 signals via a PLCgamma pathway to induce Ca2+ flux in DCs. Interestingly, treatment of DCs with particulate curdlan, which specifically engages Dectin-1, results in the phosphorylation of both PLCgamma1 and PLCgamma2. However, we show that PLCgamma2 is the critical enzyme for Dectin-1 signaling in DCs. PLCgamma2-deficient DCs have drastic impairment of Ca2+ signaling and are defective in their secretion of interleukin 2 (IL-2), IL-6, IL-10, IL-12, IL-23, and tumor necrosis factor alpha. PLCgamma2-deficient DCs also exhibit impaired activation of ERK and JNK MAPKs and AP-1 and NFAT transcription factors in response to Dectin-1 stimulation. In addition, PLCgamma2-deficient DCs are also impaired in their activation of NF-kappaB upon Dectin-1 engagement due to defective assembly of the Card9-Bcl10-Malt1 complex and impaired IKKalpha/beta activation and IkappaBalpha degradation. Thus, our data indicate that pattern recognition receptors such as Dectin-1 could elicit Ca2+ signaling and that PLCgamma2 is a critical player in the Dectin-1 signal transduction pathway.

  14. Epigenetic changes in T-cell and monocyte signatures and production of neurotoxic cytokines in ALS patients.

    PubMed

    Lam, Larry; Chin, Lydia; Halder, Ramesh C; Sagong, Bien; Famenini, Sam; Sayre, James; Montoya, Dennis; Rubbi, Liudmilla; Pellegrini, Matteo; Fiala, Milan

    2016-10-01

    We have investigated transcriptional and epigenetic differences in peripheral blood mononuclear cells (PBMCs) of monozygotic female twins discordant in the diagnosis of amyotrophic lateral sclerosis (ALS). Exploring DNA methylation differences by reduced representation bisulfite sequencing (RRBS), we determined that, over time, the ALS twin developed higher abundances of the CD14 macrophages and lower abundances of T cells compared to the non-ALS twin. Higher macrophage signature in the ALS twin was also shown by RNA sequencing (RNA-seq). Moreover, the twins differed in the methylome at loci near several genes, including EGFR and TNFRSF11A, and in the pathways related to the tretinoin and H3K27me3 markers. We also tested cytokine production by PBMCs. The ALS twin's PBMCs spontaneously produced IL-6 and TNF-α, whereas PBMCs of the healthy twin produced these cytokines only when stimulated by superoxide dismutase (SOD)-1. These results and flow cytometric detection of CD45 and CD127 suggest the presence of memory T cells in both twins, but effector T cells only in the ALS twin. The ALS twin's PBMC supernatants, but not the healthy twin's, were toxic to rat cortical neurons, and this toxicity was strongly inhibited by an IL-6 receptor antibody (tocilizumab) and less well by TNF-α and IL-1β antibodies. The putative neurotoxicity of IL-6 and TNF-α is in agreement with a high expression of these cytokines on infiltrating macrophages in the ALS spinal cord. We hypothesize that higher macrophage abundance and increased neurotoxic cytokines have a fundamental role in the phenotype and treatment of certain individuals with ALS.-Lam, L., Chin, L., Halder, R. C., Sagong, B., Famenini, S., Sayre, J., Montoya, D., Rubbi L., Pellegrini, M., Fiala, M. Epigenetic changes in T-cell and monocyte signatures and production of neurotoxic cytokines in ALS patients.

  15. Early dynamics of T helper cell cytokines and T regulatory cells in response to treatment of active Mycobacterium tuberculosis infection.

    PubMed

    Feruglio, S L; Tonby, K; Kvale, D; Dyrhol-Riise, A M

    2015-03-01

    Biomarkers that can identify tuberculosis (TB) disease and serve as markers for efficient therapy are requested. We have studied T cell cytokine production [interferon (IFN)-γ, interleukin (IL)-2, tumour necrosis factor (TNF)-α] and degranulation (CD107a) as well as subsets of CD4(+) T regulatory cells (Tregs ) after in-vitro Mycobacterium tuberculosis (Mtb) antigen stimulation [early secretory antigenic target (ESAT)-6, culture filtrate protein (CFP)-10, antigen 85 (Ag85)] in 32 patients with active tuberculosis (TB) disease throughout 24 weeks of effective TB treatment. A significant decline in the fraction of Mtb-specific total IFN-γ and single IFN-γ-producing T cells was already observed after 2 weeks of treatment, whereas the pool of single IL-2(+) cells increased over time for both CD4(+) and CD8(+) T cells. The Treg subsets CD25(high) CD127(low) , CD25(high) CD147(++) and CD25(high) CD127(low) CD161(+) expanded significantly after Mtb antigen stimulation in vitro at all time-points, whereas the CD25(high) CD127(low) CD39(+) Tregs remained unchanged. The fraction of CD25(high) CD127(low) Tregs increased after 8 weeks of treatment. Thus, we revealed an opposing shift of Tregs and intracellular cytokine production during treatment. This may indicate that functional signatures of the CD4(+) and CD8(+) T cells can serve as immunological correlates of early curative host responses. Whether such signatures can be used as biomarkers in monitoring and follow-up of TB treatment needs to be explored further.

  16. Human cytomegalovirus UL7, a homologue of the SLAM-family receptor CD229, impairs cytokine production.

    PubMed

    Engel, Pablo; Pérez-Carmona, Natàlia; Albà, M Mar; Robertson, Kevin; Ghazal, Peter; Angulo, Ana

    2011-10-01

    Human cytomegalovirus (HCMV), the β-herpesvirus prototype, has evolved a wide spectrum of mechanisms to counteract host immunity. Among them, HCMV uses cellular captured genes encoding molecules capable of interfering with the original host function or of fulfilling new immunomodulatory tasks. Here, we report on UL7, a novel HCMV heavily glycosylated transmembrane protein, containing an Ig-like domain that exhibits remarkable amino acid similarity to CD229, a cell-surface molecule of the signalling lymphocyte-activation molecule (SLAM) family involved in leukocyte activation. The UL7 Ig-like domain, which is well-preserved in all HCMV strains, structurally resembles the SLAM-family N-terminal Ig-variable domain responsible for the homophilic and heterophilic interactions that trigger signalling. UL7 is transcribed with early-late kinetics during the lytic infectious cycle. Using a mAb generated against the viral protein, we show that it is constitutively shed, through its mucine-like stalk, from the cell-surface. Production of soluble UL7 is enhanced by PMA and reduced by a broad-spectrum metalloproteinase inhibitor. Although UL7 does not hold the ability to interact with CD229 or other SLAM-family members, it shares with them the capacity to mediate adhesion to leukocytes, specifically to monocyte-derived DCs. Furthermore, we demonstrate that UL7 expression attenuates the production of proinflammatory cytokines TNF, IL-8 and IL-6 in DCs and myeloid cell lines. Thus, the ability of UL7 to interfere with cellular proinflammatory responses may contribute to viral persistence. These results enhance our understanding of those HCMV-encoded molecules involved in sustaining the balance between HCMV and the host immune system.

  17. Inhibitory effects of compounds from Phyllanthus amarus on nitric oxide production, lymphocyte proliferation, and cytokine release from phagocytes

    PubMed Central

    Yuandani; Jantan, Ibrahim; Ilangkovan, Menaga; Husain, Khairana; Chan, Kok Meng

    2016-01-01

    Standardized extract of Phyllanthus amarus has previously been shown to have a strong inhibitory effect on phagocytic activity of human neutrophils. The current study was carried out to evaluate the effects of constituents of the extract of P. amarus on nitric oxide (NO) production as well as lymphocyte proliferation and cytokine release from phagocytes. Three compounds, ethyl 8-hydroxy-8-methyl-tridecanoate, 7β,19α dihydroxy-urs-12-ene, and 1,7,8-trihydroxy-2-naphtaldehyde, together with seven known compounds were isolated from the whole plant of P. amarus. The isolated compounds and reference standards, ie, gallic acid, ellagic acid, corilagin, and geraniin, which were quantitatively analyzed in the extracts, were evaluated for their effects on immune cells. Among the compounds tested, the lignans, especially phyltetralin and phyllanthin, showed strong inhibition on lymphocyte proliferation with half maximal inhibitory concentration (IC50) values of 1.07 μM and 1.82 μM, respectively. Ethyl 8-hydroxy-8-methyl-tridecanoate and 1,7,8-trihydroxy-2-naphtaldehyde exhibited strong inhibition on nitric oxide production with IC50 values of 0.91 μM and 1.07 μM, respectively. Of all the compounds, corilagin was the strongest inhibitor of tumor necrosis factor-α release with an IC50 value of 7.39 μM, whereas geraniin depicted the strongest inhibitory activity on interleukin-1β release with an IC50 value of 16.41 μM. The compounds constituting the extract of P. amarus were able to inhibit the innate immune response of phagocytes at different steps. PMID:27354767

  18. Muscular hypertrophy and changes in cytokine production after eccentric training in the rat skeletal muscle.

    PubMed

    Ochi, Eisuke; Nakazato, Koichi; Ishii, Naokata

    2011-08-01

    We investigated the time course effects of eccentric training on muscular size, strength, and growth factor/cytokine production by using an isokinetic-exercise system for rats. Male Wistar rats (n = 34) were randomly assigned into 4 groups: 5 session eccentric-training group (ECC5S, n = 10); 5 session sham-operated group (CON5S, n = 10); 10 session eccentric-training group (ECC10S, n = 7); 10 session sham-operated group (CON10S, n = 7). In each group, a session of either training or sham operation was performed every 2 days. The training consisted of 4 sets of forced dorsiflexion (5 repetitions) combined with electric stimulation of plantar flexors. The wet weight of medial gastrocnemius muscle did not increase significantly after 5 sessions of training, whereas that after 10 sessions of training significantly increased with a concomitant increase in the cross-sectional area (CSA) of muscle fibers (weight, p < 0.05; fiber CSA, p < 0.001). Interleukin (IL)-6 in ECC5S and ECC10S groups showed significant increases (p < 0.01), whereas those of tumor necrosis factor (TNF)-α and IL-10 did not. The phospho-stat-3 showed a significant increase in ECC10S (p < 0.001) but not in ECC5S. Myostatin and follistatin also showed significant differences only between ECC10S and CON10S (p < 0.05). The results showed that repeated sessions of eccentric training for 20 days cause increases in muscular size and strength associated with increases in IL-6, follistatin, phospho-stat-3, and a decrease in myostatin. The delayed responses of IL-6, myostatin, phospho-stat-3, and follistatin would be due to the chronic effects of repeated training and possibly important for muscular hypertrophy.

  19. Impact of lithium alone and in combination with antidepressants on cytokine production in vitro.

    PubMed

    Petersein, Charlotte; Sack, Ulrich; Mergl, Roland; Schönherr, Jeremias; Schmidt, Frank M; Lichtblau, Nicole; Kirkby, Kenneth C; Bauer, Katrin; Himmerich, Hubertus

    2015-01-01

    Lithium is an important psychopharmacological agent for the treatment of unipolar as well as bipolar affective disorders. Lithium has a number of side effects such as hypothyroidism and aggravation of psoriasis. On the other hand, lithium has pro-inflammatory effects, which appear beneficial in some disorders associated with immunological deficits, such as human immunodeficiency virus (HIV) infection and systemic lupus erythematosus (SLE). Therefore, immunological characteristics of lithium may be an important consideration in individualized therapeutic decisions. We measured the levels of the cytokines interleukin (IL)-1ß, IL-2, IL-4, IL-6, IL-22, IL-17 and tumour necrosis factor (TNF)-α in the stimulated blood of thirty healthy subjects supplemented with lithium alone, the antidepressants citalopram, escitalopram or mirtazapine alone, the combination of each antidepressant with lithium, and a no drug control. These drugs were tested under three blood stimulant conditions: murine anti-human CD3 monoclonal antibody OKT3 and the 5C3 monoclonal antibody (OKT3/5C3), phytohemagglutinin (PHA), and unstimulated blood. Lithium, alone and in combination with any of the tested antidepressants, led to a consistent increase of IL-1ß, IL-6 and TNF-α levels in the unstimulated as well as the stimulated blood. In the OKT3/5C3- and PHA-stimulated blood, IL-17 production was significantly enhanced by lithium. Lithium additionally increased IL-2 concentrations significantly in PHA-stimulated blood. The data support the view that lithium has pro-inflammatory properties. These immunological characteristics may contribute to side effects of lithium, but may also explain its beneficial effects in patients suffering from HIV infection or SLE.

  20. The adaptor protein SAP regulates type II NKT-cell development, cytokine production, and cytotoxicity against lymphoma.

    PubMed

    Weng, Xiufang; Liao, Chia-Min; Bagchi, Sreya; Cardell, Susanna L; Stein, Paul L; Wang, Chyung-Ru

    2014-12-01

    CD1d-restricted NKT cells represent a unique lineage of immunoregulatory T cells that are divided into two groups, type I and type II, based on their TCR usage. Because there are no specific tools to identify type II NKT cells, little is known about their developmental requirements and functional regulation. In our previous study, we showed that signaling lymphocytic activation molecule associated protein (SAP) is essential for the development of type II NKT cells. Here, using a type II NKT-cell TCR transgenic mouse model, we demonstrated that CD1d-expressing hematopoietic cells, but not thymic epithelial cells, meditate efficient selection of type II NKT cells. Furthermore, we showed that SAP regulates type II NKT-cell development by controlling early growth response 2 protein and promyelocytic leukemia zinc finger expression. SAP-deficient 24αβ transgenic T cells (24αβ T cells) exhibited an immature phenotype with reduced Th2 cytokine-producing capacity and diminished cytotoxicity to CD1d-expressing lymphoma cells. The impaired IL-4 production by SAP-deficient 24αβ T cells was associated with reduced IFN regulatory factor 4 and GATA-3 induction following TCR stimulation. Collectively, these data suggest that SAP is critical for regulating type II NKT cell responses. Aberrant responses of these T cells may contribute to the immune dysregulation observed in X-linked lymphoproliferative disease caused by mutations in SAP.

  1. Effect of Th1/Th2 cytokine administration on proinflammatory SKOV-3 cell activation

    PubMed Central

    Sikora, Justyna; Kondera-Anasz, Zdzisława; Mickiewicz, Patrycja; Mickiewicz, Adam

    2015-01-01

    Introduction Interleukin(IL)-1β, IL-6 and IL-12 might associate with inflammatory processes in a tumor progression and create a specific microenvironment for tumor growth. The aim of the study was to assess whether the Th1 and Th2 type cytokines, such as IL-2 and IL-10, affect ovarian carcinoma continuous cell line (SKOV-3) pro-inflammatory activation. Material and methods SKOV-3 ovarian cells and peripheral blood mononuclear cells (PBMCs) were stimulated by IL-2 and IL-10. Additionally, SKOV-3 ovarian cells and PBMCs were co-cultured together. Proinflammatory activation of cancer cells was evaluated by measurement of IL-1β and IL-6 levels in culture fluid after 72 h of incubation. Results SKOV-3 cells and PBMCs secreted IL-1β and IL-6. After stimulation by IL-2 and IL-10, secretion of studied parameters was changed in a dose-dependent manner. The addition of a higher IL-2 level gave rise to an increase of IL-1β, IL-6 and IL-12 secretion in SKOV-3 cells. Stimulation by IL-10 increased only IL-1β secretion in SKOV-3 cells. However, IL-6 secretion decreased after stimulation with 25 ng/ml IL-10. Activatory effects of IL-2 and inhibitory effects of IL-10 in co-culture of SKOV-3 and PBMCs were observed. Conclusions Our results suggested that Th1/Th2 type of cytokines might influence pro-inflammatory activation of SKOV-3 ovarian cells. Co-cultures of SKOV-3 and PBMCs showed significant changes in cross-talk between cancer and immune cells. PMID:27904527

  2. Insights into cytokine-receptor interactions from cytokine engineering.

    PubMed

    Spangler, Jamie B; Moraga, Ignacio; Mendoza, Juan L; Garcia, K Christopher

    2015-01-01

    Cytokines exert a vast array of immunoregulatory actions critical to human biology and disease. However, the desired immunotherapeutic effects of native cytokines are often mitigated by toxicity or lack of efficacy, either of which results from cytokine receptor pleiotropy and/or undesired activation of off-target cells. As our understanding of the structural principles of cytokine-receptor interactions has advanced, mechanism-based manipulation of cytokine signaling through protein engineering has become an increasingly feasible and powerful approach. Modified cytokines, both agonists and antagonists, have been engineered with narrowed target cell specificities, and they have also yielded important mechanistic insights into cytokine biology and signaling. Here we review the theory and practice of cytokine engineering and rationalize the mechanisms of several engineered cytokines in the context of structure. We discuss specific examples of how structure-based cytokine engineering has opened new opportunities for cytokines as drugs, with a focus on the immunotherapeutic cytokines interferon, interleukin-2, and interleukin-4.

  3. Immunomodulatory activity of aqueous extract of Nyctanthes arbor-tristis flowers with particular reference to splenocytes proliferation and cytokines induction

    PubMed Central

    Bharshiv, Chandrabhan Kumar; Garg, Satish Kumar; Bhatia, A. K.

    2016-01-01

    Objectives: To investigate the immunomodulatory activity of aqueous extract of Nyctanthes arbor-tristis flowers (NAFE) with particular reference to splenocytes proliferation and induction of cytokines. Materials and Methods: Antibody titer was determined by tube agglutination and indirect ELISA assay in four groups of mice-control, antigen alone, and NAFE-treated (400 and 800 mg/kg for 21 days) after immunization with Salmonella antigen while cellular immunity was studied in three groups of rats (control and NAFE-treated - 400 and 800 mg/kg) following DNCB application. Splenocytes from untreated and NAFE-treated rats were stimulated using concanavalin-A (Con-A) and optical density (OD) and stimulation index were determined. Splenocytes from control rats were also treated in vitro with NAFE (50–1600 μg/ml) and Con-A to determine the effect on splenocytes proliferation. Interleukin-2 (IL-2) and IL-6 levels in splenocytes supernatant from control and NAFE-treated rats and following in vitro treatment of splenocytes with NAFE (50–1600 μg/ml) were determined using ELISA kits. Results: Marked to a significant increase in antibody titer by both the methods in NAFE-treated mice and a significant increase in skin thickness in rats after challenge with DNCB, respectively suggested humoral and cell-mediated immunostimulant potential of NAFE. Significant increase in OD and stimulation index following e x vivo and in vitro exposure of splenocytes and sensitization with Con-A and significant elevation in IL-2 and IL-6 levels in splenocytes supernantant was also observed after their ex vivo and in vitro exposure to NAFE. Conclusion: Humoral and cell-mediated immunostimulant activity of NAFE seems to be mediated through splenocytes proliferation and increased production of cytokines, especially IL-2 and IL-6. PMID:27756953

  4. Triptolide Attenuates Endotoxin- and Staphylococcal Exotoxin-Induced T-Cell Proliferation and Production of Cytokines and Chemokines

    DTIC Science & Technology

    2005-02-01

    human peripheral blood mononuclear cells (PBMC). It also blocked the production of these cytokines and chemokines by LPS- stimulated PBMC in a dose...SE- stimulated T- cell proliferation (by 98%) and expression of interleukin 1b, interleukin 6, tumor necrosis factor , gamma interferon, monocyte...chemotactic protein 1, macrophage inflammatory protein (MIP)-1a, and MIP-1b by human peripheral blood mononuclear cells (PBMC). It also blocked the

  5. Regulation of cytokine production by soluble CD23: costimulation of interferon gamma secretion and triggering of tumor necrosis factor alpha release

    PubMed Central

    1994-01-01

    Soluble CD23 (sCD23) has multiple IgE-independent biological activities. In the present study, we examined the regulatory effect of sCD23 on cytokine production by human peripheral blood mononuclear cells (PBMC). We show that sCD23 enhances by about 80-fold the interleukin 2 (IL-2)-induced interferon gamma (IFN-gamma) production and by about 10-fold the response to IL-12. This potentiating activity is time and dose dependent and is not associated with a significant effect on DNA synthesis. The sCD23 costimulatory activity for IFN-gamma synthesis is drastically reduced in monocyte-depleted PBMC, suggesting that monocytes may be the target for sCD23. This hypothesis was supported by the following observations. First, sCD23 alone is a potent inducer of tumor necrosis factor alpha (TNF-alpha) production by PBMC and this effect disappears after monocyte depletion. The triggering of TNF-alpha release is specifically inhibited by neutralizing anti-CD23 monoclonal antibody (mAb). In addition, IL-2 and IL-12 synergize with sCD23 to induce TNF-alpha production. Second, sCD23 triggers the release of other inflammatory mediators such as IL-1 alpha, IL-1 beta, and IL-6. Finally, TNF-alpha production in response to IL-2 and sCD23 precedes IFN-gamma and IFN-gamma secretion is significantly inhibited by anti-TNF-alpha mAb, indicating that the sCD23 costimulatory signal for IFN-gamma production may be partially mediated by TNF-alpha release. It is proposed that sCD23 is a proinflammatory cytokine that, in addition, may play an important role in the control of the immune response via the enhancement of IFN-gamma production. PMID:8064221

  6. Regulation of NK Cell Activation and Effector Functions by the IL-12 Family of Cytokines: The Case of IL-27.

    PubMed

    Zwirner, Norberto Walter; Ziblat, Andrea

    2017-01-01

    Natural killer (NK) cells are characterized by their ability to detect and induce apoptosis of susceptible target cells and by secretion of immunoregulatory cytokines such as IFN-γ. Activation of these effector functions is triggered upon recognition of tumor and pathogen (mostly virus)-infected cells and because of a bidirectional cross talk that NK cells establish with other cells of myeloid origin such as dendritic cells (DC) and macrophages. A common characteristic of these myeloid cells is their ability to secrete different members of the IL-12 family of cytokines such as IL-12, IL-23, and IL-27 and cytokines such as IL-15 and IL-18. Although the effect of IL-12, IL-15, and IL-18 has been characterized, the effect of IL-23 and IL-27 on NK cells (especially human) remains ill-defined. Particularly, IL-27 is a cytokine with dual functions as it has been described as pro- and as anti-inflammatory in different experimental settings. Recent evidence indicates that this cytokine indeed promotes human NK cell activation, IFN-γ secretion, NKp46-dependent NK cell-mediated cytotoxicity, and antibody (Ab)-dependent NK cell-mediated cytotoxicity (ADCC) against monoclonal Ab-coated tumor cells. Remarkably, IL-27 also primes NK cells for IL-18 responsiveness, enhancing these functional responses. Consequently, IL-27 acts as a pro-inflammatory cytokine that, in concert with other DC-derived cytokines, hierarchically contributes to NK cells activation and effector functions, which likely contributes to foster the adaptive immune response in different physiopathological conditions.

  7. Regulation of NK Cell Activation and Effector Functions by the IL-12 Family of Cytokines: The Case of IL-27

    PubMed Central

    Zwirner, Norberto Walter; Ziblat, Andrea

    2017-01-01

    Natural killer (NK) cells are characterized by their ability to detect and induce apoptosis of susceptible target cells and by secretion of immunoregulatory cytokines such as IFN-γ. Activation of these effector functions is triggered upon recognition of tumor and pathogen (mostly virus)-infected cells and because of a bidirectional cross talk that NK cells establish with other cells of myeloid origin such as dendritic cells (DC) and macrophages. A common characteristic of these myeloid cells is their ability to secrete different members of the IL-12 family of cytokines such as IL-12, IL-23, and IL-27 and cytokines such as IL-15 and IL-18. Although the effect of IL-12, IL-15, and IL-18 has been characterized, the effect of IL-23 and IL-27 on NK cells (especially human) remains ill-defined. Particularly, IL-27 is a cytokine with dual functions as it has been described as pro- and as anti-inflammatory in different experimental settings. Recent evidence indicates that this cytokine indeed promotes human NK cell activation, IFN-γ secretion, NKp46-dependent NK cell-mediated cytotoxicity, and antibody (Ab)-dependent NK cell-mediated cytotoxicity (ADCC) against monoclonal Ab-coated tumor cells. Remarkably, IL-27 also primes NK cells for IL-18 responsiveness, enhancing these functional responses. Consequently, IL-27 acts as a pro-inflammatory cytokine that, in concert with other DC-derived cytokines, hierarchically contributes to NK cells activation and effector functions, which likely contributes to foster the adaptive immune response in different physiopathological conditions. PMID:28154569

  8. Antigen-specific CD4{sup +} effector T cells: Analysis of factors regulating clonal expansion and cytokine production

    SciTech Connect

    Ohnuki, Kazunobu; Watanabe, Yuri; Takahashi, Yusuke; Kobayashi, Sakiko; Watanabe, Shiho; Ogawa, Shuhei; Kotani, Motoko; Kozono, Haruo; Tanabe, Kazunari; Abe, Ryo

    2009-03-20

    In order to fully understand T cell-mediated immunity, the mechanisms that regulate clonal expansion and cytokine production by CD4{sup +} antigen-specific effector T cells in response to a wide range of antigenic stimulation needs clarification. For this purpose, panels of antigen-specific CD4{sup +} T cell clones with different thresholds for antigen-induced proliferation were generated by repeated stimulation with high- or low-dose antigen. Differences in antigen sensitivities did not correlate with expression of TCR, CD4, adhesion or costimulatory molecules. There was no significant difference in antigen-dependent cytokine production by TG40 cells transfected with TCR obtained from either high- or low-dose-responding T cell clones, suggesting that the affinity of TCRs for their ligands is not primary determinant of T cell antigen reactivity. The proliferative responses of all T cell clones to both peptide stimulation and to TCR{beta} crosslinking revealed parallel dose-response curves. These results suggest that the TCR signal strength of effector T cells and threshold of antigen reactivity is determined by an intrinsic property, such as the TCR signalosome and/or intracellular signaling machinery. Finally, the antigen responses of high- and low-peptide-responding T cell clones reveal that clonal expansion and cytokine production of effector T cells occur independently of antigen concentration. Based on these results, the mechanisms underlying selection of high 'avidity' effector and memory T cells in response to pathogen are discussed.

  9. Proteolytic activity and cytokine up-regulation by non-albicans Candida albicans.

    PubMed

    Nawaz, Ali; Pärnänen, Pirjo; Kari, Kirsti; Meurman, Jukka H

    2015-05-01

    Mouth is an important source of infections and oral infections such as Candida infections increase the risk of mortality. Our purpose was to investigate differences in proteolytic activity of non-albicans Candida albicans (non-albicans Candida) between clinical isolates and laboratory samples. The second aim was to assess the concentration of pro- and anti-inflammatory cytokine levels IL-1β, IL-10, and TNF-α in saliva of patients with the non-albicans Candida and Candida-negative saliva samples. Clinical yeast samples from our laboratory were used for analyses. Candida strains were grown in YPG at 37 °C for 24 h in water bath with shaking. The activity of Candida proteinases of cell and cell-free fractions were analyzed by MDPF-gelatin zymography. The levels of IL-1β, IL-10, and TNF-α were measured from saliva with ELISA. The study showed differences in the proteolytic activity among the non-albicans Candida strains. C. tropicalis had higher proteolytic activity when compared to the other strains. Significant difference was found in salivary IL-1β levels between the non-albicans Candida and control strains (P < 0.002). The present findings showed differences in proteolytic activity among the non-albicans Candida strains. The increased IL-1β concentration may be one of the host response components associated with non-albicans Candida infection.

  10. Cholesterol Crystals Induce Inflammatory Cytokines Expression in nARPE-19 Cells by Activating the NF-κB Pathway

    PubMed Central

    Hu, Yijun; Lin, Haijiang; Dib, Bernard; Atik, Alp; Bouzika, Peggy; Lin, Christopher; Yan, Yueran; Tang, Shibo; Miller, Joan W.; Vavvas, Demetrios G.

    2015-01-01

    Purpose To investigate the expression of inflammatory cytokines in ARPE-19 cells after stimulation with cholesterol crystals. Methods APRE-19 cells were cultured, primed with IL-1α, and treated with cholesterol crystals under different concentrations. Inflammatory cytokines (mature-IL-1β, IL-6, and IL-8) in supernatant and inflammatory cytokines (pro-IL-1β, IL-18) in cell lysate were detected by western blot. The NF-κB pathway inhibitor BAY 11-7082 was used to determine the pathway of cytokine expression. Results Cholesterol crystals did not induce the nucleotide-binding domain leucine-rich repeat containing family, pyrin domain containing 3 (NLRP3) inflammasome, but did increase pro-IL-1β expression in ARPE-19 cells. Cholesterol crystals increased pro-IL-1β expression by activating the NF-κB pathway. Cholesterol crystal activation of the NF-κB pathway also leads to increased IL-6 and IL-8 expression. Conclusion Cholesterol crystals can induce inflammatory cytokine expression in ARPE-19 cells by activating the NF-κB pathway. PMID:25091484

  11. Immunization with Protein D from Non-Typeable Haemophilus influenzae (NTHi) Induced Cytokine Responses and Bioactive Antibody Production

    PubMed Central

    Davoudi Vijeh Motlagh, Atefeh; Siadat, Seyed Davar; Abedian Kenari, Saeid; Mahdavi, Mehdi; Behrouzi, Ava; Asgarian-Omran, Hossein

    2016-01-01

    Background Outer membrane protein D (PD) is a highly conserved and stable protein in the outer membrane of both encapsulated (typeable) and non-capsulated (non-typeable) strains of Haemophilus influenzae. As an immunogen, PD is a potential candidate vaccine against non-typeable H. influenzae (NTHi) strains. Objectives The aim of this study was to determine the cytokine pattern and the opsonic antibody response in a BALB/c mouse model versus PD from NTHi as a vaccine candidate. Methods Protein D was formulated with Freund’s and outer membrane vesicle (OMV) adjuvants and injected into experimental mice. Sera from all groups were collected. The bioactivity of the anti-PD antibody was determined by opsonophagocytic killing test. To evaluate the cytokine responses, the spleens were assembled, suspension of splenocytes was recalled with antigen, and culture supernatants were analyzed by ELISA for IL-4, IL-10, and IFN-γ cytokines. Results Anti-PD antibodies promoted phagocytosis of NTHi in both immunized mice groups (those administered PD + Freund’s and those administered PD + OMV adjuvants, 92.8% and 83.5%, respectively, compared to the control group). In addition, the concentrations of three cytokines were increased markedly in immunized mice. Conclusions We conclude that immunization with PD protects mice against NTHi. It is associated with improvements in both cellular and humoral immune responses and opsonic antibody activity. PMID:27942362

  12. Neutralizing antibodies inhibit HIV-1 infection of plasmacytoid dendritic cells by an FcγRIIa independent mechanism and do not diminish cytokines production.

    PubMed

    Lederle, Alexandre; Su, Bin; Holl, Vincent; Penichon, Julien; Schmidt, Sylvie; Decoville, Thomas; Laumond, Géraldine; Moog, Christiane

    2014-08-18

    Plasmacytoid dendritic cells (pDC) expressing FcγRIIa are antigen-presenting cells able to link innate and adaptive immunity and producing various cytokines and chemokines. Although highly restricted, they are able to replicate HIV-1. We determined the activity of anti-HIV-1 neutralizing antibodies (NAb) and non-neutralizing inhibitory antibodies (NNIAb) on the infection of primary pDC by HIV-1 primary isolates and analyzed cytokines and chemokines production. Neutralization assay was performed with primary pDC in the presence of serial antibodies (Ab) concentrations. In parallel, we measured the release of cytokines and chemokines by ELISA and CBA Flex assay. We found that NAb, but not NNIAb, inhibit HIV-1 replication in pDC. This inhibitory activity was lower than that detected for myeloid dendritic cells (mDC) infection and independent of FcγRIIa expressed on pDC. Despite the complete protection, IFN-α production was detected in the supernatant of pDC treated with NAb VRC01, 4E10, PGT121, 10-1074, 10E8, or polyclonal IgG44 but not with NAb b12. Production of MIP-1α, MIP-1β, IL-6, and TNF-α by pDC was also maintained in the presence of 4E10, b12 and VRC01. These findings suggest that pDC can be protected from HIV-1 infection by both NAb and IFN-α release triggered by the innate immune response during infection.

  13. Nicotine inhibits Fc epsilon RI-induced cysteinyl leukotrienes and cytokine production without affecting mast cell degranulation through alpha 7/alpha 9/alpha 10-nicotinic receptors.

    PubMed

    Mishra, Neerad C; Rir-sima-ah, Jules; Boyd, R Thomas; Singh, Shashi P; Gundavarapu, Sravanthi; Langley, Raymond J; Razani-Boroujerdi, Seddigheh; Sopori, Mohan L

    2010-07-01

    Smokers are less likely to develop some inflammatory and allergic diseases. In Brown-Norway rats, nicotine inhibits several parameters of allergic asthma, including the production of Th2 cytokines and the cysteinyl leukotriene LTC(4). Cysteinyl leukotrienes are primarily produced by mast cells, and these cells play a central role in allergic asthma. Mast cells express a high-affinity receptor for IgE (FcepsilonRI). Following its cross-linking, cells degranulate and release preformed inflammatory mediators (early phase) and synthesize and secrete cytokines/chemokines and leukotrienes (late phase). The mechanism by which nicotine modulates mast cell activation is unclear. Using alpha-bungarotoxin binding and quantitative PCR and PCR product sequencing, we showed that the rat mast/basophil cell line RBL-2H3 expresses nicotinic acetylcholine receptors (nAChRs) alpha7, alpha9, and alpha10; exposure to exceedingly low concentrations of nicotine (nanomolar), but not the biologically inactive metabolite cotinine, for > or = 8 h suppressed the late phase (leukotriene/cytokine production) but not degranulation (histamine and hexosaminidase release). These effects were unrelated to those of nicotine on intracellular free calcium concentration but were causally associated with the inhibition of cytosolic phospholipase A(2) activity and the PI3K/ERK/NF-kappaB pathway, including phosphorylation of Akt and ERK and nuclear translocation of NF-kappaB. The suppressive effect of nicotine on the late-phase response was blocked by the alpha7/alpha9-nAChR antagonists methyllycaconitine and alpha-bungarotoxin, as well as by small interfering RNA knockdown of alpha7-, alpha9-, or alpha10-nAChRs, suggesting a functional interaction between alpha7-, alpha9-, and alpha10-nAChRs that might explain the response of RBL cells to nanomolar concentrations of nicotine. This "hybrid" receptor might serve as a target for novel antiallergic/antiasthmatic therapies.

  14. Interleukin-32 production associated with biliary innate immunity and proinflammatory cytokines contributes to the pathogenesis of cholangitis in biliary atresia.

    PubMed

    Okamura, A; Harada, K; Nio, M; Nakanuma, Y

    2013-08-01

    Biliary atresia (BA) is thought to be associated with infections by viruses such as Reoviridae and is characterized histologically by fibrosclerosing cholangitis with proinflammatory cytokine-mediated inflammation. Interleukin (IL)-32 affects the continuous inflammation by increasing the production of proinflammatory cytokines. In this study, the role of IL-32 in the cholangitis of BA was examined. Immunohistochemistry for IL-32 and caspase 1 was performed using 21 samples of extrahepatic bile ducts resected from BA patients. Moreover, using cultured human biliary epithelial cells (BECs), the expression of IL-32 and its induction on stimulation with a Toll-like receptor [(TLR)-3 ligand (poly(I:C)] and proinflammatory cytokines was examined. BECs composing extrahepatic bile ducts showing cholangitis expressed IL-32 in BA, but not in controls. Caspase 1 was expressed constantly on BECs of both BA and control subjects. Furthermore, poly(I:C) and proinflammatory cytokines [(IL-1β, interferon (IFN)-γ and tumour necrosis factor (TNF)-α] induced IL-32 expression strongly in cultured BECs, accompanying the constant expression of TLR-3 and caspase 1. Our results imply that the expression of IL-32 in BECs was found in the damaged bile ducts of BA and induced by biliary innate immunity via TLR-3 and proinflammatory cytokines. These findings suggest that IL-32 is involved initially in the pathogenic mechanisms of cholangitis in BA and also plays an important role in the amplification and continuance of periductal inflammatory reactions. It is therefore tempting to speculate that inhibitors of IL-32 could be useful for attenuating cholangitis in BA.

  15. Dimethyl fumarate blocks pro-inflammatory cytokine production via inhibition of TLR induced M1 and K63 ubiquitin chain formation

    PubMed Central

    McGuire, Victoria A.; Ruiz-Zorrilla Diez, Tamara; Emmerich, Christoph H.; Strickson, Sam; Ritorto, Maria Stella; Sutavani, Ruhcha V.; Weiβ, Anne; Houslay, Kirsty F.; Knebel, Axel; Meakin, Paul J.; Phair, Iain R.; Ashford, Michael L. J.; Trost, Matthias; Arthur, J. Simon C.

    2016-01-01

    Dimethyl fumarate (DMF) possesses anti-inflammatory properties and is approved for the treatment of psoriasis and multiple sclerosis. While clinically effective, its molecular target has remained elusive - although it is known to activate anti-oxidant pathways. We find that DMF inhibits pro-inflammatory cytokine production in response to TLR agonists independently of the Nrf2-Keap1 anti-oxidant pathway. Instead we show that DMF can inhibit the E2 conjugating enzymes involved in K63 and M1 polyubiquitin chain formation both in vitro and in cells. The formation of K63 and M1 chains is required to link TLR activation to downstream signaling, and consistent with the block in K63 and/or M1 chain formation, DMF inhibits NFκB and ERK1/2 activation, resulting in a loss of pro-inflammatory cytokine production. Together these results reveal a new molecular target for DMF and show that a clinically approved drug inhibits M1 and K63 chain formation in TLR induced signaling complexes. Selective targeting of E2s may therefore be a viable strategy for autoimmunity. PMID:27498693

  16. Isoquercitrin suppresses the expression of histamine and pro-inflammatory cytokines by inhibiting the activation of MAP Kinases and NF-κB in human KU812 cells.

    PubMed

    Li, Li; Zhang, Xiao-Hui; Liu, Guang-Rong; Liu, Chang; Dong, Yin-Mao

    2016-06-01

    Mast cells and basophils are multifunctional effector cells that contain abundant secretory granules in their cytoplasm. Both cell types are involved in a variety of inflammatory and immune events, producing an array of inflammatory mediators, such as cytokines. The aim of the study was to examine whether isoquercitrin modulates allergic and inflammatory reactions in the human basophilic KU812 cells and to elucidate its influence on the phosphorylation of mitogen-activated protein kinase (MAPK) and nuclear factor (NF)-κB activation. The KU812 cells were stimulated with phorbol-12-myristate 13-acetate plus the calcium ionophore A23187 (PMACI). The inhibitory effects of isoquercitrin on the productions of histamine and pro-inflammatory cytokines in the stimulated KU812 cells were measured using cytokine-specific enzyme-linked immunosorbent (ELISA) assays. Western blotting analysis was used to assess the effects of isoquercitrin on the MAPKs and NF-κB protein levels. Our results indicated that the isoquercitrin treatment of PMACI-stimulated KU812 cells significantly reduced the production of histamine and the pro-inflammatory cytokines, such as interleukin (IL)-6, IL-8, IL-1β, and tumor necrosis factor (TNF)-α. The treated cells exhibited decreased phosphorylation of extracellular signal-regulated kinase (ERK), revealing the role of ERK MAPK in isoquercitrin-mediated allergy inhibition. Furthermore, isoquercitrin suppressed the PMACI-mediated activation of NF-κB in the human basophil cells. In conclusion, the results from the present study provide insights into the potential therapeutic use of isoquercitrin for the treatment of inflammatory and allergic reactions.

  17. Common γ-chain cytokine signaling is required for macroautophagy induction during CD4+ T-cell activation

    PubMed Central

    Botbol, Yair; Patel, Bindi; Macian, Fernando

    2015-01-01

    Macroautophagy is a cellular process that mediates degradation in the lysosome of cytoplasmic components including proteins and organelles. Previous studies have shown that macroautophagy is induced in activated T cells to regulate organelle homeostasis and the cell's energy metabolism. However, the signaling pathways that initiate and regulate activation-induced macroautophagy in T cells have not been identified. Here, we show that activation-induced macroautophagy in T cells depends on signaling from common γ-chain cytokines. Consequently, inhibition of signaling through JAK3, induced downstream of cytokine receptors containing the common γ-chain, prevents full induction of macroautophagy in activated T cells. Moreover, we found that common γ-chain cytokines are not only required for macroautophagy upregulation during T cell activation but can themselves induce macroautophagy. Our data also show that macroautophagy induction in T cells is associated with an increase of LC3 expression that is mediated by a post-transcriptional mechanism. Overall, our findings unveiled a new role for common γ-chain cytokines as a molecular link between autophagy induction and T-cell activation. PMID:26391567

  18. Effects of Omega-3-Rich Harp Seal Oil on the Production of Pro-Inflammatory Cytokines in Mouse Peritoneal Macrophages.

    PubMed

    Choi, Myungwon; Ju, Jaehyun; Suh, Jae Soo; Park, Kun-Young; Kim, Kwang Hyuk

    2015-06-01

    Omega-3, a polyunsaturated fatty acid, is an essential fatty acid necessary for human health, and it protects against cardiovascular disease, inflammation, autoimmune diseases, and cancer. In the present study, we investigated the effects of omega-3-rich harp seal oil (HSO) on the production of nitric oxide (NO) and cytokines, such as tumor necrosis factor (TNF)-α, interleukin-(IL)-1β, IL-6, and IL-12/IL-23 (p40) in peritoneal macrophages of mice. The culture supernatants of murine macrophages exposed to lipopolysaccharide (LPS), HSO, or HSO+LPS were harvested to assay IL-1β, TNF-α, IL-6, and IL-12/IL-23 (p40) cytokines and NO. TNF-α, IL-1 β, and IL-12/IL-23 (p40) levels, except IL-6, were lower in the culture supernatants of mouse peritoneal macrophages exposed to LPS plus HSO than those of the groups exposed to LPS alone. These observations demonstrate that omega-3-rich harp seal oil downregulates the production of the pro-inflammatory cytokines such as IL-1β, TNF-α, and IL-12/IL-23 (p40). These results suggest that HSO could be potentially used as a preventive agent or as an adjunct in anti-inflammatory therapy, if more research results were accumulated.

  19. SARM is required for neuronal injury and cytokine production in response to central nervous system viral infection.

    PubMed

    Hou, Ying-Ju; Banerjee, Rebecca; Thomas, Bobby; Nathan, Carl; García-Sastre, Adolfo; Ding, Aihao; Uccellini, Melissa B

    2013-07-15

    Four of the five members of the Toll/IL-1R domain-containing adaptor family are required for signaling downstream of TLRs, promoting innate immune responses against different pathogens. However, the role of the fifth member of this family, sterile α and Toll/IL-1R domain-containing 1 (SARM), is unclear. SARM is expressed primarily in the CNS where it is required for axonal death. Studies in Caenorhabditis elegans have also shown a role for SARM in innate immunity. To clarify the role of mammalian SARM in innate immunity, we infected SARM(-/-) mice with a number of bacterial and viral pathogens. SARM(-/-) mice show normal responses to Listeria monocytogenes, Mycobacterium tuberculosis, and influenza virus, but show dramatic protection from death after CNS infection with vesicular stomatitis virus. Protection correlates with reduced CNS injury and cytokine production by nonhematopoietic cells, suggesting that SARM is a positive regulator of cytokine production. Neurons and microglia are the predominant source of cytokines in vivo, supporting a role for SARM as a link between neuronal injury and innate immunity.

  20. Effects of Omega-3-Rich Harp Seal Oil on the Production of Pro-Inflammatory Cytokines in Mouse Peritoneal Macrophages

    PubMed Central

    Choi, Myungwon; Ju, Jaehyun; Suh, Jae Soo; Park, Kun-Young; Kim, Kwang Hyuk

    2015-01-01

    Omega-3, a polyunsaturated fatty acid, is an essential fatty acid necessary for human health, and it protects against cardiovascular disease, inflammation, autoimmune diseases, and cancer. In the present study, we investigated the effects of omega-3-rich harp seal oil (HSO) on the production of nitric oxide (NO) and cytokines, such as tumor necrosis factor (TNF)-α, interleukin-(IL)-1β, IL-6, and IL-12/IL-23 (p40) in peritoneal macrophages of mice. The culture supernatants of murine macrophages exposed to lipopolysaccharide (LPS), HSO, or HSO+LPS were harvested to assay IL-1β, TNF-α, IL-6, and IL-12/IL-23 (p40) cytokines and NO. TNF-α, IL-1 β, and IL-12/IL-23 (p40) levels, except IL-6, were lower in the culture supernatants of mouse peritoneal macrophages exposed to LPS plus HSO than those of the groups exposed to LPS alone. These observations demonstrate that omega-3-rich harp seal oil downregulates the production of the pro-inflammatory cytokines such as IL-1β, TNF-α, and IL-12/IL-23 (p40). These results suggest that HSO could be potentially used as a preventive agent or as an adjunct in anti-inflammatory therapy, if more research results were accumulated. PMID:26175994

  1. Cell cycle progression following naive T cell activation is independent of Jak3/common gamma-chain cytokine signals.

    PubMed

    Shi, Min; Lin, Tsung H; Appell, Kenneth C; Berg, Leslie J

    2009-10-01

    T cell proliferation following activation is an essential aspect of the adaptive immune response. Multiple factors, such as TCR signaling, costimulation, and signals from cytokines, each contribute to determine the magnitude of T cell expansion. In this report, we examine in detail the role of Jak3/common gamma-chain-dependent cytokines in promoting cell cycle progression and proliferation of naive T cells. Using naive CD4+ T cells from Jak3-deficient mice and wild-type CD4+ T cells treated with a small molecule inhibitor of Jak3, we find that these cytokine signals are not required for proliferation; instead, they are important for the survival of activated T cells. In addition, we show that the percentage of cells entering the cell cycle and the percentage of cells in each round of cell division are comparable between Jak3-deficent and wild-type T cells. Furthermore, cell cycle progression and the regulated expression of key cell cycle proteins are independent of Jak3/common gamma-chain cytokine signals. These findings hold true over a wide range of TCR signal strengths. However, when CD28 costimulatory signals, but not TCR signals, are limiting, Jak3-dependent cytokine signals become necessary for the proliferation of naive T cells. Because CD28 signaling has been found to be dispensable for autoreactive T cell responses, these data suggest the potential for interfering with autoimmune T cell responses by inhibition of Jak3 signaling.

  2. Vaccenic acid-mediated reduction in cytokine production is independent of c9,t11-CLA in human peripheral blood mononuclear cells.

    PubMed

    Jaudszus, Anke; Jahreis, Gerhard; Schlörmann, Wiebke; Fischer, Janine; Kramer, Ronny; Degen, Christian; Rohrer, Carsten; Roth, Alexander; Gabriel, Holger; Barz, Dagmar; Gruen, Michael

    2012-10-01

    The ruminant trans fatty acid vaccenic acid (tVA) favorably alters markers of inflammation. However, it is not yet clear whether these effects are attributed to its endogenous partial conversion to c9,t11-CLA, which is known to possess anti-inflammatory properties. We compared the cytokine reducing potential of tVA to c9,t11-CLA in human T-helper (Th) cells as a main source of cytokine production during inflammation. Secondly, we assessed whether a bioconversion of tVA to c9,t11-CLA via stearoyl-CoA desaturase (SCD) encoded activity takes place in peripheral blood mononuclear cells (PBMC) in order to relate the outcomes of intracellular cytokine measurement to the degree of conversion. TVA reduced the percentage of both IL-2 and TNF-α expressing Th cells significantly, but to a lesser extent compared to c9,t11-CLA, as determined by flow cytometry after alloreactive stimulation of PBMC. Pre-treatment with the selective PPARγ antagonist T0070907 largely re-established the IL-2 and TNF-α positive Th cell population in both tVA and c9,t11-CLA treated cultures. Interestingly, while the portion of tVA dose-dependently increased within the cellular lipid fraction, the initially marginal amount of c9,t11-CLA remained unaltered. However, SCD mRNA although abundantly expressed in PBMC was not regulated by tVA. Conclusively, these results suggest that the cytokine reducing effect of tVA in human T cells is independent of c9,t11-CLA, since no bioconversion occurred. Moreover, the data provide evidence that tVA mechanistically acts in a manner similar to c9,t11-CLA.

  3. Glibenclamide reduces pro-inflammatory cytokine production by neutrophils of diabetes patients in response to bacterial infection

    NASA Astrophysics Data System (ADS)

    Kewcharoenwong, Chidchamai; Rinchai, Darawan; Utispan, Kusumawadee; Suwannasaen, Duangchan; Bancroft, Gregory J.; Ato, Manabu; Lertmemongkolchai, Ganjana

    2013-11-01

    Type 2 diabetes mellitus is a major risk factor for melioidosis, which is caused by Burkholderia pseudomallei. Our previous study has shown that polymorphonuclear neutrophils (PMNs) from diabetic subjects exhibited decreased functions in response to B. pseudomallei. Here we investigated the mechanisms regulating cytokine secretion of PMNs from diabetic patients which might contribute to patient susceptibility to bacterial infections. Purified PMNs from diabetic patients who had been treated with glibenclamide (an ATP-sensitive potassium channel blocker for anti-diabetes therapy), showed reduction of interleukin (IL)-1β and IL-8 secretion when exposed to B. pseudomallei. Additionally, reduction of these pro-inflammatory cytokines occurred when PMNs from diabetic patients were treated in vitro with glibenclamide. These findings suggest that glibenclamide might be responsible for the increased susceptibility of diabetic patients, with poor glycemic control, to bacterial infections as a result of its effect on reducing IL-1β production by PMNs.

  4. Capsaicin attenuates LPS-induced inflammatory cytokine production by upregulation of LXRα.

    PubMed

    Tang, Jing; Luo, Kang; Li, Yan; Chen, Quan; Tang, Dan; Wang, Deming; Xiao, Ji

    2015-09-01

    Here, we investigated the role of LXRα in capsaicin mediated anti-inflammatory effects. Results revealed that capsaicin inhibits LPS-induced IL-1β, IL-6 and TNF-α production in a time- and dose-dependent manner. Moreover, capsaicin increases LXRα expression through PPARγ pathway. Inhibition of LXRα activation by siRNA diminished the inhibitory action of capsaicin on LPS-induced IL-1β, IL-6 and TNF-α production. Additionally, LXRα siRNA abrogated the inhibitory action of capsaicin on p65 NF-κB protein expression. Thus, we propose that the anti-inflammatory effects of capsaicin are LXRα dependent, and LXRα may potentially link the capsaicin mediated PPARγ activation and NF-κB inhibition in LPS-induced inflammatory response.

  5. Cytokine-mediated induction of endothelial adhesion molecule and histocompatibility leukocyte antigen expression by cytomegalovirus-activated T cells.

    PubMed Central

    Waldman, W. J.; Knight, D. A.

    1996-01-01

    Cytomegalovirus (CMV) has been associated with allograft rejection and transplantation-associated arteriosclerosis. CMV infects endothelium, the interface between allograft tissue and the host immune system; however, mechanisms by which such interaction might exacerbate the rejection process remain unresolved. Here we test the hypothesis that host immune activity, triggered by CMV-infected graft endothelial cells (ECs), can result in the production of cytokines capable of enhancing the alloimmunogenicity of nearby uninfected endothelia. To model these phenomena in vitro, confluent monolayers of ECs derived from human umbilical vein or adult gonadal vein were incubated 5 days beneath trans-well culture inserts containing CMV-seropositive or CMV-seronegative donor-derived CD3+ or CD4+ T cells alone or in combination with CMV-infected or uninfected allogeneic ECs. The extent of T cell proliferation was determined by [3H]thymidine labeling of trans-well contents after transfer to microtiter plates. Endothelial responses to soluble factors elaborated by CMV-activated T cells were determined by immunohistochemical staining and immunofluorescence flow cytometric analysis of underlying EC monolayers. Results of experiments with CMV-seropositive donor-derived CD4+ T cells demonstrated enhancement of ICAM-1 and histocompatibility leukocyte antigen class I, as well as induction of histocompatibility leukocyte antigen DR on ECs incubated beneath T cell/EC/CMV trans-well co-cultures. Total (CD3+) T cells co-cultured with EC/CMV induced VCAM-1 as well. Furthermore, [3H]thymidine incorporation by these T cells indicated a strong proliferative response. Endothelial responses to T cells alone or in combination with uninfected ECs were minimal, and T cells cultured under these conditions showed little proliferative activity. Similarly, little or no endothelial responses were apparent in monolayers beneath trans-wells containing T cells isolated from CMV-seronegative individuals

  6. The Role of Cytokines in the Functional Activity of Phagocytes in Blood and Colostrum of Diabetic Mothers

    PubMed Central

    Fagundes, Danny Laura Gomes; França, Eduardo Luzía; Morceli, Glilciane; Rudge, Marilza Vieira Cunha; Calderon, Iracema de Mattos Paranhos; Honorio-França, Adenilda Cristina

    2013-01-01

    Immune response changes induced by diabetes are a risk factor for infections during pregnancy and may modify the development of the newborn's immune system. The present study analyzed colostrum and maternal and cord blood of diabetic women to determine (1) the levels of the cytokines IFN-γ and TGF-β and (2) phagocytic activity after incubation with cytokines. Methods. Colostrum and maternal and cord blood samples were classified into normoglycemic (N = 20) and diabetic (N = 19) groups. Cytokine levels, superoxide release, rate of phagocytosis, bactericidal activity, and intracellular Ca2+ release by phagocytes were analyzed in the samples. Irrespective of glycemic status, IFN-γ and TGF-β levels were not changed in colostrum and maternal and cord blood. In maternal blood and colostrum, superoxide release by cytokine-stimulated phagocytes was similar between the groups. Compared to spontaneous release, superoxide release was stimulated by IFN-γ and TGF-β in normoglycemic and diabetic groups. In the diabetic group, cord blood phagocytes incubated with IFN-γ exhibited higher phagocytic activity in response to EPEC, and maternal blood exhibited lower microbicidal activity. These data suggest that diabetes interferes in maternal immunological parameters and that IFN-γ and TGF-β modulate the functional activity of phagocytes in the colostrum, maternal blood, and cord blood of pregnant diabetic women. PMID:24489577

  7. Long-term study of the impact of methotrexate on serum cytokines and lymphocyte subsets in patients with active rheumatoid arthritis: correlation with pharmacokinetic measures

    PubMed Central

    Kremer, Joel M; Lawrence, David A; Hamilton, Robert; McInnes, Iain B

    2016-01-01

    Objective To describe changes in immune parameters observed during long-term methotrexate (MTX) therapy in patients with active rheumatoid arthritis (RA) and explore correlations with simultaneously measured MTX pharmacokinetic (PKC) parameters. Design Prospective, open-label, long-term mechanism of action study. Setting University clinic. Methods MTX was initiated at a single weekly oral dose of 7.5 mg and dose adjusted for efficacy and toxicity for the duration of the study. Standard measures of disease activity were performed at baseline and every 6–36 months. Serum cytokine measurements in blood together with lymphocyte surface immunophenotypes and stimulated peripheral blood mononuclear cell (PBMC) cytokine production were assessed at each clinical evaluation. Results Cytokine concentrations exhibited multiple significant correlations with disease activity measures over time. The strongest correlations observed were for interleukin (IL)-6 (r=0.45, p<0.0001 for swollen joints and r=0.32, p=0.002 for tender joints) and IL-8 (r=0.25, p=0.01 for swollen joints). Significant decreases from baseline were observed in serum IL-1B, IL-6 and IL-8 concentrations. The most significant changes were observed for IL-6 (p<0.001). Significant increases from baseline were observed in IL-2 release from PBMCs ex vivo (p<0.01). In parallel, multiple statistically significant correlations were observed between MTX PKC measures and immune parameters. The change in swollen joint count correlated inversely with the change in area under the curve (AUC) for MTX (r=−0.63, p=0.007). Conclusions MTX therapy of patients with RA is accompanied by a variety of changes in serum cytokine expression, which in turn correlate strongly with clinical disease activity and MTX pharmacokinetics (PKCs). These data strongly support the notion that MTX mediates profound and functionally relevant effects on the immunological hierarchy in the RA lesion. PMID:27335660

  8. Analysis of the expression of toll-like receptors 2 and 4 and cytokine production during experimental Leishmania chagasi infection.

    PubMed

    Cezário, Glaucia Aparecida Gomes; de Oliveira, Larissa Ragozo Cardoso; Peresi, Eliana; Nicolete, Vanessa Cristina; Polettini, Jossimara; de Lima, Carlos Roberto Gonçalves; Gatto, Mariana; Calvi, Sueli Aparecida

    2011-08-01

    Toll-like receptors (TLRs) recognise pathogen-derived molecules and influence immunity to control parasite infections. This study aimed to evaluate the mRNA expression of TLRs 2 and 4, the expression and production of the cytokines interleukin (IL)-12, interferon (IFN)-γ, tumor necrosis factor (TNF)-α, IL-17, IL-10 and transforming growth factor (TGF)-β and the production of nitric oxide (NO) in the spleen of mice infected with Leishmania chagasi. It also aimed to evaluate any correlations between mRNA expression TLR2 and 4 and cytokines and NO production. Infection resulted in increased TLR2-4, IL-17, TNF-α and TGF-β mRNA expression during early infection, with decreased expression during late infection correlating with parasite load. IFN-γ and IL-12 mRNA expression decreased at the peak of parasitism. IL-10 mRNA expression increased throughout the entire time period analysed. Although TGF-β, TNF-α and IL-17 were highly produced during the initial phase of infection, IFN-γ and IL-12 exhibited high production during the final phase of infection. IL-10 and NO showed increased production throughout the evaluated time period. In the acute phase of infection, there was a positive correlation between TLR2-4, TNF-α, IL-17, NO, IL-10 and TGF-β expression and parasite load. During the chronic phase of infection, there was a positive correlation between TLR2-4, TNF-α, IL-17 and TGF-β expression and parasite load. Our data suggest that infection by L. chagasi resulted in modulation of TLRs 2 and 4 and cytokines.

  9. Differential effects of CD28 costimulation upon cytokine production by CD4+ and CD8+ T cells.

    PubMed

    Rochford, Rosemary; Riggs, James E; Clavo, Anaira; Ernst, David N; Hobbs, Monte V

    2004-01-01

    The impact of CD28 ligation upon CD4+ and CD8+ T lymphocyte proliferation and cytokine production was assessed. Although costimulation increased the proliferative response of both T cell subsets, cytokine production was most markedly increased in the CD4+ subset, as evidenced by a 40-fold increase in interleukin-2 (IL-2), a 14-fold increase in interleukin-3 (IL-3) and 5-fold increases in interferon gamma and GM-colony-stimulating factor (CSF) production. The CD8+ T cell response to CD28 ligation was less marked, maxima being a 5-fold increase in IL-2 production and 2-fold increases in IL-3 and GM-CSF production. Resolution of CD4+ and CD8+ T cells into their CD44lo (naïve) and CD44hi (memory/effector) subsets revealed that naive CD4+ T cells were the most CD28-responsive subsets. CD28-mediated costimulation promotes distinct differentiation programs in CD4+ versus CD8+ T cells.

  10. MicroRNA-22 negatively regulates poly(I:C)-triggered type I interferon and inflammatory cytokine production via targeting mitochondrial antiviral signaling protein (MAVS)

    PubMed Central

    Ye, Jing; Duan, Xiaodong; Zohaib, Ali; Wang, Wentao; Chen, Zheng; Zhu, Bibo; Li, Yunchuan; Chen, Huanchun; Cao, Shengbo

    2016-01-01

    MicroRNAs (miRNAs) are small non-coding RNAs that play important roles in regulating the host immune response. Here we found that miR-22 is induced in glial cells upon stimulation with poly(I:C). Overexpression of miR-22 in the cultured cells resulted in decreased activity of interferon regulatory factor-3 and nuclear factor-kappa B, which in turn led to reduced expression of interferon-β and inflammatory cytokines, including tumor necrosis factor-α, interleukin-1β, interleukin-6, and chemokine (C-C motif) ligand 5, upon stimulation with poly(I:C), whereas knockdown of miR-22 had the opposite effect. We used a combination of bioinformatics and experimental techniques to demonstrate that mitochondrial antiviral signaling protein (MAVS), which positively regulates type I interferon production, is a novel target of miR-22. Overexpression of miR-22 decreased the activity of a luciferase reporter containing the MAVS 3′-untranslated region and led to decreased MAVS mRNA and protein levels. In contrast, ectopic expression of miR-22 inhibitor led to elevated MAVS expression. Collectively, our results demonstrate that miR-22 negatively regulates poly(I:C)-induced production of type I interferon and inflammatory cytokines via targeting MAVS. PMID:27705941

  11. Cytokines in sleep regulation.

    PubMed

    Krueger, J M; Takahashi, S; Kapás, L; Bredow, S; Roky, R; Fang, J; Floyd, R; Renegar, K B; Guha-Thakurta, N; Novitsky, S

    1995-01-01

    The central thesis of this essay is that the cytokine network in brain is a key element in the humoral regulation of sleep responses to infection and in the physiological regulation of sleep. We hypothesize that many cytokines, their cellular receptors, soluble receptors, and endogenous antagonists are involved in physiological sleep regulation. The expressions of some cytokines are greatly amplified by microbial challenge. This excess cytokine production during infection induces sleep responses. The excessive sleep and wakefulness that occur at different times during the course of the infectious process results from dynamic changes in various cytokines that occur during the host's response to infectious challenge. Removal of any one somnogenic cytokine inhibits normal sleep, alters the cytokine network by changing the cytokine mix, but does not completely disrupt sleep due to the redundant nature of the cytokine network. The cytokine network operates in a paracrine/autocrine fashion and is responsive to neuronal use. Finally, cytokines elicit their somnogenic actions via endocrine and neurotransmitter systems as well as having direct effects neurons and glia. Evidence in support of these postulates is reviewed in this essay.

  12. Ultrafiltered pig leukocyte extract (IMUNOR) decreases nitric oxide formation and hematopoiesis-stimulating cytokine production in lipopolysaccharide-stimulated RAW 264.7 macrophages.

    PubMed

    Hofer, Michal; Vacek, Antonín; Lojek, Antonín; Holá, Jirina; Streitová, Denisa

    2007-10-01

    A low-molecular-weight (<12 kDa) ultrafiltered pig leukocyte extract, IMUNOR, was tested in experiments in vitro on non-stimulated and lipopolysaccharide (LPS)-stimulated murine RAW 264.7 macrophages in order to assess modulation of nitric oxide (NO) production (measured indirectly as the concentration of nitrites), hematopoiesis-stimulating activity of the supernatant of the macrophage cells (ascertained by counting cell colonies growing from progenitor cells for granulocytes and macrophages (GM-CFC) in vitro), and the release of hematopoiesis-stimulating cytokines. No hematopoiesis-stimulating activity and cytokine or NO production were found in the supernatant of non-stimulated macrophages. It was found that IMUNOR does not influence this status. Supernatant of LPS-stimulated macrophages was characterized by hematopoiesis-stimulating activity, as well as by the presence of nitrites, interleukin-6 (IL-6), and granulocyte colony-stimulating factor (G-CSF). A key role in the hematopoiesis-stimulating activity of the supernatant of LPS-stimulated macrophages could be ascribed to G-CSF since the formation of the colonies could be abrogated nearly completely by monoclonal antibodies against G-CSF. IMUNOR was found to suppress all the mentioned manifestations of the LPS-activated macrophages. When considering these results together with those from our previous in vivo study revealing stimulatory effects of IMUNOR on radiation-suppressed hematopoiesis, a hypothesis may be formulated which postulates a homeostatic role of IMUNOR, consisting in stimulation of impaired immune and hematopoietic systems but also in cutting back the production of proinflammatory mediators in cases of overstimulation which threats with undesirable consequences.

  13. Melanoma tumors alter proinflammatory cytokine production and monoamine brain function, and induce depressive-like behavior in male mice.

    PubMed

    Lebeña, Andrea; Vegas, Oscar; Gómez-Lázaro, Eneritz; Arregi, Amaia; Garmendia, Larraitz; Beitia, Garikoitz; Azpiroz, Arantza

    2014-10-01

    Depression is a commonly observed disorder among cancer patients; however, the mechanisms underlying the relationship between these disorders are not well known. We used an animal model to study the effects of tumor development on depressive-like behavior manifestation, proinflammatory cytokine expression, and central monoaminergic activity. Male OF1 mice were inoculated with B16F10 melanoma tumor cells and subjected to a 21-day behavioral evaluation comprising the novel palatable food (NPF) test and tail suspension test (TST). The mRNA expression levels of proinflammatory cytokines, interleukin (IL)-1β and IL-6, and tumor necrosis factor-alpha (TNF-α), were measured in the hypothalamus and hippocampus and the levels of IL-6 and TNF-α were measured in the blood plasma. We similarly determined the monoamine turnover in various brain areas. The tumors resulted in increasing the immobility in TST and the expression level of IL-6 in the hippocampus. These increases corresponded with a decrease in dopaminergic activity in the striatum and a decrease in serotonin turnover in the prefrontal cortex. Similarly, a high level of tumor development produced increases in the brain expression levels of IL-6 and TNF-α and plasma levels of IL-6. Our findings suggest that these alterations in inflammatory cytokines and monoaminergic system function might be responsible for the manifestation of depressive-like behaviors in tumor-bearing mice.

  14. N-Stearoylethanolamine suppresses the pro-inflammatory cytokines production by inhibition of NF-κB translocation.

    PubMed

    Berdyshev, Andrey G; Kosiakova, Halyna V; Onopchenko, Oleksandra V; Panchuk, Rostislav R; Stoika, Rostislav S; Hula, Nadiya M

    2015-09-01

    N-Stearoylethanolamine (NSE) is a minor lipid that belongs to the N-Acylethanolamines family that mediates a wide range of biological processes. This study investigates the mechanisms of anti-inflammatory action of NSE on different model systems. Namely, we estimated the effect of NSE on inflammatory cytokines mRNA level (leukemia cells L1210), cytokines content (serum and LPS-stimulated macrophages) and nuclear translocation of NF-κB (peritoneal macrophages LPS-stimulated and isolated from rats with obesity-induced insulin resistance). The results indicated that NSE dose-dependently inhibits the IL-1 and IL-6 mRNA level in L1210 cells. Furthermore, the NSE treatment triggered a normalization of serum TNF-α level in insulin resistant rats and a reduction of medium IL-1 level in LPS-activated peritoneal macrophages. These NSE's effects were associated with the inhibition of nuclear NF-κB translocation in rat peritoneal macrophages.

  15. Hepatoblasts comprise a niche for fetal liver erythropoiesis through cytokine production.

    PubMed

    Sugiyama, Daisuke; Kulkeaw, Kasem; Mizuochi, Chiyo; Horio, Yuka; Okayama, Satoko

    2011-07-01

    In mammals, definitive erythropoiesis first occurs in fetal liver (FL), although little is known about how the process is regulated. FL consists of hepatoblasts, sinusoid endothelial cells and hematopoietic cells. To determine niche cells for fetal liver erythropoiesis, we isolated each FL component by flow cytometry. mRNA analysis suggested that Dlk-1-expressing hepatoblasts primarily expressed EPO and SCF, genes encoding erythropoietic cytokines. EPO protein was detected predominantly in hepatoblasts, as assessed by ELISA and immunohistochemistry, and was not detected in sinusoid endothelial cells and hematopoietic cells. To characterize hepatoblast function in FL, we analyzed Map2k4(-/-) mouse embryos, which lack hepatoblasts, and observed down-regulation of EPO and SCF expression in FL relative to wild-type mice. Our observations demonstrate that hepatoblasts comprise a niche for erythropoiesis through cytokine secretion.

  16. Differential Regulation of Proinflammatory Cytokine Expression by Mitogen-Activated Protein Kinases in Macrophages in Response to Intestinal Parasite Infection

    PubMed Central

    Lim, Mei Xing; Png, Chin Wen; Tay, Crispina Yan Bing; Teo, Joshua Ding Wei; Jiao, Huipeng; Lehming, Norbert

    2014-01-01

    Blastocystis is a common enteric protistan parasite that can cause acute, as well as chronic, infection and is associated with irritable bowel syndrome (IBS). However, the pathogenic status of Blastocystis infection remains unclear. In this study, we found that Blastocystis antigens induced abundant expression of proinflammatory cytokines, including interleukin 1β (IL-1β), IL-6, and tumor necrosis factor alpha (TNF-α), in mouse intestinal explants, in mouse colitis colon, and in macrophages. Further investigation utilizing RAW264.7 murine macrophages showed that Blastocystis treatment in RAW264.7 macrophages induced the activation of ERK, JNK, and p38, the three major groups of mammalian mitogen-activated protein (MAP) kinases that play essential roles in the expression of proinflammatory cytokines. ERK inhibition in macrophages significantly suppressed both mRNA and protein expression of IL-6 and TNF-α and mRNA expression of IL-1β. On the other hand, JNK inhibition resulted in reductions in both c-Jun and ERK activation and significant suppression of all three proinflammatory cytokines at both the mRNA and protein levels. Inhibition of p38 suppressed only IL-6 protein expression with no effect on the expression of IL-1β and TNF-α. Furthermore, we found that serine proteases produced by Blastocystis play an important role in the induction of ERK activation and proinflammatory cytokine expression by macrophages. Our study thus demonstrated for the first time that Blastocystis could induce the expression of various proinflammatory cytokines via the activation of MAP kinases and that infection with Blastocystis may contribute to the pathogenesis of inflammatory intestinal diseases through the activation of inflammatory pathways in host immune cells, such as macrophages. PMID:25156742

  17. The Structure of the GM-CSF Receptor Complex Reveals a Distinct Mode of Cytokine Receptor Activation

    SciTech Connect

    Hansen, Guido; Hercus, Timothy R.; McClure, Barbara J.; Stomski, Frank C.; Dottore, Mara; Powell, Jason; Ramshaw, Hayley; Woodcock, Joanna M.; Xu, Yibin; Guthridge, Mark; McKinstry, William J.; Lopez, Angel F.; Parker, Michael W.

    2008-08-11

    Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a pleiotropic cytokine that controls the production and function of blood cells, is deregulated in clinical conditions such as rheumatoid arthritis and leukemia, yet offers therapeutic value for other diseases. Its receptors are heterodimers consisting of a ligand-specific {alpha} subunit and a {beta}c subunit that is shared with the interleukin (IL)-3 and IL-5 receptors. How signaling is initiated remains an enigma. We report here the crystal structure of the human GM-CSF/GM-CSF receptor ternary complex and its assembly into an unexpected dodecamer or higher-order complex. Importantly, mutagenesis of the GM-CSF receptor at the dodecamer interface and functional studies reveal that dodecamer formation is required for receptor activation and signaling. This unusual form of receptor assembly likely applies also to IL-3 and IL-5 receptors, providing a structural basis for understanding their mechanism of activation and for the development of therapeutics.

  18. Th17 cells demonstrate stable cytokine production in a proallergic environment.

    PubMed

    Glosson-Byers, Nicole L; Sehra, Sarita; Stritesky, Gretta L; Yu, Qing; Awe, Olufolakemi; Pham, Duy; Bruns, Heather A; Kaplan, Mark H

    2014-09-15

    Th17 cells are critical for the clearance of extracellular bacteria and fungi, but also contribute to the pathology of autoimmune diseases and allergic inflammation. After exposure to an appropriate cytokine environment, Th17 cells can acquire a Th1-like phenotype, but less is known about their ability to adopt Th2 and Th9 effector programs. To explore this in more detail, we used an IL-17F lineage tracer mouse strain that allows tracking of cells that formerly expressed IL-17F. In vitro-derived Th17 cells adopted signature cytokine and transcription factor expression when cultured under Th1-, Th2-, or Th9-polarizing conditions. In contrast, using two models of allergic airway disease, Th17 cells from the lungs of diseased mice did not adopt Th1, Th2, or Th9 effector programs, but remained stable IL-17 secretors. Although in vitro-derived Th17 cells expressed IL-4Rα, those induced in vivo during allergic airway disease did not, possibly rendering them unresponsive to IL-4-induced signals. However, in vitro-derived, Ag-specific Th17 cells transferred in vivo to OVA and aluminum hydroxide-sensitized mice also maintained IL-17 secretion and did not produce alternative cytokines upon subsequent OVA challenge. Thus, although Th17 cells can adopt new phenotypes in response to some inflammatory environments, our data suggest that in allergic inflammation, Th17 cells are comparatively stable and retain the potential to produce IL-17. This might reflect a cytokine environment that promotes Th17 stability, and allow a broader immune response at tissue barriers that are susceptible to allergic inflammation.

  19. IL-13 induces disease-promoting type 2 cytokines, alternatively activated macrophages and allergic inflammation during pulmonary infection of mice with Cryptococcus neoformans.

    PubMed

    Müller, Uwe; Stenzel, Werner; Köhler, Gabriele; Werner, Christoph; Polte, Tobias; Hansen, Gesine; Schütze, Nicole; Straubinger, Reinhard K; Blessing, Manfred; McKenzie, Andrew N J; Brombacher, Frank; Alber, Gottfried

    2007-10-15

    In the murine model of Cryptococcus neoformans infection Th1 (IL-12/IFN-gamma) and Th17 (IL-23/IL-17) responses are associated with protection, whereas an IL-4-dependent Th2 response exacerbates disease. To investigate the role of the Th2 cytokine IL-13 during pulmonary infection with C. neoformans, IL-13-overexpressing transgenic (IL-13Tg(+)), IL-13-deficient (IL-13(-/-)), and wild-type (WT) mice were infected intranasally. Susceptibility to C. neoformans infection was found when IL-13 was induced in WT mice or overproduced in IL-13Tg(+) mice. Infected IL-13Tg(+) mice had a reduced survival time and higher pulmonary fungal load as compared with WT mice. In contrast, infected IL-13(-/-) mice were resistant and 89% of these mice survived the entire period of the experiment. Ag-specific production of IL-13 by susceptible WT and IL-13Tg(+) mice was associated with a significant type 2 cytokine shift but only minor changes in IFN-gamma production. Consistent with enhanced type 2 cytokine production, high levels of serum IgE and low ratios of serum IgG2a/IgG1 were detected in susceptible WT and IL-13Tg(+) mice. Interestingly, expression of IL-13 by susceptible WT and IL-13Tg(+) mice was associated with reduced IL-17 production. IL-13 was found to induce formation of alternatively activated macrophages expressing arginase-1, macrophage mannose receptor (CD206), and YM1. In addition, IL-13 production led to lung eosinophilia, goblet cell metaplasia and elevated mucus production, and enhanced airway hyperreactivity. This indicates that IL-13 contributes to fatal allergic inflammation during C. neoformans infection.

  20. Leishmania pifanoi Proteoglycolipid Complex P8 Induces Macrophage Cytokine Production through Toll-Like Receptor 4▿

    PubMed Central

    Whitaker, Shanta M.; Colmenares, Maria; Pestana, Karen Goldsmith; McMahon-Pratt, Diane

    2008-01-01

    The P8 proteoglycolipid complex (P8 PGLC) is a glyconjugate expressed by Leishmania mexicana complex parasites. We previously have shown that vaccination with P8 PGLC provides protection against cutaneous leishmaniasis in susceptible BALB/c mice. However, the biological importance of this complex remains unknown. Here we show that P8 PGLC localizes to the surface of Leishmania pifanoi amastigotes and that upon exposure to macrophages, P8 PGLC binds and induces inflammatory cytokine and chemokine mRNAs such as tumor necrosis factor alpha and RANTES early after stimulation. Our studies indicate that cytokine and chemokine induction is dependent upon Toll-like receptor 4 (TLR4). Interestingly, key inflammatory cytokines and chemokines (such as interleukin-6 [IL-6], macrophage inflammatory protein 1β, and beta interferon [IFN-β]) that can be induced through TLR4 activation were not induced or only slightly upregulated by P8 PGLC. Activation by P8 PGLC does not occur in the presence of TLR4 alone and requires both CD14 and myeloid differentiation protein 2 for signaling; this requirement may be responsible for the limited TLR4 response. This is the first characterization of a TLR4 ligand for Leishmania. In vitro experiments indicate that L. pifanoi amastigotes induce lower levels of cytokines in macrophages in the absence of TLR4; however, notably higher IL-10/IFN-γ ratios were found for TLR4-deficient mice than for BALB/c mice. Further, increased levels of parasites persist in BALB/c mice deficient in TLR4. Taken together, these results suggest that TLR4 recognition of Leishmania pifanoi amastigotes is important for the control of infection and that this is mediated, in part, through the P8 PGLC. PMID:18299340

  1. Prolactin modulates cytokine production induced by culture filtrate proteins of M. bovis through different signaling mechanisms in THP1 cells.

    PubMed

    Martínez-Neri, Priscila A; López-Rincón, Gonzalo; Mancilla-Jiménez, Raúl; del Toro-Arreola, Susana; Muñoz-Valle, José Francisco; Fafutis-Morris, Mary; Bueno-Topete, Miriam Ruth; Estrada-Chávez, Ciro; Pereira-Suárez, Ana Laura

    2015-01-01

    The immunomodulatory functions of prolactin (PRL) are well recognized. Augmented PRL plasma levels were observed in patients with advanced tuberculosis (TB). Recently, we have reported that LPS and Mycobacterium bovis (M. bovis) induced differential expression of PRL receptor (PRLR) isoforms in THP-1 cells and bovine macrophages, respectively. The aim of this work was to determine whether PRL should be considered as a potential modulator of the signaling pathways and cytokine synthesis, induced by culture filtrate protein (CFP) from M. bovis in THP-1 monocytes. The THP-1 cells were stimulated with PRL (20ng/mL), M. bovis CFP (50μg/mL). PRLR as well as phosphorylated STAT3, STAT5, Akt1/2/3, ERK1/2 and p38 expression were evaluated by Western blot. IL1-β, TNF-α, IL-6, IL-12, IL-8, and IL-10 concentrations were measured by ELISA. Our results demonstrated that the expression pattern of PRLR short isoforms is induced by M. bovis CFP. M bovis CFP induced phosphorylation of Akt2, ERK1/2, p38, STAT3, and STAT5 pathways. In turn, PRL only activated the JAK2/STAT3-5 signaling pathway. However, when combined both stimuli, PRL significantly increased STAT3-5 phosphorylation and downregulated Akt2, ERK1/2, and p38 phosphorylation. As expected, M. bovis CFP induced substantial amounts of IL1-β, IL-6, TNF-α, IL-8, IL-12, and IL-10. However, the PRL costimulation considerably decreased IL1-β, TNF-α, and IL-12 secretion, and increased IL-10 production. This results suggest that up-regulation of IL-10 by PRL might be modulating the pro-inflammatory response against mycobacterial antigens through the MAPK pathway.

  2. Increased production of proinflammatory cytokines following infection with porcine reproductive and respiratory syndrome virus and Mycoplasma hyopneumoniae.

    PubMed

    Thanawongnuwech, Roongroje; Thacker, Brad; Halbur, Patrick; Thacker, Eileen L

    2004-09-01

    Induction of the proinflammatory cytokines interleukin-1 (IL-1) (alpha and beta), IL-6, IL-8, IL-10, IL-12, and tumor necrosis factor alpha (TNF-alpha) in pulmonary alveolar macrophages (PAMs) was assessed following experimental infection with porcine reproductive and respiratory syndrome virus (PRRSV) and/or Mycoplasma hyopneumoniae by using in vivo and in vitro models. The in vivo model consisted of pigs infected with PRRSV and/or M. hyopneumoniae and necropsied at 10, 28, or 42 days postinfection. Pigs infected with both pathogens had a greater percentage of macroscopic lung lesions, increased clinical disease, and slower viral clearance than pigs infected with either pathogen alone. The pigs infected with both PRRSV and M. hyopneumoniae had significantly increased levels of mRNA for many proinflammatory cytokines in PAMs collected by bronchoalveolar lavage (BAL) at all necropsy dates compared to those in uninfected control pigs. Increased levels of IL-1beta, IL-8, IL-10, and TNF-alpha proteins in BAL fluid, as measured by enzyme-linked immunosorbent assay, confirmed the increased cytokine induction induced by the pathogens. An in vitro model consisted of M. hyopneumoniae-inoculated tracheal ring explants cultured with PRRSV-infected PAMs. PAMs were harvested at 6 or 15 h postinfection with either or both pathogens. The in vitro study detected increased IL-10 and IL-12 mRNA levels in PAMs infected with PRRSV at all time periods. In addition, IL-10 protein levels were significantly elevated in the culture supernatants in the presence of M. hyopneumoniae-inoculated tracheal ring explants. The increased production of proinflammatory cytokines in vivo and in vitro associated with concurrent M. hyopneumoniae and PRRSV infection may play a role in the increased rates of pneumonia associated with PRRSV infection. The increased levels of IL-10 may be a possible mechanism that PRRSV and M. hyopneumoniae use to exacerbate the severity and duration of pneumonia induced by

  3. The effects of vitamin A supplementation with measles vaccine on leucocyte counts and in vitro cytokine production.

    PubMed

    Jensen, Kristoffer Jarlov; Fisker, Ane Bærent; Andersen, Andreas; Sartono, Erliyani; Yazdanbakhsh, Maria; Aaby, Peter; Erikstrup, Christian; Benn, Christine Stabell

    2016-02-28

    As WHO recommends vitamin A supplementation (VAS) at vaccination contacts after age 6 months, many children receive VAS together with measles vaccine (MV). We aimed to investigate the immunological effect of VAS given with MV. Within a randomised placebo-controlled trial investigating the effect on overall mortality of providing VAS with vaccines in Guinea-Bissau, we conducted an immunological sub-study of VAS v. placebo with MV, analysing leucocyte counts, whole blood in vitro cytokine production, vitamin A status and concentration of C-reactive protein (CRP). VAS compared with placebo was associated with an increased frequency of CRP ≥ 5 mg/l (28 v. 12%; P=0·005). Six weeks after supplementation, VAS had significant sex-differential effects on leucocyte, lymphocyte, monocyte and basophil cell counts, decreasing them in males but increasing them in females. Mainly in females, the effect of VAS on cytokine responses differed by previous VAS: in previous VAS recipients, VAS increased the pro-inflammatory and T helper cell type 1 (Th1) cytokine responses, whereas VAS decreased these responses in previously unsupplemented children. In previous VAS recipients, VAS was associated with increased IFN-γ responses to phytohaemagglutinin in females (geometric mean ratio (GMR): 3·97; 95% CI 1·44, 10·90) but not in males (GMR 0·44; 95% CI 0·14, 1·42); the opposite was observed in previously unsupplemented children. Our results corroborate that VAS provided with MV has immunological effects, which may depend on sex and previous VAS. VAS may increase the number of leucocytes, but also repress both the innate and lymphocyte-derived cytokine responses in females, whereas this repression may be opposite if the females have previously received VAS.

  4. Fluoxetine stimulates anti-inflammatory IL-10 cytokine production and attenuates sensory deficits in a rat model of decompression sickness.

    PubMed

    Blatteau, Jean-Eric; de Maistre, Sébastien; Lambrechts, Kate; Abraini, Jacques; Risso, Jean-Jacques; Vallée, Nicolas

    2015-12-15

    Despite "gold standard" hyperbaric oxygen treatment, 30% of patients suffering from neurological decompression sickness still exhibit incomplete recovery, including sensory impairments. Fluoxetine, a well-known antidepressant, is recognized as having anti-inflammatory effects in the setting of cerebral ischemia. In this study, we focused on the assessment of sensory neurological deficits and measurement of circulating cytokines after decompression in rats treated or not with fluoxetine. Seventy-eight rats were divided into a clinical (n = 38) and a cytokine (n = 40) group. In both groups, the rats were treated with fluoxetine (30 mg/kg po, 6 h beforehand) or with a saccharine solution. All of the rats were exposed to 90 m seawater for 45 min before staged decompression. In the clinical group, paw withdrawal force after mechanical stimulation and paw withdrawal latency after thermal stimulation were evaluated before and 1 and 48 h after surfacing. At 48 h, a dynamic weight-bearing device was used to assess postural stability, depending on the time spent on three or four paws. For cytokine analysis, blood samples were collected from the vena cava 1 h after surfacing. Paw withdrawal force and latency were increased after surfacing in the controls, but not in the fluoxetine group. Dynamic weight-bearing assessment highlighted a better stability on three paws for the fluoxetine group. IL-10 levels were significantly decreased after decompression in the controls, but maintained at baseline level with fluoxetine. This study suggests that fluoxetine has a beneficial effect on sensory neurological recovery. We hypothesize that the observed effect is mediated through maintained anti-inflammatory cytokine IL-10 production.

  5. Changes in TNF and IL-6 production after diphtheria toxoid vaccination: drug modulation of the cytokine levels

    PubMed Central

    Bliacher, M. S.; Danilina, A. V.; Kalashnikova, E. A.; Lopatina, T. K.; Fedorova, I. M.

    1996-01-01

    The effect of vaccination with diphtheria toxoid (AD-M) on TNF and IL-6 production has been studied in humans. In the present study it was demonstrated that immunization with AD-M resulted in changes of in vitro TNF and IL-6 production by peripheral blood mononuclear cells. TNF release was suppressed but IL-6 production was stimulated. On the other hand, serum levels of TNF were markedly increased over a period of 3 weeks. It was also demonstrated that the postvaccinal cytokine production disturbances may be corrected by pretreatment with a new synthetic hexapeptid (Imunofan®). It is possible that the imunofan treatment could prevent some postvaccinal complications. PMID:18475748

  6. Cytokine-induced activation of glial cells in the mouse brain is enhanced at an advanced age.

    PubMed

    Deng, X-H; Bertini, G; Xu, Y-Z; Yan, Z; Bentivoglio, M

    2006-08-25

    Numerous neurological diseases which include neuroinflammatory components exhibit an age-related prevalence. The aging process is characterized by an increase of inflammatory mediators both systemically and in the brain, which may prime glial cells. However, little information is available on age-related changes in the glial response of the healthy aging brain to an inflammatory challenge. This problem was here examined using a mixture of the proinflammatory cytokines interferon-gamma and tumor necrosis factor-alpha, which was injected intracerebroventricularly in young (2-3.5 months), middle-aged (10-11 months) and aged (18-21 months) mice. Vehicle (phosphate-buffered saline) was used as control. After a survival of 1 or 2 days (all age groups) or 4 days (young and middle-aged animals), immunohistochemically labeled astrocytes and microglia were investigated both qualitatively and quantitatively. In all age groups, astrocytes were markedly activated in periventricular as well as in deeper brain regions 2 days following cytokine treatment, whereas microglia activation was already evident at 24 h. Interestingly, cytokine-induced activation of both astrocytes and microglia was significantly more marked in the brain of aged animals, in which it included numerous ameboid microglia, than of younger age groups. Moderate astrocytic activation was also seen in the hippocampal CA1 field of vehicle-treated aged mice. FluoroJade B histochemistry and the terminal deoxynucleotidyl transferase-mediated UTP nick-end labeling technique, performed at 2 days after cytokine administration, did not reveal ongoing cell death phenomena in young or aged animals. This indicated that glial cell changes were not secondary to neuronal death. Altogether, the findings demonstrate for the first time enhanced activation of glial cells in the old brain, compared with young and middle-aged subjects, in response to cytokine exposure. Interestingly, the results also suggest that such enhancement

  7. Commercial Product Activation Using RFID

    NASA Technical Reports Server (NTRS)

    Jedrey, Thomas

    2008-01-01

    Radio-frequency identification (RFID) would be used for commercial product activation, according to a proposal. What is new here is the concept of combining RFID with activation - more specifically, using RFID for activating commercial products (principally, electronic ones) and for performing such ancillary functions as tracking individual product units on production lines, tracking shipments, and updating inventories. According to the proposal, an RFID chip would be embedded in each product. The information encoded in the chip would include a unique number for identifying the product. An RFID reader at the point of sale would record the number of the product and would write digital information to the RFID chip for either immediate activation of the product or for later interrogation and processing. To be practical, an RFID product-activation system should satisfy a number of key requirements: the system should be designed to be integrable into the inventory-tracking and the data-processing and -communication infrastructures of businesses along the entire supply chain from manufacture to retail; the system should be resistant to sophisticated hacking; activation codes should be made sufficiently complexity to minimize the probability of activating stolen products; RFID activation equipment at points of sale must be capable to two-way RF communication for the purposes of reading information from, and writing information to, embedded RFID chips; the equipment at points of sale should be easily operable by sales clerks with little or no training; the point-of-sale equipment should verify activation and provide visible and/or audible signals indicating verification or lack thereof; and, the system should be able to handle millions of products per year with minimal human intervention, among other requirements.

  8. Recombinant cytokines from plants.

    PubMed

    Sirko, Agnieszka; Vaněk, Tomas; Góra-Sochacka, Anna; Redkiewicz, Patrycja

    2011-01-01

    Plant-based platforms have been successfully applied for the last two decades for the efficient production of pharmaceutical proteins. The number of commercialized products biomanufactured in plants is, however, rather discouraging. Cytokines are small glycosylated polypeptides used in the treatment of cancer, immune disorders and various other related diseases. Because the clinical use of cytokines is limited by high production costs they are good candidates for plant-made pharmaceuticals. Several research groups explored the possibilities of cost-effective production of animal cytokines in plant systems. This review summarizes recent advances in this field.

  9. Amelioration of adjuvant-induced arthritis by ursolic acid through altered Th1/Th2 cytokine production.

    PubMed

    Ahmad, Sheikh Fayaz; Khan, Beenish; Bani, Sarang; Suri, K A; Satti, N K; Qazi, G N

    2006-03-01

    The objective of the study was to investigate the activity of ursolic acid (UA) on proinflammatory (Th1) and anti-inflammatory (Th2) cytokines in the peripheral blood of arthritic balb/c mice. Ursolic acid is ubiquitous in the plant kingdom and is a constituent of numerous plants which are having diversified phylogenetic origin and taxonomic position. We applied Cytometric bead array (CBA) technology for simultaneously measurement of these cytokines in adjuvant inflammatory arthritis induced mice treated with ursolic acid in graded oral doses. Cytometric bead array uses the sensitivity of amplified fluorescence detection by flowcytometer to measure soluble analytes in a particle based immune assay. This assay can accurately quantitate five cytokines in a 50 microl sample volume. The T-helper (Th1) deviated cells produce detectable level of tumor necrosis factor (TNF-alpha), interleukin-2 (IL-2) and interferon-gamma (IFN-gamma), while the Th2 deviated cells produce significant amount of interleukin-4 (IL-4) and interleukin-5 (IL-5). Oral administration of UA at doses of 10, 20, 40, 80 and 160 mg kg(-1) per oral dose inhibited the presence of IL-2, IFN-gamma and TNF-alpha in the peripheral blood.

  10. Effect of zinc supplementation on E-ADA activity, seric zinc, and cytokines levels of Trypanosoma evansi infected Wistar rats.

    PubMed

    Bottari, Nathieli B; Baldissera, Matheus D; Oliveira, Camila B; Duarte, Thiago; Duarte, Marta M M F; Leal, Marta L R; Thomé, Gustavo R; Zanini, Daniela; Schetinger, Maria Rosa C; Nunes, Matheus A G; Dressler, Valderi L; Monteiro, Silvia G; Tonin, Alexandre A; Da Silva, Aleksandro S

    2014-09-01

    The aim of this study was to evaluate the effect of zinc supplementation on the ecto-adenosine deaminase activity (E-ADA), zinc seric levels and cytokines (TNF-α, IL-1, IL-6, and IL -10) on rats experimentally infected by Trypanosoma evansi. Four groups with 10 rats each were used as negative controls (groups A and B), while the animals from the groups C and D were infected intraperitoneally with 0.1 mL of cryopreserved blood containing 1.4 × 10(4) of trypanosomes. Animals of groups B and D received two doses of Zinc (Zn) at 5 mg kg(-1), subcutaneously, on the 2nd and 7th day post-infection (PI). Blood samples were collected on days 5 (n = 5) and 15 PI (n = 5). Zn supplementation was able to increase the rat's longevity and to reduce their parasitemia. It was observed that seric Zn levels were increased on infected animals under Zn supplementation. Animals that were infected and supplemented with Zn showed changes in E-ADA activity and in cytokine levels (P < 0.05). Zn supplementation of healthy animals (Group B), increased the E-ADA activity, as well as reduced the concentration of cytokines. Infected animals from groups C and D showed increased levels of cytokines. Finally, we observed that Zn supplementation led to a modulation on cytokine's level in rats infected by T. evansi, as well as in E-ADA activity.

  11. Chicken guanylate-binding protein. Conservation of GTPase activity and induction by cytokines.

    PubMed

    Schwemmle, M; Kaspers, B; Irion, A; Staeheli, P; Schultz, U

    1996-04-26

    To gain further insights into the cytokine network of birds, we used polymerase chain reaction technology to clone a cDNA that codes for a chicken homolog of the interferon-induced guanylate-binding proteins (GBPs). In its N-terminal moiety, the 64-kDa chicken GBP contains two sequence blocks of 100 and 19 amino acids, respectively, that are about 70% identical to mammalian GBPs. The first region includes two motifs of the canonical GTP-binding consensus element. The other parts of chicken GBP are poorly conserved, except for a CAAX motif at the extreme C terminus which might signal isoprenylation. Like mammalian GBPs, recombinant chicken GBP specifically bound to agarose-immobilized guanine nucleotides and hydrolyzed GTP to both GDP and GMP. Regulation by interferons was also conserved: chicken GBP RNA was barely detectable in uninduced chicken cells. Low GBP RNA levels were found in cells treated with type I interferon, whereas very high levels were observed in cells treated with supernatant of a chicken T cell line that secretes a gamma-interferon-like activity. Together with recent phylogenetic studies of interferon genes, these results suggest that in spite of low sequence conservation, the various components of the avian interferon system are functionally well conserved.

  12. Abrogation of streptococcal pyrogenic exotoxin B-mediated suppression of phagocytosis in U937 cells by Cordyceps sinensis mycelium via production of cytokines.

    PubMed

    Kuo, Chih-Feng; Chen, Cheng-Chih; Lin, Chiou-Feng; Jan, Ming-Shiou; Huang, Robert Y; Luo, Yueh-Hsia; Chuang, Woei-Jer; Sheu, Chia-Chin; Lin, Yee-Shin

    2007-02-01

    Streptococcal pyrogenic exotoxin B (SPE B) is a virulent factor in group A streptococcal infection. We previously showed that SPE B reduced phagocytosis in human monocytic U937 cells. Here we show that the mycelium extract of Cordyceps sinensis (CS), a Chinese immunomodulatory herbal medicine, increased phagocytosis in U937 cells. Neither heat nor trypsin pretreatment prevented CS extract from causing this increase. Further studies indicated that SPE B-mediated suppression of U937 cell phagocytic activity was abrogated by CS extract. Factors in the conditioned medium from CS-extract-treated U937 cells were responsible for blocking the SPE B-mediated suppression of phagocytosis. Heating the conditioned medium eliminated the increase, which suggested that the U937-cell protein products augmented phagocytosis. Analyzing cytokine mRNA expression of U937 cells revealed increases in interferon-gamma (IFN-gamma), interleukin (IL)-12 p35 and p40, and tumor necrosis factor-alpha (TNF-alpha), but not in IL-1beta, IL-6, or IL-8. Treating U937 cells with anti-IFN-gamma, IL-12, and TNF-alpha antibodies also eliminated the conditioned medium-induced increase in phagocytosis. Taken together, SPE B inhibited phagocytosis, but CS mycelium extract abrogated this inhibition by causing cytokine production.

  13. Mycobacterial heat shock protein 70 induces interleukin-10 production: immunomodulation of synovial cell cytokine profile and dendritic cell maturation

    PubMed Central

    DETANICO, T; RODRIGUES, L; SABRITTO, A C; KEISERMANN, M; BAUER, M E; ZWICKEY, H; BONORINO, C

    2004-01-01

    Cytokines are key modulators of the immune responses that take place in the inflamed synovium of arthritis patients. Consequently, substances that can reverse the inflammatory profile of the inflamed joint are potential tools for clinical management of the disease. Mycobacterial heat shock protein 70 (MTBHSP70) has been found to protect rats from experimentally induced arthritis through the induction of interleukin (IL)-10-producing T cells. In this study, we have demonstrated that MTBHSP70 induces IL-10 production in synoviocytes from arthritis patients and peripheral blood monoculear cells (PBMCs) from both patients and healthy controls. IL-10 production was accompanied by a decrease in tumour necrosis factor (TNF)-α production by synovial cells. Separation studies showed that the target cells were mainly monocytes. Accordingly, we observed that MTBHSP70 delayed maturation of murine bone marrow-derived dendritic cells. Our results suggest that MTBHSP may act on antigen-presenting cells (APCs) to modulate the cytokine response in arthritis and support an anti-inflammatory role for this protein, suggesting that it may be of therapeutic use in the modulation of arthritis. PMID:14738465

  14. Mature IgM-expressing plasma cells sense antigen and develop competence for cytokine production upon antigenic challenge

    PubMed Central

    Blanc, Pascal; Moro-Sibilot, Ludovic; Barthly, Lucas; Jagot, Ferdinand; This, Sébastien; de Bernard, Simon; Buffat, Laurent; Dussurgey, Sébastien; Colisson, Renaud; Hobeika, Elias; Fest, Thierry; Taillardet, Morgan; Thaunat, Olivier; Sicard, Antoine; Mondière, Paul; Genestier, Laurent; Nutt, Stephen L.; Defrance, Thierry

    2016-01-01

    Dogma holds that plasma cells, as opposed to B cells, cannot bind antigen because they have switched from expression of membrane-bound immunoglobulins (Ig) that constitute the B-cell receptor (BCR) to production of the secreted form of immunoglobulins. Here we compare the phenotypical and functional attributes of plasma cells generated by the T-cell-dependent and T-cell-independent forms of the hapten NP. We show that the nature of the secreted Ig isotype, rather than the chemical structure of the immunizing antigen, defines two functionally distinct populations of plasma cells. Fully mature IgM-expressing plasma cells resident in the bone marrow retain expression of a functional BCR, whereas their IgG+ counterparts do not. Antigen boost modifies the gene expression profile of IgM+ plasma cells and initiates a cytokine production program, characterized by upregulation of CCL5 and IL-10. Our results demonstrate that IgM-expressing plasma cells can sense antigen and acquire competence for cytokine