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Sample records for human cytokine synthesis

  1. Synthesis of hyaluronic acid by human peritoneal mesothelial cells: effect of cytokines and dialysate.

    PubMed

    Breborowicz, A; Korybalska, K; Grzybowski, A; Wieczorowska-Tobis, K; Martis, L; Oreopoulos, D G

    1996-01-01

    To assess effects of the inflammatory cytokines (IL-1-beta, TNF-alpha, TGF-beta 1) and dialysate effluent on synthesis of hyaluronic acid by human peritoneal mesothelial cells (HMC) in in vitro culture. Dialysate effluent was collected after the overnight dwell of Dianeal 1.5% from patients during CAPD training. HMC were obtained from omentum from nonuremic donors or were harvested from the dialysate effluent from CAPD patients. Synthesis of hyaluronic acid was studied on monolayers of HMC, which were deprived of serum 48 hours prior to experiment. Effects of cytokines were tested in a medium with low serum concentration (0.1%) or in medium mixed (1:1 v/v) with the autologous dialysate. Hyaluronic acid level in medium was measured with radioimmunoassay. Cytokines enhanced synthesis of hyaluronic acid by HMC, and the strongest effect was induced by IL-1. Effluent dialysate stimulates synthesis of hyaluronic acid stronger than 10% FCS. Effluent dialysate and IL-1 synergistically enhance synthesis of hyaluronic acid by HMC. Effluent dialysate from CAPD patients stimulates production of hyaluronic acid by HMC and acts synergistically with cytokines.

  2. Leptin enhances survival and induces migration, degranulation, and cytokine synthesis of human basophils.

    PubMed

    Suzukawa, Maho; Nagase, Hiroyuki; Ogahara, Ikuko; Han, Kaiyu; Tashimo, Hiroyuki; Shibui, Akiko; Koketsu, Rikiya; Nakae, Susumu; Yamaguchi, Masao; Ohta, Ken

    2011-05-01

    Basophils are the rarest leukocytes in human blood, but they are now recognized as one of the most important immunomodulatory as well as effector cells in allergic inflammation. Leptin, a member of the IL-6 cytokine family, has metabolic effects as an adipokine, and it is also known to participate in the pathogenesis of inflammatory reactions. Because there is an epidemiologic relationship between obesity and allergy, we examined whether basophil functions are modified by leptin. We found that human basophils express leptin receptor (LepR) at both the mRNA and surface protein levels, which were upregulated by IL-33. Leptin exerted strong effects on multiple basophil functions. It induced a strong migratory response in human basophils, similar in potency to that of basophil-active chemokines. Also, leptin enhanced survival of human basophils, although its potency was less than that of IL-3. Additionally, CD63, a basophil activation marker expressed on the cell surface, was upregulated by leptin, an effect that was neutralized by blocking of LepR. Assessments of basophil degranulation and cytokine synthesis found that leptin showed a strong priming effect on human basophil degranulation in response to FcεRI aggregation and induced Th2, but not Th1, cytokine production by the cells. In summary, the present findings indicate that leptin may be a key molecule mediating the effects of adipocytes on inflammatory cells such as basophils by binding to LepR and activating the cellular functions, presumably exacerbating allergic inflammation.

  3. Spermine Inhibits Proinflammatory Cytokine Synthesis in Human Mononuclear Cells: A Counterregulatory Mechanism that Restrains the Immune Response

    PubMed Central

    Zhang, Minghuang; Caragine, Theresa; Wang, Haichao; Cohen, Pamela S.; Botchkina, Galina; Soda, Kuniyasu; Bianchi, Marina; Ulrich, Peter; Cerami, Anthony; Sherry, Barbara; Tracey, Kevin J.

    1997-01-01

    The local production of proinflammatory cytokines mediates the host response to inflammation, infection, and injury, whereas an overexpression of these mediators can injure or kill the host. Recently, we identified a class of multivalent guanylhydrazone compounds that are effective inhibitors of proinflammatory cytokine synthesis in monocytes/macrophages. The structure of one such cationic molecule suggested a molecular mimicry with spermine, a ubiquitous endogenous biogenic amine that increases significantly at sites of inflammation and infection. Here, we addressed the hypothesis that spermine might counterregulate the innate immune response by downregulating the synthesis of potentially injurious cytokines. When spermine was added to cultures of human peripheral blood mononuclear cells stimulated with lipopolysaccharide (LPS), it effectively inhibited the synthesis of the proinflammatory cytokines tumor necrosis factor (TNF), interleukin-1 (IL-1), IL-6, MIP-1α, and MIP-1β. The inhibition of cytokine synthesis was specific and reversible, with significant inhibition of TNF synthesis occurring even when spermine was added after LPS. The mechanism of spermine-mediated cytokine suppression was posttranscriptional and independent of polyamine oxidase activity. Local administration of spermine in vivo protected mice against the development of acute footpad inflammation induced by carrageenan. These results identify a distinct molecular counterregulatory role for spermine in downregulating the monocyte proinflammatory cytokine response. PMID:9151701

  4. Calcium dependent and independent cytokine synthesis by air pollution particle-exposed human bronchial epithelial cells

    SciTech Connect

    Sakamoto, Noriho; Hayashi, Shizu; Gosselink, John; Ishii, Hiroshi; Ishimatsu, Yuji; Mukae, Hiroshi; Hogg, James C.; Eeden, Stephan F. van

    2007-12-01

    Exposure to ambient air pollution particles with a diameter of < 10 {mu}m (PM{sub 10}) has been associated with increased cardiopulmonary morbidity and mortality. We have shown that human bronchial epithelial cells (HBECs) exposed to PM{sub 10} produce pro-inflammatory mediators that contribute to a local and systemic inflammatory response. Changes in intracellular calcium concentrations ([Ca{sup 2+}]{sub i}) have been demonstrated to regulate several functions of the airway epithelium including the production of pro-inflammatory mediators. The aim of the present study was to determine the nature and mechanism of calcium responses induced by PM{sub 10} in HBECs and its relationship to cytokine synthesis. Methods: Primary HBECs were exposed to urban air pollution particles (EHC-93) and [Ca{sup 2+}]{sub i} responses were measured using the fluoroprobe (Fura-2). Cytokine levels were measured at mRNA and protein levels using real-time PCR and ELISA. Results: PM{sub 10} increased [Ca{sup 2+}]{sub i} in a dose-dependent manner. This calcium response was reduced by blocking the influx of calcium into cells (i.e. calcium-free medium, NiCl{sub 2}, LaCl{sub 3}). PM{sub 10} also decreased the activity of calcium pumps. PM{sub 10} increased the production of IL-1{beta}, IL-8, GM-CSF and LIF. Preincubation with intracellular calcium chelator (BAPTA-AM) attenuated IL-1{beta} and IL-8 production, but not GM-CSF and LIF production. Conclusion: We conclude that exposure to PM{sub 10} induces an increase in cytosolic calcium and cytokine production in bronchial epithelial cells. Our results also suggest that PM{sub 10} induces the production of pro-inflammatory mediators via either intracellular calcium-dependent (IL-1{beta}, IL-8) or -independent (GM-CSF, LIF) pathways.

  5. Cytokines in human milk.

    PubMed

    Srivastava, M D; Srivastava, A; Brouhard, B; Saneto, R; Groh-Wargo, S; Kubit, J

    1996-09-01

    Breast feeding improves the health of children. The greatest significance is to host defense, prevention of autoimmunity, and development of the digestive system; however, the underlying mechanisms for these effects are not well understood. Based on recent evidence that cytokines might be important in these processes, we have used ELISA to quantitate the cytokines in human colostrum, transitional, and mature milk from mothers delivering preterm or at term. We also used reverse transcription PCR to test breast milk cells for the production of cytokine mRNA. No significant (< 10 pg/ml) GM-CSF, SCF, LIF, MIP-1 alpha, IL-2, IL-4, IL-11, IL-12, IL-13, IL-15, sIL-2R, or IFN-gamma was detected. And, in contrast to earlier studies using bioassays or RIA, no significant IL-1 beta, TNF-alpha, or IL-6 was present; nor was IL-10, which had been tested using less specific antibodies. We did confirm the presence of high levels of M-CSF, which remained high throughout lactation. Human milk contained latent, but not free, TGF-beta 1, and especially TGF-beta 2, both of which may be activated by gastric acid pH. High levels of IL-1RA were detected, and like activated TGF-beta, may protect against autoimmunity. Chemokines, particularly GRO-alpha and MCP-1, but also RANTES and IL-8, were present and could protect against infection. Maternal cells in breast milk expressed mRNA for MCP-1 (20/20), IL-8 (14/20), TGF-beta 1 (14/16), TGF-beta 2 (4/6), M-CSF (9/12), IL-6 (6/12) and IL-1 beta (7/12), and may be a source of these cytokines. mRNA for IL-2, IL-10, IFN-gamma, TNF-alpha was not detected and only weak expression was found for RANTES (1/18). There was considerable variability between individual women, and women delivering preterm had lower levels of several cytokines in colostrum than women delivering at term. Yet, cytokine levels remained high months to years into lactation, providing immunological benefit to the breastfed infant/child.

  6. An Autocrine Cytokine/JAK/STAT-Signaling Induces Kynurenine Synthesis in Multidrug Resistant Human Cancer Cells

    PubMed Central

    Campia, Ivana; Buondonno, Ilaria; Castella, Barbara; Rolando, Barbara; Kopecka, Joanna; Gazzano, Elena; Ghigo, Dario; Riganti, Chiara

    2015-01-01

    Background Multidrug resistant cancer cells are hard to eradicate for the inefficacy of conventional anticancer drugs. Besides escaping the cytotoxic effects of chemotherapy, they also bypass the pro-immunogenic effects induced by anticancer drugs: indeed they are not well recognized by host dendritic cells and do not elicit a durable anti-tumor immunity. It has not yet been investigated whether multidrug resistant cells have a different ability to induce immunosuppression than chemosensitive ones. We addressed this issue in human and murine chemosensitive and multidrug resistant cancer cells. Results We found that the activity and expression of indoleamine 2,3-dioxygenase 1 (IDO1), which catalyzes the conversion of tryptophan into the immunosuppressive metabolite kynurenine, was higher in all the multidrug resistant cells analyzed and that IDO1 inhibition reduced the growth of drug-resistant tumors in immunocompetent animals. In chemoresistant cells the basal activity of JAK1/STAT1 and JAK1/STAT3 signaling was higher, the STAT3 inhibitor PIAS3 was down-regulated, and the autocrine production of STAT3-target and IDO1-inducers cytokines IL-6, IL-4, IL-1β, IL-13, TNF-α and CD40L, was increased. The disruption of the JAK/STAT signaling lowered the IDO1 activity and reversed the kynurenine-induced pro-immunosuppressive effects, as revealed by the restored proliferation of T-lymphocytes in STAT-silenced chemoresistant cells. Conclusions Our work shows that multidrug resistant cells have a stronger immunosuppressive attitude than chemosensitive cells, due to the constitutive activation of the JAK/STAT/IDO1 axis, thus resulting chemo- and immune-evasive. Disrupting this axis may significantly improve the efficacy of chemo-immunotherapy protocols against resistant tumors. PMID:25955018

  7. Isolation and expression of human cytokine synthesis inhibitory factor cDNA clones: Homology to Epstein-Barr virus open reading frame BCRFI

    SciTech Connect

    Vieira, P.; De Waal-Malefyt, R.; Dang, M.N.; Johnson, K.E.; Kastelein, R.; Fiorentino, D.F.; DeVries, J.E.; Roncarolo, M.G.; Mosmann, T.R.; Moore, K.W. )

    1991-02-15

    The authors demonstrated the existence of human cytokine synthesis inhibitory factor (DSIF) (interleukin 10 (IL-10)). cDNA clones encoding human IL-10 (hIL-10) were isolated from a tetanus toxin-specific human T-cell clone. Like mouse IL-10, hIL-10 exhibits strong DNA and amino acid sequence homology to an open reading frame in the Epstein-Barr virus, BDRFL. hIL-10 and the BCRFI product inhibit cytokine synthesis by activated human peripheral blood mononuclear cells and by a mouse Th1 clone. Both hIL-10 and mouse IL-10 sustain the viability of a mouse mast cell line in culture, but BCRFI lacks comparable activity in this way, suggesting that BCRFI may have conserved only a subset of hIL-10 activities.

  8. Cytokine synthesis in human peripheral blood mononuclear cells after oral administration of polyenzyme preparations.

    PubMed

    Desser, L; Rehberger, A; Kokron, E; Paukovits, W

    1993-01-01

    Pharmaceutical preparations containing mixtures of various proteolytic and nonproteolytic enzymes have been suggested for use in the treatment of malignant diseases. However, the mode of action of such preparations was not clear. We have shown before that intact bromelain, papain or amylase, which are components of a commercial polyenzyme preparation, induce cytokine production in peripheral blood mononuclear cells in vitro. IFN-alpha and IFN-gamma which had no effect alone, synergistically increased TNF production when applied together with the enzymes. Here we show that trypsin alone had only a small inducing effect. The tryptic but not the autolytic fragments of papain and bromelain have a higher (10- to 40-fold) inducing capacity for TNF production than the untreated enzyme. Additionally we demonstrate that after ingestion of milligram doses of the polyenzyme preparation (as recommended for clinical use), PBMNC of healthy donors acquire the ability to produce TNF-alpha, IL-1 beta and IL-6 when incubated ex vivo with IFN-gamma. Our results indicate that the biological effects observed after oral administration of polyenzyme preparations are related to their ability to induce cytokine production. This may explain the antitumor effects of such enzymes. Our results also suggest that polyenzyme preparations may have a stronger immunomodulary effect when used in combination with IFN-gamma.

  9. Inhibition by rebamipide of cytokine-induced or lipopolysaccharide-induced chemokine synthesis in human corneal fibroblasts.

    PubMed

    Fukuda, Ken; Ishida, Waka; Tanaka, Hiroshi; Harada, Yosuke; Fukushima, Atsuki

    2014-12-01

    The dry-eye drug rebamipide has mucin secretagogue activity in and anti-inflammatory effects on corneal epithelial cells. Corneal stromal fibroblasts (transdifferentiated keratocytes) function as immune modulators in the pathogenesis of chronic ocular allergic inflammation and in innate immune responses at the ocular surface. The possible anti-inflammatory effects of rebamipide on human corneal stromal fibroblasts were examined. Serum-deprived cells were incubated for 1 h with rebamipide and then for various times in the additional absence or presence of cytokines or bacterial lipopolysaccharide (LPS). The release of chemokines into culture supernatants was determined with ELISAs. The intracellular abundance of chemokine mRNAs was quantitated by reverse transcription and real-time PCR analysis. Degradation of the nuclear factor κB (NFκB) inhibitor IκBα was detected by immunoblot analysis. Rebamipide suppressed the release of interleukin (IL)-8 and the upregulation of IL-8 mRNA induced by tumour necrosis factor α (TNF-α) or LPS in corneal fibroblasts. It also inhibited eotaxin-1 (CCL-11) expression at the protein and mRNA levels induced by the combination of TNF-α and IL-4. In addition, rebamipide attenuated the degradation of IκBα induced by TNF-α or LPS. Rebamipide inhibited the synthesis of chemokines by corneal fibroblasts in association with suppression of NFκB signalling. Rebamipide may therefore prove effective for the treatment of corneal stromal inflammation associated with allergy or bacterial infection. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.

  10. Cytokines in human leptospirosis.

    PubMed

    Papa, Anna; Kotrotsiou, Tzimoula

    2015-12-01

    Leptospirosis is a zoonotic disease with increased public health concern. Cytokines produced in response to the infection with pathogenic leptospires have been proposed to be involved in the pathogenesis of the disease. The aim of the study was to measure and evaluate the levels of 27 cytokines in patients with acute leptospirosis. The levels of 27 cytokines were measured from 42 acute leptospirosis cases; 47 samples were obtained from severe cases. Statistical analysis was performed using SPSS. IL-6, IL-8, GM-CSF, IP-10, MCP-1, and VEGF levels differed significantly between the severe cases and the control group, while GM-CSF levels differed significantly between the mild cases and the control group (p<0.05). IL-6, IP-10 and MCP-1 were elevated in most cases. IP-10 was significantly higher in severe than in non-severe cases (p<0.05). The high IP-10 levels suggest a cellular immune response, despite the fact that leptospires are not intracellular organisms. IL-1ra, MCP-1, MIP-1b and TNF-α peaked 1-5 days post onset of illness (p.o.i.), IL-1b, IL-6, IL-8, IL-9, GM-CSF, IP-10 and MIP-1a peaked 6-10 days p.o.i., while VEGF peaked later (11-15 days p.o.i.). TNF-α was significantly lower in the severe cases with pulmonary involvement (p<0.05). Having a better insight into the host immune response in leptospirosis could be the basis for immunotherapeutic targets, especially for the severe cases in which antibiotic treatment is not enough. © The Author 2015. Published by Oxford University Press on behalf of Royal Society of Tropical Medicine and Hygiene. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  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. Monocyte cytokine synthesis in response to extracellular cell stress proteins suggests these proteins exhibit network behaviour.

    PubMed

    Kaiser, Frank; Steptoe, Andrew; Thompson, Stephen; Henderson, Brian

    2014-01-01

    Human peripheral blood monocytes were exposed to single or pairs of cell stress proteins (CSPs), specifically Hsp10, Hsp27, Hsp60 and Hsp70-the former two having anti-inflammatory actions while the latter pair being assumed to be pro-inflammatory in activity. This study was to test if these proteins exhibited any network behaviour. To control for possible lipopolysaccharide contamination, polymyxin B was used. Surprisingly, at concentrations higher than 1 μg/ml, polymyxin B itself could induce cytokine synthesis. A number of commercial sources of the molecular chaperones were tested, and marked variations in monocyte cytokine synthesis were found. All four CSPs stimulated the same profile of IL-1/IL-6 synthesis and IL-10/TNF-α synthesis although the kinetics of production of these two pairs of cytokines were very different. A key question was whether extracellular molecular chaperones exhibited network behaviour. To test this, monocytes were cultured with suboptimal concentrations of single CSP and pairs of CSP to look for additive, synergistic or antagonistic cell responses. The major finding was that pairs of molecular chaperones, including chaperones thought to stimulate monocyte cytokine synthesis, could produce significant antagonistic cellular responses. This demonstrates that extracellular CSPs constitute an additional potent layer within the complex cytokine network and furthermore suggests that monocytes have evolved to dampen their immune responses upon exposure to extracellular networks of CSPs-perhaps as a mechanism for protecting cells against detrimental cellular stress responses.

  13. Human herpesvirus 8-encoded cytokines.

    PubMed

    Nicholas, John

    2010-03-01

    Human herpesvirus (HHV)-8, also called Kaposi's sarcoma-associated herpesvirus, was discovered in 1994 and was rapidly sequenced, revealing several unique and surprising features of its genetic makeup. Among these discoveries was the identification of the first viral homolog of IL-6 and three CC/beta-chemokine ligands (viral CCL-1, -2 and -3), not previously found in gamma-herpesviruses. Viral IL-6 was immediately recognized as a potential contributor to HHV-8 pathogenesis, specifically endothelial-derived Kaposi's sarcoma and the B-cell malignancy multicentric Castleman's disease with which IL-6, a proangiogenic and B-cell growth factor, had previously been implicated. The roles of the viral chemokines were speculated to involve immune evasion; however, like viral IL-6, the viral chemokines have the potential to contribute to pathogenesis through their shared angiogenic activities, known to be important for Kaposi's sarcoma and HHV-8-associated primary effusion lymphoma, and also via direct prosurvival activities. This article will discuss the molecular properties, activities and functions of viral IL-6 and the viral CCLs, proteins that could provide appropriate targets for antiviral and therapeutic strategies.

  14. Cytokines and immune surveillance in humans

    NASA Technical Reports Server (NTRS)

    Sonnenfeld, Gerald

    1994-01-01

    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 explore further the effects of space flight on cyotokines and cytokine-directed immunological function. Among the tests carried out are interferon-alpha production, interferon-gamma production, interleukin-1 and -2 production, signal transduction in neutrophils, signal transduction in monocytes, and monocyte phagocytic activity. The experiments will be performed using peripheral blood obtained from human subjects. It is our intent to eventually carry out these experiments using astronauts as subjects to determine the effects of space flight on cytokine production and activity. However, these subjects are not currently available. Until they become available, we will carry out these experiments using subjects maintained in the bed-rest model for microgravity.

  15. A possible role of autogenous IFN-beta for cytokine productions in human fibroblasts.

    PubMed

    Sano, Emiko; Ohashi, Kensaku; Sato, Yuichiro; Kashiwagi, Masamitsu; Joguchi, Atsuhiro; Naruse, Norio

    2007-04-15

    It has been already known that human diploid fibroblasts are able to produce not only high levels of IFN-beta but also various kinds of cytokines by poly rI: poly rC, and some inflammatory cytokines are induced by IFN-beta gene activation. We also obtained similar results. However, in our system, cytokine productions were extremely enhanced by treating the cells with a low dose of type 1 IFN and the priming effects on cytokine productions were blocked by cycloheximide similar to those on IFN-beta productions. Most of cytokines were produced later than IFN-beta and synthesis patterns of their mRNA showed the same phenomena. We made clear that cytokine productions by poly rI: poly rC are mediated by secreted IFN-beta at a protein level using a monoclonal antibody against human IFN-beta. Further, it was shown that intra-cellular IFN-beta which is not secreted might also participate in cytokine productions. Meanwhile, IL-1beta induced various kinds of cytokines in human fibroblasts and production time courses of these cytokines were similar to those of poly rI: poly rC induced cytokines. Although secreted IFN-beta was not detected in IL-1beta stimulated culture, expression of IFN-beta mRNA was augmented. These results showed that priming effects of type 1 IFN on cytokine productions by poly rI: poly rC might not be the direct action, but successive IFN-beta production might be essential in the production processes of other cytokines. Further, it was suggested that inducible IFN-beta might also take part in IL-1beta-induced cytokine productions.

  16. Production of interleukin-4, interferon (IFN)-gamma and IFN-alpha in human immunodeficiency virus-1 infection: an imbalance of type 1 and type 2 cytokines may reduce the synthesis of IFN-alpha.

    PubMed

    Hober, D; Benyoucef, S; Chehadeh, W; Chieux, V; De La Tribonniere, X; Mouton, Y; Bocket, L; Wattre, P

    1998-10-01

    Interferon-alpha (IFN-alpha) is an important molecule in the antiviral response, but cells from HIV-1-infected individuals show a reduced ability to secrete IFN-alpha. We investigated an association between an imbalance of type 1/type2 cytokines and the production of IFN-alpha in HIV-1 infection. We used whole blood culture to study the cytokine production profile, interferon-gamma (IFN-gamma) and interleukin-4 (IL-4), in response to HIV-1 antigens and to study the Sendai Virus and HSV-1-induced-production of IFN-alpha in seven HIV-1-infected patients. An impaired synthesis of IFN-alpha was obtained in patients with a predominant IL-4 production (IL-4 > IFN-gamma), and we found a positive correlation between the ex vivo production of IFN-alpha and the IFN-gamma/IL-4 ratio but not with the HIV RNA copy number in plasma. We investigated the role of T-cell-derived cytokines in the in vitro production of IFN-alpha by PBMC from eight healthy donors, activated with Sendai Virus or HSV-1. Whereas type 2 cytokines (IL-4, IL-13) inhibited virus-induced IFN-alpha synthesis, on the contrary, type 1 cytokines (IL-2, IFN-gamma) enhanced it. A disarray in the T-cell-derived cytokine response may play a role in the defect of IFN-alpha production in HIV-1-infected individuals. Further investigations are needed to explore this hypothesis.

  17. Inhibition of substance P-induced cytokine synthesis by St. John's wort extracts.

    PubMed

    Fiebich, B L; Höllig, A; Lieb, K

    2001-07-01

    We tested the hypothesis that extracts from St. John's wort interfere with protein synthesis induced by substance P (SP), a neuropeptide which has been implicated in the etiopathology of depression and anxiety. Using human astrocytoma cells, which express functional neurokinin (NK)-1-receptors, we investigated whether extracts from St. John's wort are able to inhibit SP-induced synthesis of the cytokine interleukin-6 (IL-6). We found a potent and dose-dependent inhibition of SP-induced IL-6 synthesis by various extracts from St. John's wort. These results do not only give further evidence of the anti-inflammatory effects of St. John's wort, but also lend support to the hypothesis that the antidepressant effect of St. John's wort is, at least in part, a result of its inhibitory effects on SP-induced protein synthesis.

  18. Temporal Trends in the Inflammatory Cytokine Profile of Human Breastmilk

    PubMed Central

    Chollet-Hinton, Lynn S.; Stuebe, Alison M.; Casbas-Hernandez, Patricia; Chetwynd, Ellen

    2014-01-01

    Abstract A longer lifetime duration of breastfeeding may decrease the risk of breast cancer by reducing breast inflammation and mitigating inflammatory cytokine expression during postlactational involution. However, little is known about how the inflammatory cytokine profile in human breastmilk changes over time. To study temporal trends in breastmilk cytokine expression, we measured 80 human cytokines in the whey fraction of breastmilk samples from 15 mothers at 1, 4, 8, and 12 weeks postpartum. We used mixed models to identify temporal changes in cytokine expression and investigated parity status (multiparous vs. primiparous) as a potential confounder. Nine cytokines (monocyte chemoattractant protein-1, epithelial-derived neutrophil-activating protein-78, hepatocyte growth factor, insulin-like growth factor-binding protein-1, interleukin-16, interleukin-8, macrophage colony-stimulating factor, osteoprotegerin, and tissue inhibitor of metallopeptidase-2) had significantly decreased expression with increasing breastfeeding duration; all nine have known roles in breast involution, inflammation, and cancer and may serve as biomarkers of changing breast microenvironment. No cytokine significantly increased in level over the study period. Total protein concentration significantly decreased over time (p<0.0001), which may mediate the association between length of breastfeeding and inflammatory cytokine expression. Parity status did not confound temporal trends, but levels of several cytokines were significantly higher among multiparous versus primiparous women. Our results suggest that inflammatory cytokine expression during lactation is dynamic, and expressed milk may provide a noninvasive window into the extensive biological changes that occur in the postpartum breast. PMID:25380323

  19. Temporal trends in the inflammatory cytokine profile of human breastmilk.

    PubMed

    Chollet-Hinton, Lynn S; Stuebe, Alison M; Casbas-Hernandez, Patricia; Chetwynd, Ellen; Troester, Melissa A

    2014-12-01

    A longer lifetime duration of breastfeeding may decrease the risk of breast cancer by reducing breast inflammation and mitigating inflammatory cytokine expression during postlactational involution. However, little is known about how the inflammatory cytokine profile in human breastmilk changes over time. To study temporal trends in breastmilk cytokine expression, we measured 80 human cytokines in the whey fraction of breastmilk samples from 15 mothers at 1, 4, 8, and 12 weeks postpartum. We used mixed models to identify temporal changes in cytokine expression and investigated parity status (multiparous vs. primiparous) as a potential confounder. Nine cytokines (monocyte chemoattractant protein-1, epithelial-derived neutrophil-activating protein-78, hepatocyte growth factor, insulin-like growth factor-binding protein-1, interleukin-16, interleukin-8, macrophage colony-stimulating factor, osteoprotegerin, and tissue inhibitor of metallopeptidase-2) had significantly decreased expression with increasing breastfeeding duration; all nine have known roles in breast involution, inflammation, and cancer and may serve as biomarkers of changing breast microenvironment. No cytokine significantly increased in level over the study period. Total protein concentration significantly decreased over time (p<0.0001), which may mediate the association between length of breastfeeding and inflammatory cytokine expression. Parity status did not confound temporal trends, but levels of several cytokines were significantly higher among multiparous versus primiparous women. Our results suggest that inflammatory cytokine expression during lactation is dynamic, and expressed milk may provide a noninvasive window into the extensive biological changes that occur in the postpartum breast.

  20. AMBIENT PARTICULATE MATTER DECREASED IN HUMAN ALVEOLAR MACHROPHAGE CYTOKINE RELEASE

    EPA Science Inventory

    Human exposure to ambient airborne particulate matter (PM) is associated with cardiopulmonary mortality and morbidity, including increased hospitalizations for lung infection. Normal lung immune responses to bacterial infection include alveolar macrophage cytokine production and...

  1. Evaluation of Cytokine Synthesis in Human Whole Blood by Enzyme Linked Immunoassay (ELISA), Reverse Transcriptase Polymerase Chain Reaction (RT-PCR), and Flow Cytometry

    DTIC Science & Technology

    2007-05-08

    following incubation with low doses of LPS. MATERIALS AND METHODS Whole Blood Treatment Human blood was obtained following informed consent and was collected...isolated RNA with 2.5 N lithium chloride (LiCI). 9 For subsequent RT-PCR studies, this method was used. The dose dependence of LPS stimulation of TNF...copy number was 10-fold lower than that used in the dose -response IL-8 PCR, and thus the target in the cDNA was estimated to be roughly ten times

  2. Differentiation and cytokine synthesis of human alveolar osteoblasts compared to osteoblast-like cells (MG63) in response to titanium surfaces.

    PubMed

    Rausch-fan, Xiaohui; Qu, Zhe; Wieland, Marco; Matejka, Michael; Schedle, Andreas

    2008-01-01

    The aim of this study was to investigate the influence of different implant surface topographies and chemistries on the expression of differentiation/proliferation markers on MG63 cells and primary human alveolar osteoblasts. Hydrophobic acid-etched (A) and hydrophobic coarse-grit-blasted, acid-etched (SLA) surfaces and hydrophilic acid-etched (modA) and hydrophilic coarse-grit-blasted (modSLA) surfaces were produced. Thereby, modA and modSLA surfaces were rinsed under nitrogen protection and stored in a sealed glass tube containing isotonic NaCl solution at pH 4-6. Tissue culture plates without specimens served as controls. The behavior of MG63 cells and primary human alveolar osteoblasts (AOB) grown on all surfaces was compared through determination of alkaline phosphatase (ALP) activity, cell proliferation ((3)H-thymidin incorporation, MTT colorimetric assay) and expression of osteocalcin (OC), osteoprotegerin (OPG), transforming growth factor-beta1 (TGF-beta(1)) and vascular endothelial growth factor (VEGF), detected with commercial available test kits. Proliferation of MG63 and primary cells was highest on controls, followed by A surfaces, modA and SLA surfaces being almost on the same level and lowest on modSLA surfaces. modSLA surfaces exhibited highest ALP and OC production, followed by SLA, modA and A surfaces. Proliferation and OC production were comparable for MG63 cells and AOB. OPG, TGF-beta(1) and VEGF produced on primary cells showed a slightly different rank order on different surfaces compared to MG63 cells. modSLA still showed the highest production of OPG, TGF-beta(1) and VEGF, but was followed by modA, SLA and A. Statistical significance was checked by ANOVA (p<0.0035). MG63 cells and primary human alveolar osteoblasts showed similar proliferation and differentiation characteristics on different titanium surfaces. Only modA surfaces showed enhanced expression of OPG, TGF-beta(1) and VEGF on MG63 cells compared to primary human alveolar

  3. Liver X receptor is a key regulator of cytokine release in human monocytes.

    PubMed

    Myhre, Anders E; Agren, Joanna; Dahle, Maria K; Tamburstuen, Margareth V; Lyngstadaas, Ståle P; Collins, A Jon L; Foster, Simon J; Thiemermann, Christoph; Aasen, Ansgar O; Wang, Jacob E

    2008-04-01

    Aberrant regulation of innate immune responses and uncontrolled cytokine bursts are hallmarks of sepsis and endotoxemia. Activation of the nuclear liver X receptor (LXR) was recently demonstrated to suppress inflammatory genes. Our aim was to investigate the expression of LXR in human monocytes under normal and endotoxemic conditions and to study the influence of LXR activation on endotoxin-induced cytokine synthesis and release. Adherent human monocytes or whole blood were pretreated with a synthetic LXR agonist (3-{3-[(2-chloro-3-trifluoromethyl-benzyl)-(2,2-diphenyl-ethyl)-amino]-propoxy}-phenyl)-acetic acid) and subsequently challenged with LPS (from Escherichia coli) or peptidoglycan (from Staphylococcus aureus). Cytokine release was assessed by a Multiplex antibody bead kit, and cytokine mRNA levels were measured by real-time reverse-transcriptase-polymerase chain reaction. We found that LXRalpha mRNA was up-regulated in CD14+ monocytes in LPS-challenged blood, whereas LXRbeta mRNA was not altered. Addition of 3-{3-[(2-chloro-3-trifluoromethyl-benzyl)-(2,2-diphenyl-ethyl)-amino]-propoxy}-phenyl)-acetic acid to monocytes suppressed the LPS-induced release of IL-1beta, IL-6, IL-8, IL-10, IL-12p40, TMF-alpha, macrophage inflammatory protein 1alpha, macrophage inflammatory protein 1beta, and monocyte chemoattractant protein 1 in a concentration-dependent manner. Surprisingly, an accompanying decrease in cytokine mRNA accumulation was not observed. The suppressed cytokine release could not be explained by a diminished transport of mRNA out of the nucleus or a decreased secretion of cytokines. We propose that LXR is a key regulator of cytokine release in LPS-challenged human monocytes, possibly by interfering with translational events.

  4. The Vitronectin Receptor and its Associated CD47 Molecule Mediates Proinflammatory Cytokine Synthesis in Human Monocytes by Interaction with Soluble CD23

    PubMed Central

    Hermann, P.; Armant, M.; Brown, E.; Rubio, M.; Ishihara, H.; Ulrich, D.; Caspary, R.G.; Lindberg, F.P.; Armitage, R.; Maliszewski, C.; Delespesse, G.; Sarfati, M.

    1999-01-01

    The vitronectin receptor, αvβ3 integrin, plays an important role in tumor cell invasion, angiogenesis, and phagocytosis of apoptotic cells. CD47, a member of the multispan transmembrane receptor family, physically and functionally associates with vitronectin receptor (VnR). Although vitronectin (Vn) is not a ligand of CD47, anti-CD47 and β3 mAbs suppress Vn, but not fibronectin (Fn) binding and function. Here, we show that anti-CD47, anti-β3 mAb and Vn, but not Fn, inhibit sCD23-mediated proinflammatory function (TNF-α, IL-12, and IFN-γ release). Surprisingly, anti-CD47 and β3 mAbs do not block sCD23 binding to αv+β3+ T cell lines, whereas Vn and an αv mAb (clone AMF7) do inhibit sCD23 binding, suggesting the VnR complex may be a functional receptor for sCD23. sCD23 directly binds αv+β3+/CD47− cell lines, but coexpression of CD47 increases binding. Moreover, sCD23 binds purified αv protein and a single human αv chain CHO transfectant. We conclude that the VnR and its associated CD47 molecule may function as a novel receptor for sCD23 to mediate its proinflammatory activity and, as such, may be involved in the inflammatory process of the immune response. PMID:10037797

  5. Regulation of Human Helper T Cell Subset Differentiation by Cytokines

    PubMed Central

    Schmitt, Nathalie; Ueno, Hideki

    2015-01-01

    Since the discovery of Th1 and Th2 cells in the late 80’s, the family of effector CD4+ helper T (Th) cell subsets has expanded. The differentiation of naïve CD4+ T cells is largely determined when they interact with dendritic cells in lymphoid organs, and cytokines play a major role in the regulation of Th differentiation in the early stages. Recent studies show that the developmental mechanism of certain Th subsets is not fully shared between mice and humans. Here we will review recent discoveries on the roles of cytokines in the regulation of Th differentiation in humans, and discuss the differences between mice and humans in the developmental mechanisms of several Th subsets, including Th17 cells and T follicular helper (Tfh) cells. We propose that the differentiation of human Th subsets is largely regulated by the three cytokines, IL-12, IL-23, and TGF-β. PMID:25879814

  6. Stabilization of a hydrophobic recombinant cytokine by human serum albumin.

    PubMed

    Hawe, Andrea; Friess, Wolfgang

    2007-11-01

    The objective was to evaluate the impact of pH and NaCl content on aggregation, particle formation, and solubility of a hydrophobic recombinant human cytokine in formulations with human serum albumin (HSA) as stabilizing excipient. While cytokine-HSA formulations were stable at physiological pH, a tremendous increase in turbidity at pH 5.0, close to the isoelectric point of HSA was caused by a partially irreversible precipitation. By dynamic light scattering (DLS), disc centrifugation, atomic force microscopy (AFM), and light obscuration it could be shown that the turbidity was mainly caused by particles larger than 120 nm. SDS-PAGE provided evidence that the precipitation at pH 5.0 was mainly caused by the cytokine. The HSA-stabilizers Na-octanoate and Na-N-acetyltryptophante were less effective in preventing the turbidity increase of unstabilized-HSA compared to NaCl. The interactions between HSA and cytokine were weakened by NaCl, as determined by fluorescence spectroscopy. The positive effect of NaCl on the formulation could be attributed to a direct stabilization of HSA and weaker interactions between HSA and the cytokine, which in consequence provided an overall stabilization of the cytokine. Copyright 2007 Wiley-Liss, Inc.

  7. Human colonic intraepithelial lymphocytes regulate the cytokines produced by lamina propria mononuclear cells

    PubMed Central

    Dehennin, J. P.; Li, Li; Sibille, C.; Geubel, A.; Vaerman, J. P.

    1997-01-01

    Using an in vitro autologous human system, the immunomodulatory function of colonic intraepithelial lymphocytes (IEL) on cytokine production by lamina propria mononuclear cells (LPMNC) has been investigated. In contrast to LPMNC, colonic IEL produced only low amounts of IL-10, interferon-γ and interleukin-2. However, co-culture experiments (IEL + LPMNC) have shown that IEL can enhance the PHA-induced synthesis of IL-2 and interferon-γ, but not IL-10 by LPMNC. Using a transwell filter culture system apparatus, this effect was shown not to require a cell-to-cell interaction. Thus, IEL in vitro may modulate the cytokine synthesis of LPMNC, through the production of soluble factors. This may prove highly relevant in the in vivo immune activation of the gastrointestinal mucosa. PMID:18472843

  8. Collection of aerosolized human cytokines using Teflon® filters.

    PubMed

    McKenzie, Jennifer H; McDevitt, James J; Fabian, M Patricia; Hwang, Grace M; Milton, Donald K

    2012-01-01

    Collection of exhaled breath samples for the analysis of inflammatory biomarkers is an important area of research aimed at improving our ability to diagnose, treat and understand the mechanisms of chronic pulmonary disease. Current collection methods based on condensation of water vapor from exhaled breath yield biomarker levels at or near the detection limits of immunoassays contributing to problems with reproducibility and validity of biomarker measurements. In this study, we compare the collection efficiency of two aerosol-to-liquid sampling devices to a filter-based collection method for recovery of dilute laboratory generated aerosols of human cytokines so as to identify potential alternatives to exhaled breath condensate collection. Two aerosol-to-liquid sampling devices, the SKC® Biosampler and Omni 3000™, as well as Teflon® filters were used to collect aerosols of human cytokines generated using a HEART nebulizer and single-pass aerosol chamber setup in order to compare the collection efficiencies of these sampling methods. Additionally, methods for the use of Teflon® filters to collect and measure cytokines recovered from aerosols were developed and evaluated through use of a high-sensitivity multiplex immunoassay. Our results show successful collection of cytokines from pg/m(3) aerosol concentrations using Teflon® filters and measurement of cytokine levels in the sub-picogram/mL concentration range using a multiplex immunoassay with sampling times less than 30 minutes. Significant degradation of cytokines was observed due to storage of cytokines in concentrated filter extract solutions as compared to storage of dry filters. Use of filter collection methods resulted in significantly higher efficiency of collection than the two aerosol-to-liquid samplers evaluated in our study. The results of this study provide the foundation for a potential new technique to evaluate biomarkers of inflammation in exhaled breath samples.

  9. Collection of Aerosolized Human Cytokines Using Teflon® Filters

    PubMed Central

    McKenzie, Jennifer H.; McDevitt, James J.; Fabian, M. Patricia; Hwang, Grace M.; Milton, Donald K.

    2012-01-01

    Background Collection of exhaled breath samples for the analysis of inflammatory biomarkers is an important area of research aimed at improving our ability to diagnose, treat and understand the mechanisms of chronic pulmonary disease. Current collection methods based on condensation of water vapor from exhaled breath yield biomarker levels at or near the detection limits of immunoassays contributing to problems with reproducibility and validity of biomarker measurements. In this study, we compare the collection efficiency of two aerosol-to-liquid sampling devices to a filter-based collection method for recovery of dilute laboratory generated aerosols of human cytokines so as to identify potential alternatives to exhaled breath condensate collection. Methodology/Principal Findings Two aerosol-to-liquid sampling devices, the SKC® Biosampler and Omni 3000™, as well as Teflon® filters were used to collect aerosols of human cytokines generated using a HEART nebulizer and single-pass aerosol chamber setup in order to compare the collection efficiencies of these sampling methods. Additionally, methods for the use of Teflon® filters to collect and measure cytokines recovered from aerosols were developed and evaluated through use of a high-sensitivity multiplex immunoassay. Our results show successful collection of cytokines from pg/m3 aerosol concentrations using Teflon® filters and measurement of cytokine levels in the sub-picogram/mL concentration range using a multiplex immunoassay with sampling times less than 30 minutes. Significant degradation of cytokines was observed due to storage of cytokines in concentrated filter extract solutions as compared to storage of dry filters. Conclusions Use of filter collection methods resulted in significantly higher efficiency of collection than the two aerosol-to-liquid samplers evaluated in our study. The results of this study provide the foundation for a potential new technique to evaluate biomarkers of inflammation in

  10. Host and Environmental Factors Influencing Individual Human Cytokine Responses.

    PubMed

    Ter Horst, Rob; Jaeger, Martin; Smeekens, Sanne P; Oosting, Marije; Swertz, Morris A; Li, Yang; Kumar, Vinod; Diavatopoulos, Dimitri A; Jansen, Anne F M; Lemmers, Heidi; Toenhake-Dijkstra, Helga; van Herwaarden, Antonius E; Janssen, Matthijs; van der Molen, Renate G; Joosten, Irma; Sweep, Fred C G J; Smit, Johannes W; Netea-Maier, Romana T; Koenders, Mieke M J F; Xavier, Ramnik J; van der Meer, Jos W M; Dinarello, Charles A; Pavelka, Norman; Wijmenga, Cisca; Notebaart, Richard A; Joosten, Leo A B; Netea, Mihai G

    2016-11-03

    Differences in susceptibility to immune-mediated diseases are determined by variability in immune responses. In three studies within the Human Functional Genomics Project, we assessed the effect of environmental and non-genetic host factors of the genetic make-up of the host and of the intestinal microbiome on the cytokine responses in humans. We analyzed the association of these factors with circulating mediators and with six cytokines after stimulation with 19 bacterial, fungal, viral, and non-microbial metabolic stimuli in 534 healthy subjects. In this first study, we show a strong impact of non-genetic host factors (e.g., age and gender) on cytokine production and circulating mediators. Additionally, annual seasonality is found to be an important environmental factor influencing cytokine production. Alpha-1-antitrypsin concentrations partially mediate the seasonality of cytokine responses, whereas the effect of vitamin D levels is limited. The complete dataset has been made publicly available as a comprehensive resource for future studies. PAPERCLIP. Copyright © 2016 Elsevier Inc. All rights reserved.

  11. Comparison of serum and cell-specific cytokines in humans.

    PubMed

    Jason, J; Archibald, L K; Nwanyanwu, O C; Byrd, M G; Kazembe, P N; Dobbie, H; Jarvis, W R

    2001-11-01

    Cytokines function at the cellular, microenvironmental level, but human cytokine assessment is most commonly done at the macro level, by measuring serum cytokines. The relationships between serum and cellular cytokines, if there are any, are undefined. In a study of hospitalized patients in Malawi, we compared cytometrically assessed, cell-specific cytokine data to serum interleukin 2 (IL-2), IL-4, IL-6, IL-8, IL-10, gamma interferon (IFN-gamma), and tumor necrosis factor alpha (TNF-alpha) levels in 16 children and 71 (IL-2, -4, -6, -10) or 159 (IL-8, IFN-gamma, and TNF-alpha) adults, using Wilcoxon rank sum tests and Pearson's (r(p)) and Spearman's (r(s)) rank correlations. For the entire study group, correlations between identical serum and cellular cytokines mainly involved IL-8 and IFN-gamma, were few, and were weakly positive (r < 0.40). Blood culture-positive persons had the most and strongest correlations, including those between serum IL-2 levels and the percentages of lymphocytes spontaneously making IL-2 (r(s) = +0.74), serum IL-8 levels and the percentages of lymphocytes spontaneously making IL-8 (r(p) = +0.66), and serum IL-10 levels and the percentages of CD8(+) T cells making TNF-alpha (r(p) = +0.89). Human immunodeficiency virus (HIV)-positive persons had the next largest number of correlations, including several serum IL-8 level correlations, correlation of serum IL-10 levels with the percentages of lymphocytes producing induced IL-10 (r(s) = +0.36), and correlation of serum IFN-gamma levels and the percentages of lymphocytes spontaneously making both IL-6 and IFN-gamma in the same cell (r(p) = +0.59). HIV-negative, malaria smear-positive, and pediatric patients had few significant correlations; for the second and third of these subgroups, serum IL-8 level was correlated with the percentage of CD8(-) T cells producing induced IL-8 (r(s) = +0.40 and r(s) = +0.56, respectively). Thus, the strength of associations between serum and cellular cytokines

  12. Analysis of inflammatory cytokines in human blood, breath ...

    EPA Pesticide Factsheets

    A change in the expression of cytokines in human biological media indicates an inflammatory response to external stressors and reflects an early step along the adverse outcome pathway (AOP) for various health endpoints. To characterize and interpret this inflammatory response, methodology was developed for measuring a suite of 10 different cytokines in human blood, exhaled breath condensate (EBC), and urine using an electrochemiluminescent multiplex Th1/Th2 cytokine immunoassay platform. Measurement distributions and correlations for eight interleukins (IL) (1β, 2, 4, 5, 8, 10, 12p70 and 13), interferon-γ (IFN-γ), and tumor necrosis factor-α (TNF-α) were evaluated using 90 blood plasma, 77 EBC, and 400 urine samples collected from nominally healthy adults subjects in North Carolina in 2008-2012. The in vivo results show that there is sufficient sensitivity for characterizing all 10 cytokines at levels of 0.05-0.10 ρg/ml with a dynamic range up to 100 ng/ml across all three of these biological media. The measured in vivo results also show that the duplicate analysis of blood, EBC and urine samples have average estimated fold ranges of 2.21, 3.49, and 2.50, respectively, which are similar to the mean estimated fold range (2.88) for the lowest concentration (0.610ρg/ml) from a series of spiked control samples; the cytokine method can be used for all three biological media. Nine out of the 10 cytokines measured in EBC were highly correlated within one a

  13. In situ expression of cytokines in human heart allografts.

    PubMed

    Van Hoffen, E; Van Wichen, D; Stuij, I; De Jonge, N; Klöpping, C; Lahpor, J; Van Den Tweel, J; Gmelig-Meyling, F; De Weger, R

    1996-12-01

    Although allograft rejection, the major complication of human organ transplantation, has been extensively studied, little is known about the exact cellular localization of the cytokine expression inside the graft during rejection. Therefore, we used in situ hybridization and immunohistochemistry to study local cytokine mRNA and protein expression in human heart allografts, in relation to the phenotypical characteristics of the cellular infiltrate. Clear expression of mRNA for interleukin (IL)-6, IL-8, IL-9, and IL-10 and weak expression for IL-2, IL-4, IL-5, and tumor necrosis factor (TNF)-alpha was detected in biopsies exhibiting high rejection grades (grade 3A/B). Also at lower grades of rejection, mRNA for IL-6 and IL-9 was present. Some mRNA for IL-1 beta, TNF-beta, and interferon (IFN)-gamma was detected in only a few biopsies. Using immunohistochemistry, IL-2, IL-3, and IL-10 protein was detected in biopsies with high rejection grades, whereas few cells expressed IL-6, IL-8, and IFN-gamma. In biopsies with lower grades of rejection, a weaker expression of these cytokines was observed. IL-4 was hardly detected in any of the biopsies. The level of IL-12 expression was equal in all biopsies. Although mRNA expression of several cytokines was expressed at a low level compared with the protein level of those cytokines, there was a good correlation between localization of cytokine mRNA and protein. Expression of IL-2, IL-4, IL-5, TNF-alpha, and IFN-gamma was mainly detected in lymphocytes. IL-3, IL-6, IL-10, and IL-12 were not detected or not only detected in lymphocytes but also in other stromal elements (eg, macrophages). Macrophage production of IL-3 and IL-12 was confirmed by immunofluorescent double labeling with CD68. We conclude that cardiac allograft rejection is not simply regulated by T helper cell cytokine production, but other intragraft elements contribute considerably to this process.

  14. Human malarial disease: a consequence of inflammatory cytokine release

    PubMed Central

    Clark, Ian A; Budd, Alison C; Alleva, Lisa M; Cowden, William B

    2006-01-01

    Malaria causes an acute systemic human disease that bears many similarities, both clinically and mechanistically, to those caused by bacteria, rickettsia, and viruses. Over the past few decades, a literature has emerged that argues for most of the pathology seen in all of these infectious diseases being explained by activation of the inflammatory system, with the balance between the pro and anti-inflammatory cytokines being tipped towards the onset of systemic inflammation. Although not often expressed in energy terms, there is, when reduced to biochemical essentials, wide agreement that infection with falciparum malaria is often fatal because mitochondria are unable to generate enough ATP to maintain normal cellular function. Most, however, would contend that this largely occurs because sequestered parasitized red cells prevent sufficient oxygen getting to where it is needed. This review considers the evidence that an equally or more important way ATP deficency arises in malaria, as well as these other infectious diseases, is an inability of mitochondria, through the effects of inflammatory cytokines on their function, to utilise available oxygen. This activity of these cytokines, plus their capacity to control the pathways through which oxygen supply to mitochondria are restricted (particularly through directing sequestration and driving anaemia), combine to make falciparum malaria primarily an inflammatory cytokine-driven disease. PMID:17029647

  15. Cytokines and Cytokine Profiles in Human Autoimmune Diseases and Animal Models of Autoimmunity

    PubMed Central

    Kunz, Manfred; Ibrahim, Saleh M.

    2009-01-01

    The precise pathomechanisms of human autoimmune diseases are still poorly understood. However, a deepened understanding of these is urgently needed to improve disease prevention and early detection and guide more specific treatment approaches. In recent years, many new genes and signalling pathways involved in autoimmunity with often overlapping patterns between different disease entities have been detected. Major contributions were made by experiments using DNA microarray technology, which has been used for the analysis of gene expression patterns in chronic inflammatory and autoimmune diseases, among which were rheumatoid arthritis, systemic lupus erythematosus, psoriasis, systemic sclerosis, multiple sclerosis, and type-1 diabetes. In systemic lupus erythematosus, a so-called interferon signature has been identified. In psoriasis, researchers found a particular immune signalling cluster. Moreover the identification of a new subset of inflammatory T cells, so-called Th17 T cells, secreting interleukin (IL)-17 as one of their major cytokines and the identification of the IL-23/IL-17 axis of inflammation regulation, have significantly improved our understanding of autoimmune diseases. Since a plethora of new treatment approaches using antibodies or small molecule inhibitors specifically targeting cytokines, cellular receptors, or signalling mechanisms has emerged in recent years, more individualized treatment for affected patients may be within reach in the future. PMID:19884985

  16. Human Mesenchymal Stem Cells Protect Human Islets from Pro-Inflammatory Cytokines

    PubMed Central

    Yeung, Telford Y.; Seeberger, Karen L.; Kin, Tatsuya; Adesida, Adetola; Jomha, Nadr; Shapiro, A. M. James; Korbutt, Gregory S.

    2012-01-01

    Transplantation of human islets is an attractive alternative to daily insulin injections for patients with type 1 diabetes. However, the majority of islet recipients lose graft function within five years. Inflammation is a primary contributor to graft loss, and inhibiting pro-inflammatory cytokine activity can reverse inflammation mediated dysfunction of islet grafts. As mesenchymal stem cells (MSCs) possess numerous immunoregulatory properties, we hypothesized that MSCs could protect human islets from pro-inflammatory cytokines. Five hundred human islets were co-cultured with 0.5 or 1.0×106 human MSCs derived from bone marrow or pancreas for 24 hours followed by 48 hour exposure to interferon-γ, tumor necrosis factor-α and interleukin 1β. Controls include islets cultured alone (± cytokines) and with human dermal fibroblasts (± cytokines). For all conditions, glucose stimulated insulin secretion (GSIS), total islet cellular insulin content, islet β cell apoptosis, and potential cytoprotective factors secreted in the culture media were determined. Cytokine exposure disrupted human islet GSIS based on stimulation index and percentage insulin secretion. Conversely, culture with 1.0×106 bMSCs preserved GSIS from cytokine treated islets. Protective effects were not observed with fibroblasts, indicating that preservation of human islet GSIS after exposure to pro-inflammatory cytokines is MSC dependent. Islet β cell apoptosis was observed in the presence of cytokines; however, culture of bMSCs with islets prevented β cell apoptosis after cytokine treatment. Hepatocyte growth factor (HGF) as well as matrix metalloproteinases 2 and 9 were also identified as putative secreted cytoprotective factors; however, other secreted factors likely play a role in protection. This study, therefore, demonstrates that MSCs may be beneficial for islet engraftment by promoting cell survival and reduced inflammation. PMID:22666480

  17. Synthesis of cytokines during tumour development in mice immunized with the mycobacterial antigen complex A60.

    PubMed

    Maes, H; Cocito, C

    1996-10-01

    The authors have previously reported on the ability of A60, an immunodominant antigenic complex of Mycobacterium bovis BCG, to prevent cancer development in mice challenged with EMT 6 tumour cells. Such effect proved to rely on neoplastic cell lysis by cytolytic T lymphocytes and activated macrophages. The involvement of cytokines in triggering the immune response leading to tumour rejection is analysed in the present work. The synthesis of IL-2, IFN-gamma and TNF-alpha was strongly increased in A60-primed mice. Cancer development depressed the blood levels of these three cytokines. In vitro cultures of lymphocytes from lymph nodes and blood of A60-primed mice produced higher levels of these cytokines in the presence of A60, as compared to cultures lacking A60. Such effect was inhibited by co-incubation of lymphocytes with EMT 6 tumour cells In vitro cultures of macrophages yielded higher levels of TNF-alpha in the presence of A60 and co-incubation of these cells with EMT 6 tumour cells also inhibited TNF-alpha production. The enhanced synthesis of IL-2 and IFN-gamma, which promote activation of cytolytic T lymphocytes and macrophages, accounts for the increased tumour cell lysis induced in vivo by A60. The A60-promoted synthesis of TNF-alpha is partly responsible for the latter effect. The inhibitory action of EMT-6 tumour cells on cytokine synthesis is a powerful mechanism of tumour escape from the immune system's control.

  18. Th17 cytokines induce pro-fibrotic cytokines release from human eosinophils

    PubMed Central

    2013-01-01

    Background Subepithelial fibrosis is one of the most critical structural changes affecting bronchial airway function during asthma. Eosinophils have been shown to contribute to the production of pro-fibrotic cytokines, TGF-β and IL-11, however, the mechanism regulating this process is not fully understood. Objective In this report, we investigated whether cytokines associated with inflammation during asthma may induce eosinophils to produce pro-fibrotic cytokines. Methods Eosinophils were isolated from peripheral blood of 10 asthmatics and 10 normal control subjects. Eosinophils were stimulated with Th1, Th2 and Th17 cytokines and the production of TGF-β and IL-11 was determined using real time PCR and ELISA assays. Results The basal expression levels of eosinophil derived TGF-β and IL-11 cytokines were comparable between asthmatic and healthy individuals. Stimulating eosinophils with Th1 and Th2 cytokines did not induce expression of pro-fibrotic cytokines. However, stimulating eosinophils with Th17 cytokines resulted in the enhancement of TGF-β and IL-11 expression in asthmatic but not healthy individuals. This effect of IL-17 on eosinophils was dependent on p38 MAPK activation as inhibiting the phosphorylation of p38 MAPK, but not other kinases, inhibited IL-17 induced pro-fibrotic cytokine release. Conclusions Th17 cytokines might contribute to airway fibrosis during asthma by enhancing production of eosinophil derived pro-fibrotic cytokines. Preventing the release of pro-fibrotic cytokines by blocking the effect of Th17 cytokines on eosinophils may prove to be beneficial in controlling fibrosis for disorders with IL-17 driven inflammation such as allergic and autoimmune diseases. PMID:23496774

  19. Th17 cytokines induce pro-fibrotic cytokines release from human eosinophils.

    PubMed

    Al-Muhsen, Saleh; Letuve, Severine; Vazquez-Tello, Alejandro; Pureza, Mary Angeline; Al-Jahdali, Hamdan; Bahammam, Ahmed S; Hamid, Qutayba; Halwani, Rabih

    2013-03-13

    Subepithelial fibrosis is one of the most critical structural changes affecting bronchial airway function during asthma. Eosinophils have been shown to contribute to the production of pro-fibrotic cytokines, TGF-β and IL-11, however, the mechanism regulating this process is not fully understood. In this report, we investigated whether cytokines associated with inflammation during asthma may induce eosinophils to produce pro-fibrotic cytokines. Eosinophils were isolated from peripheral blood of 10 asthmatics and 10 normal control subjects. Eosinophils were stimulated with Th1, Th2 and Th17 cytokines and the production of TGF-β and IL-11 was determined using real time PCR and ELISA assays. The basal expression levels of eosinophil derived TGF-β and IL-11 cytokines were comparable between asthmatic and healthy individuals. Stimulating eosinophils with Th1 and Th2 cytokines did not induce expression of pro-fibrotic cytokines. However, stimulating eosinophils with Th17 cytokines resulted in the enhancement of TGF-β and IL-11 expression in asthmatic but not healthy individuals. This effect of IL-17 on eosinophils was dependent on p38 MAPK activation as inhibiting the phosphorylation of p38 MAPK, but not other kinases, inhibited IL-17 induced pro-fibrotic cytokine release. Th17 cytokines might contribute to airway fibrosis during asthma by enhancing production of eosinophil derived pro-fibrotic cytokines. Preventing the release of pro-fibrotic cytokines by blocking the effect of Th17 cytokines on eosinophils may prove to be beneficial in controlling fibrosis for disorders with IL-17 driven inflammation such as allergic and autoimmune diseases.

  20. A new family of synthetic diterpenes that regulates cytokine synthesis by inhibiting IkappaBalpha phosphorylation.

    PubMed

    Chao, Ta-Hsiang; Lam, Thanh; Vong, Binh G; Través, Paqui G; Hortelano, Sonsoles; Chowdhury, Chinmay; Bahjat, F Rena; Lloyd, G Kenneth; Moldawer, Lyle L; Boscá, Lisardo; Palladino, Michael A; Theodorakis, Emmanuel A

    2005-01-01

    The synthesis and the biological evaluation of a new family diterpenes are presented. The synthetic studies were inspired by the structural framework of acanthoic acid (1) and yielded a family of compounds that were evaluated as anti-inflammatory agents. Among them, compounds 2, 10, 12, and 16 exhibited a very low nonspecific cytotoxicity and inhibited the synthesis of TNF-alpha with greater than 65 % efficacy at low micromolar concentrations. Cytokine-specificity studies revealed that these compounds also inhibited the synthesis of the proinflammatory cytokines IL-1beta and IL-6, while inhibition of IL-1ra and IL-8 synthesis was marginal and only occurred at high concentrations. Further studies, through EMSA and Western blot analyses, indicated that these compounds decreased the extent of phosphorylation of IkappaBalpha; this suggests that they exert their anti-inflammatory profile by inhibiting NF-kappaB-mediated cytokine synthesis. These findings imply that these diterpenes represent promising leads for the development of novel anti-inflammatory agents.

  1. Selective regulation of T cell IL-5 synthesis by OM-01, JTE-711 and p38 MAP kinase inhibitor: independent control of Th2 cytokines, IL-4 and IL-5.

    PubMed

    Mori, A; Okudaira, H; Kobayashi, N; Akiyama, K

    2001-01-01

    Helper T cells are involved in chronic eosinophilic inflammation. Control of cytokine production seems to be an effective management. The effect of nonactin, SB203580, a p38 MAP kinase inhibitor, and JTE-711 on the cytokine production of allergen-specific T cell clones was determined. The effect of nonactin on the airway eosinophilia was investigated using murine asthma model. Nonactin suppressed IL-5 synthesis by human Th cells in a dose-dependent manner without affecting IL-2 or IL-4 synthesis, and still significantly suppressed murine airway eosinophilia induced by the antigen inhalation. SB203580 and JTE-711 also selectively inhibited IL-5 synthesis in vitro. Synthesis of IL-5 by human Th cells can be differentially regulated from that of other major T cell cytokines. The in vivo effects of selective IL-5 synthesis inhibitors suggest that IL-5 is the reasonable target for the regulation of allergic disorders accompanied by eosinophilic inflammation. Copyright 2001 S. Karger AG, Basel

  2. 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.

  3. Interleukin 17, a nitric oxide-producing cytokine with a peroxynitrite-independent inhibitory effect on proteoglycan synthesis.

    PubMed

    Pacquelet, Sandrine; Presle, Nathalie; Boileau, Christelle; Dumond, Hélène; Netter, Patrick; Martel-Pelletier, Johanne; Pelletier, Jean-Pierre; Terlain, Bernard; Jouzeau, Jean-Yves

    2002-12-01

    To compare the potency of 2 cytokines, interleukin 17 (IL-17) and IL-1beta, on rat cartilage proteoglycan synthesis with special attention to nitric oxide (NO) and peroxynitrite formation. Chondrocytes in alginate beads were stimulated with human recombinant (rh) IL-17 (0.03 to 300.0 ng/ml) and/or rhIL-1beta (0.25 to 25.0 ng/ml) in the presence or not of L-NMMA or CuDips. Alternatively, rats were injected with either IL-17 (10.0 micro g) or IL-1beta (1.0 micro g) into each knee joint. NO concentrations were determined by a spectrofluorimetric assay, proteoglycan synthesis by 35SO4-2 incorporation, peroxynitrite generation by immunostaining for 3-nitrotyrosine, and IL-1beta mRNA expression by reverse transcription-polymerase chain reaction. IL-17 inhibited proteoglycan synthesis and increased NO production, both in vitro and in vivo, without inducing expression of IL-1beta mRNA in cartilage. Additive effects were observed when IL-17 was combined with low concentrations of IL-1. Surprisingly, a similar NO synthesis between IL-1 and IL-17 led to a less suppressive effect of IL-17 on cartilage anabolism than with IL-1. Both in vitro and in vivo, peroxynitrite formation was extensive with IL-1beta, but negligible or nonexistent with IL-17. L-NMMA and CuDips completely corrected the suppressive effect of IL-1beta on proteoglycan synthesis, unlike with IL-17. These data showed that NO is weakly involved in the IL-17 mediated inhibition of proteoglycan synthesis in rat. NO overload may not be predictive of any inhibitory effect on cartilage anabolism, but instead superoxide is a key regulator of NO contribution to chondrocyte dysfunction. Since IL-17 is a NO-producing cytokine with additive effects when combined with IL-1, it may play a pivotal role in cartilage destruction during rheumatoid arthritis, for which infiltrating cells produce high levels of superoxide and proinflammatory cytokines.

  4. LIGHT is associated with hypertriglyceridemia in obese subjects and increased cytokine secretion from cultured human adipocytes.

    PubMed

    Bassols, J; Moreno-Navarrete, J M; Ortega, F; Ricart, W; Fernandez-Real, J M

    2010-01-01

    LIGHT (lymphotoxin-like inducible protein that competes with glycoprotein D for herpesvirus entry on T cells) is a member of the tumor necrosis factor (TNF) family, primarily expressed in lymphocytes, which was associated with the induction of pro-inflammatory cytokines and alterations of lipid homeostasis in animal models. We aimed to analyze whether LIGHT has a role in the human obesity-associated inflammatory status. The association between circulating LIGHT concentrations and clinical variables was studied in 190 subjects with different degrees of obesity and glucose tolerance. The expression and release of 21 different cytokines, and the expression of genes involved in lipid metabolism were also evaluated after stimulation with LIGHT in cultured human differentiated adipocytes. Serum LIGHT concentrations positively associated with body mass index (BMI), fat mass, glycated hemoglobin and fasting triglycerides, and negatively with high-density lipoprotein cholesterol. Circulating LIGHT concentrations were significantly increased in morbidly obese subjects and in patients with type 2 diabetes. LIGHT induced the secretion of several cytokines and upregulated the expression and secretion of interleukin-6 (IL-6), IL-8, Growth Regulated Oncogene (GRO) and monocyte chemotactic protein-1 (MCP-1). These observations were concomitant with the activation of nuclear factor (NF)-kappaB signalling in human differentiated adipocytes. LIGHT also upregulated the expression and synthesis of its own receptor (herpesvirus entry mediator (HVEM)) and decreased the expression of peroxisome proliferator-activated receptor-gamma (PPAR-gamma) and fatty acid synthase. These data suggest that LIGHT may have a role in mediating chronic inflammation and alterations of lipid metabolism in obese subjects.

  5. Profiling of cytokines in human epithelial ovarian cancer ascites

    PubMed Central

    Matte, Isabelle; Lane, Denis; Laplante, Claude; Rancourt, Claudine; Piché, Alain

    2012-01-01

    Background The behavior of tumor cells is influenced by the composition of the surrounding tumor environment. The importance of ascites in ovarian cancer (OC) progression is being increasingly recognized. The characterization of soluble factors in ascites is essential to understand how this environment affects OC progression. The development of cytokine arrays now allows simultaneous measurement of multiple cytokines per ascites using a single array. Methods We applied a multiplex cytokine array technology that simultaneously measures the level of 120 cytokines in ascites from 10 OC patients. The ascites concentration of a subset (n = 5) of cytokines that was elevated based on the multiplex array was validated by commercially available ELISA. The ascites level of these 5 cytokines was further evaluated by ELISA in a cohort of 38 patients. Kaplan-Meier analysis was used to assess the association of cytokine expression with progression-free survival (PFS) in this cohort. Results We observed a wide variability of expression between different cytokines and levels of specific cytokines also varied in the 10 malignant ascites tested. Fifty-three (44%) cytokines were not detected in any of the 10 ascites. The level of several factors including, among others, angiogenin, angiopoietin-2, GRO, ICAM-1, IL-6, IL-6R, IL-8, IL-10, leptin, MCP-1, MIF NAP-2, osteprotegerin (OPG), RANTES, TIMP-2 and UPAR were elevated in most malignant ascites. Higher levels of OPG, IL-10 and leptin in OC ascites were associated with shorter PFS. IL-10 was shown to promote the anti-apoptotic activity of malignant ascites whereas OPG did not. Conclusion Our data demonstrated that there is a complex network of cytokine expression in OC ascites. Characterization of cytokine profiles in malignant ascites may provide information from which to prioritize key functional cytokines and understand the mechanism by which they alter tumor cells behavior. A better understanding of the cytokine network is

  6. Endocrine and cytokine responses in humans with pulmonary tuberculosis.

    PubMed

    Rey, Adriana Del; Mahuad, Carolina V; Bozza, Verónica V; Bogue, Cristina; Farroni, Miguel A; Bay, María Luisa; Bottasso, Oscar A; Besedovsky, Hugo O

    2007-02-01

    Endocrine responses during chronic infections such as lung tuberculosis are poorly characterized. Hormonal changes are likely to occur since some of the cytokines produced during this disease could affect endocrine mechanisms that, in turn, influence the course of infectious/inflammatory processes. A main purpose of this work was to study endocrine responses involving pituitary, adrenal, gonadal, and thyroid hormones in parallel to IFN-gamma, IL-10, and IL-6 levels in tuberculosis patients with different degree of pulmonary involvement. We have also studied whether products derived from peripheral immune cells obtained from the patients can affect the in vitro production of adrenal steroids. The population studied comprised HIV-negative newly diagnosed, untreated male patients with mild, moderate, and advanced lung tuberculosis, and matched, healthy controls. IFN-gamma, IL-10, and IL-6 levels were elevated in patients with tuberculosis. Dehydroepiandrosterone and testosterone levels were profoundly decreased and growth hormone levels were markedly elevated in patients, in parallel to modest increases in cortisol, estradiol, prolactin, and thyroid hormone concentrations. Supernatants of peripheral blood mononuclear cells obtained from the patients and stimulated in vitro with Mycobacterium tuberculosis antigens significantly inhibited dehydroepiandrosterone secretion by the human adrenal cell line NCI-H295-R. These results support the hypothesis that at least some of the endocrine changes observed in the patients may be mediated by endogenous cytokines. The endocrine profile of tuberculosis patients would favor a reduction of protective cell-mediated immunity and an exacerbation of inflammation leading to perpetuation of the lung injury and to the hypercatabolic condition that characterizes this disease.

  7. Chemotherapeutic drugs and human tumor cells cytokine network

    PubMed Central

    Levina, Vera; Su, Yunyun; Nolen, Brian; Liu, Xiaoning; Gordin, Yuri; Lee, Melissa; Lokshin, Anna; Gorelik, Elieser

    2008-01-01

    The ability of human tumor cell lines to produce various cytokines, chemokines, angiogenic and growth factors was investigated using Luminex multiplex technology. Media conditioned by tumor cells protected tumor cells from drug-induced apoptosis and stimulated tumor cell proliferation. Antibodies neutralizing IL-6, CXCL8, CCL2 and CCL5 blocked this stimulation. Treatment of tumor cells with doxorubicin and cisplatin resulted in a substantial increase in the production of IL-6, CXCL8, CCL2, CCL5, BFGF, G-CSF, and VEGF. This stimulation was associated with drug-induced activation of NF-κB, AP-1, AP-2, CREB, HIF-1, STAT-1, STAT-3, STAT-5 and ATF-2 transcription factors and up-regulation of IL-6, CXCL8, FGF-2, CSF-3 and CCL5 gene expression. Treatment of tumor cells with doxorubicin and antibodies neutralizing G-CSF, CCL2 or CCL5 had higher inhibitory effects than each modality used alone. These results indicate that chemokines and growth factors produced by tumor by binding to the cognate receptors on tumor and stroma cells could provide proliferative and anti-apoptotic signals helping tumor to escape drug-mediated destruction. Clinical studies showed that antibodies neutralizing VEGF (Avastin/ Bevacizumab) or blocking HER2/neu signaling (Herceptin/ Trastuzumab) could increase the efficacy of chemotherapy although these beneficial effects have been limited. It is possible that drug-stimulated production of growth and pro-angiogenic factors could counterbalance the effects of antibody therapy. In addition, numerous growth factors and chemokines share angiogenic and growth-stimulating properties, and thus reduction of a single factor is insufficient to completely block tumor growth. Thus, a broad disruption of tumor cytokine network is needed to further increase the efficacy of cancer therapy. PMID:18697197

  8. Modulation of cytokine expression in human macrophages by endocrine-disrupting chemical Bisphenol-A

    SciTech Connect

    Liu, Yanzhen; Mei, Chenfang; Liu, Hao; Wang, Hongsheng; Zeng, Guoqu; Lin, Jianhui; Xu, Meiying

    2014-09-05

    Highlights: • Effects of BPA on the cytokines expression of human macrophages were investigated. • BPA increased pro-inflammation cytokines TNF-α and IL-6 production. • BPA decreased anti-inflammation IL-10 and TGF-β production. • ERα/β/ERK/NF-κB signaling involved in BPA-mediated cytokines expression. - Abstract: Exposure to environmental endocrine-disrupting chemical Bisphenol-A (BPA) is often associated with dysregulated immune homeostasis, but the mechanisms remain unclear. In the present study, the effects of BPA on the cytokines responses of human macrophages were investigated. Treatment with BPA increased pro-inflammation cytokines tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) production, but decreased anti-inflammation cytokines interleukin-10 (IL-10) and transforming growth factor-β (TGF-β) production in THP1 macrophages, as well as in primary human macrophages. BPA effected cytokines expression through estrogen receptor α/β (ERα/β)-dependent mechanism with the evidence of ERα/β antagonist reversed the expression of cytokines. We also identified that activation of extracellular regulated protein kinases (ERK)/nuclear factor κB (NF-κB) signal cascade marked the effects of BPA on cytokines expression. Our results indicated that BPA effected inflammatory responses of macrophages via modulating of cytokines expression, and provided a new insight into the link between exposure to BPA and human health.

  9. Proinflammatory cytokines induce amelotin transcription in human gingival fibroblasts.

    PubMed

    Nakayama, Yohei; Takai, Hideki; Matsui, Sari; Matsumura, Hiroyoshi; Zhou, Liming; Kato, Ayako; Ganss, Bernhard; Ogata, Yorimasa

    2014-12-01

    Amelotin (AMTN) is a secreted protein transcribed predominantly during the maturation stage of enamel formation and localized in the junctional epithelium. We investigated differences in the levels of AMTN gene expression between non-inflamed gingiva and inflamed gingiva from patients with chronic periodontitis. Total RNAs were isolated from these tissues and their gene expression profiles were monitored by DNA microarray. The observed induction of AMTN mRNA in inflamed gingiva and cultured human gingival fibroblasts (HGF) was confirmed by real-time PCR. Transient transfection assays were performed using chimeric constructs of mouse AMTN gene promoter fragments linked to a luciferase reporter gene. Immunohistochemical localization of AMTN in inflamed and non-inflamed gingiva was assessed by immunohistochemistry. Among many differentially expressed genes, the level of AMTN mRNA was significantly increased in inflamed gingiva. Treatment of HGF with interleukin-1β (IL-1β), IL-6 and tumor necrosis factor-α (TNF-α) induced the expression of AMTN mRNA, and increased the luciferase activities of the AMTN promoter constructs. AMTN protein was detected in inflamed gingival connective tissue and junctional epithelium. These findings demonstrate that proinflammatory cytokines induce AMTN gene expression in human gingival fibroblasts and suggest a role for AMTN in gingival inflammation.

  10. Cytokine regulation of granulocyte-macrophage colony-stimulating factor (GM-CSF) production by human retinal pigment epithelial cells

    PubMed Central

    Crane, I J; Kuppner, M C; Mckillop-Smith, S; Wallace, C A; Forrester, J V

    1999-01-01

    GM-CSF is an important regulator of macrophage, granulocyte and dendritic cell behaviour and function. These cell types have been implicated in the retinal damage characteristic of endogenous posterior uveitis. Dendritic cells in the choroid have access to retinal antigens processed by the retinal pigment epithelial (RPE) cells of the blood–retinal barrier and are thought to be candidates for the presentation of antigen in uveoretinitis. We therefore investigated the production of GM-CSF and its regulation in human RPE cells. IL-1β, tumour necrosis factor-alpha (TNF-α) and transforming growth factor-beta (TGF-β) all stimulated GM-CSF production by RPE cells and a combination of these cytokines increased GM-CSF production over five-fold compared with that with the individual cytokines alone. Interferon-gamma (IFN-γ) rapidly down-regulated these responses. IFN-γ did not appear to be acting directly on IL-1β or via the synthesis of another protein. GM-CSF mRNA expression showed the same pattern of response to these cytokines, indicating transcriptional or pre-transcriptional regulation, and there was no evidence that IFN-γ was acting by destabilizing GM-CSF mRNA. These results are generally important in understanding the ways in which cytokine regulation differs between different cell types and also more specifically for determining ways in which a cytokine with a significant role in the development of autoimmune uveoretinitis may be manipulated. PMID:9933455

  11. 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

  12. Identification of three related human GRO genes encoding cytokine functions

    SciTech Connect

    Haskill, S.; Peace, A.; Morris, J.; Sporn, S.A. ); Anisowicz, A.; Lee, S.W.; Sager, R. ); Smith, T. ); Martin, G.; Ralph, P. )

    1990-10-01

    The product of the human GRO gene is a cytokine with inflammatory and growth-regulatory properties; GRO is also called MGSA for melanoma growth-stimulatory activity. The authors have identified two additional genes, GRO{beta} and GRO{gamma}, that share 90{percent} and 86{percent} identity at the deduced amino acid level with the original GRO{alpha} isolate. One amino acid substitution of proline in GRO{alpha} by leucine in GRO{beta} and GRO{gamma} leads to a large predicted change in protein conformation. Significant differences also exist in the 3' untranslated region, including different numbers of ATTTA repeats associated with mRNA instability. A 122-base-pair region in the 3' region is conserved among the three GRO genes, and a part of it is also conserved in the Chinese hamster genome, suggesting a role in regulation. DNA hybridization with oligonucleotide probes and partial sequence analysis of the genomic clones confirm that the three forms are derived from related but different genes. Only one chromosomal locus has been identified, at 4q21, by using a GRO{alpha} cDNA clone that hybridized to all three genes. Expression studies reveal tissue-specific regulation as well as regulation by specific inducing agents, including interleukin 1, tumor necrosis factor, phorbol 12-myristate 13-acetate, and lipopolysaccharide.

  13. Postoperative pain management and proinflammatory cytokines: animal and human studies.

    PubMed

    Shavit, Yehuda; Fridel, Keren; Beilin, Benzion

    2006-12-01

    The postoperative period is associated with neuroendocrine, metabolic, and immune alterations, which are the combined result of tissue damage, anesthesia, postoperative pain, and psychological stress. Limited evidence indicates that pain management in the postoperative period can affect the outcome of the surgery, reducing cardiac, pulmonary, and metabolic complications. Recent evidence indicates that pain and immune factors, especially proinflammatory cytokines, mutually interact and influence each other. A series of animal studies demonstrates that effective preemptive analgesia improved postoperative recovery, and this effect was enhanced by coadministration of IL-1ra together with the preemptive analgesics. Furthermore, preemptive analgesia attenuated surgery-induced PGE(2) production in the amygdala and the activation of the HPA axis. IL-1 signaling is required for the production of amygdala PGE(2) in response to surgical stress, and may thus affect the physiological and psychological aspects of surgical stress. These reports suggest that short-term effective analgesia can have long-lasting beneficial effects on surgery recovery. They further suggest that IL-1 blockade should be considered in the clinical management of pain associated with peripheral or nerve injury. Another series of human studies describes an interaction between the effectiveness of postoperative pain relief and surgery-associated immune alterations: In three separate studies, the more effective pain management technique was associated with diminished surgery-induced immune alterations, especially diminished elevation of IL-1. Reduced elevation of postoperative IL-1 and effective pain relief may both contribute to an attenuated illness response and a better surgery outcome.

  14. Cytokine-Mediated Regulation of Human Lymphocyte Development and Function: Insights from Primary Immunodeficiencies.

    PubMed

    Tangye, Stuart G; Pelham, Simon J; Deenick, Elissa K; Ma, Cindy S

    2017-09-15

    Cytokine-mediated intracellular signaling pathways are fundamental for the development, activation, and differentiation of lymphocytes. These distinct processes underlie protection against infectious diseases after natural infection with pathogens or immunization, thereby providing the host with long-lived immunological memory. In contrast, aberrant cytokine signaling can also result in conditions of immune dysregulation, such as early-onset autoimmunity. Thus, balanced signals provided by distinct cytokines, and delivered to specific cell subsets, are critical for immune homeostasis. The essential roles of cytokines in human immunity have been elegantly and repeatedly revealed by the discovery of individuals with mutations in cytokine ligands, receptors, and downstream transcription factors that cause primary immunodeficiency or autoimmune conditions. In this article, we review how the discovery and characterization of such individuals has identified nonredundant, and often highly specialized, functions of specific cytokines and immune cell subsets in human lymphocyte biology, host defense against infections, and immune regulation. Copyright © 2017 by The American Association of Immunologists, Inc.

  15. The Retinoic Acid Receptor-alpha mediates human T-cell activation and Th2 cytokine and chemokine production.

    PubMed

    Dawson, Harry D; Collins, Gary; Pyle, Robert; Key, Michael; Taub, Dennis D

    2008-04-16

    We have recently demonstrated that all-trans-retinoic acid (ATRA) and 9-cis-retinoic acid (9-cis RA) promote IL-4, IL-5 and IL-13 synthesis, while decreasing IFN-gamma and TNF-alpha expression by activated human T cells and reduces the synthesis of IL-12p70 from accessory cells. Here, we have demonstrated that the observed effects using ATRA and 9-cis RA are shared with the clinically useful RAR ligand, 13-cis retinoic acid (13-cis RA), and the retinoic acid receptor-alpha (RAR-alpha)-selective agonist, AM580 but not with the RAR-beta/gamma ligand, 4-hydroxyphenylretinamide (4-HPR). The increase in type 2 cytokine production by these retinoids correlated with the expression of the T cell activation markers, CD69 and CD38. The RAR-alpha-selective agonist, AM580 recapitulated all of the T cell activation and type 2 cytokine-inducing effects of ATRA and 9-cis-RA, while the RAR-alpha-selective antagonist, RO 41-5253, inhibited these effects. These results strongly support a role for RAR-alpha engagement in the regulation of genes and proteins involved with human T cell activation and type 2 cytokine production.

  16. The Retinoic Acid Receptor-α mediates human T-cell activation and Th2 cytokine and chemokine production

    PubMed Central

    Dawson, Harry D; Collins, Gary; Pyle, Robert; Key, Michael; Taub, Dennis D

    2008-01-01

    Background We have recently demonstrated that all-trans-retinoic acid (ATRA) and 9-cis-retinoic acid (9-cis RA) promote IL-4, IL-5 and IL-13 synthesis, while decreasing IFN-γ and TNF-α expression by activated human T cells and reduces the synthesis of IL-12p70 from accessory cells. Here, we have demonstrated that the observed effects using ATRA and 9-cis RA are shared with the clinically useful RAR ligand, 13-cis retinoic acid (13-cis RA), and the retinoic acid receptor-α (RAR-α)-selective agonist, AM580 but not with the RAR-β/γ ligand, 4-hydroxyphenylretinamide (4-HPR). Results The increase in type 2 cytokine production by these retinoids correlated with the expression of the T cell activation markers, CD69 and CD38. The RAR-α-selective agonist, AM580 recapitulated all of the T cell activation and type 2 cytokine-inducing effects of ATRA and 9-cis-RA, while the RAR-α-selective antagonist, RO 41–5253, inhibited these effects. Conclusion These results strongly support a role for RAR-α engagement in the regulation of genes and proteins involved with human T cell activation and type 2 cytokine production. PMID:18416830

  17. Fenretinide inhibited de novo ceramide synthesis and proinflammatory cytokines induced by Aggregatibacter actinomycetemcomitans

    PubMed Central

    Yu, Hong; Valerio, Michael; Bielawski, Jacek

    2013-01-01

    Ceramides play an essential role in modulating immune signaling pathways and proinflammatory cytokine production in response to infectious pathogens, stress stimuli, or chemotherapeutic drugs. In this study, we demonstrated that Aggregatibacter actinomycetemcomitans, the pathogen for aggressive periodontitis, induced de novo synthesis of ceramide in Raw 264.7 cells. In addition, we identified that fenretinide, a synthetic retinoid, suppressed the de novo synthesis of ceramide induced by A. actinomycetemcomitans. Moreover, fenretinide attenuated interleukin (IL)-1β, IL-6, and cyclooxygenase-2 mRNA expression induced by A. actinomycetemcomitans. Fenretinide also decreased IL-1β, IL-6, and prostaglandin E2 proinflammatory cytokine levels in Raw 264.7 cells induced by A. actinomycetemcomitans. However, fenretinide had no significant effects on tumor necrosis factor alpha mRNA or protein levels. Furthermore, we showed that fenretinide inhibited the janus kinase-signal transducer and activator of transcription, phosphatidylinositol 3-kinase-Akt, protein kinase C, and nuclear factor-kappaB signaling pathways, whereas fenretinide up-regulated the mitogen-activated protein kinase signaling pathways after bacterial stimulation. This study emphasizes the de novo ceramide synthesis pathway in response to bacterial stimulation and demonstrates the anti-inflammatory role of fenretinide in the bacteria-induced immune response. PMID:23139430

  18. Fenretinide inhibited de novo ceramide synthesis and proinflammatory cytokines induced by Aggregatibacter actinomycetemcomitans.

    PubMed

    Yu, Hong; Valerio, Michael; Bielawski, Jacek

    2013-01-01

    Ceramides play an essential role in modulating immune signaling pathways and proinflammatory cytokine production in response to infectious pathogens, stress stimuli, or chemotherapeutic drugs. In this study, we demonstrated that Aggregatibacter actinomycetemcomitans, the pathogen for aggressive periodontitis, induced de novo synthesis of ceramide in Raw 264.7 cells. In addition, we identified that fenretinide, a synthetic retinoid, suppressed the de novo synthesis of ceramide induced by A. actinomycetemcomitans. Moreover, fenretinide attenuated interleukin (IL)-1β, IL-6, and cyclooxygenase-2 mRNA expression induced by A. actinomycetemcomitans. Fenretinide also decreased IL-1β, IL-6, and prostaglandin E2 proinflammatory cytokine levels in Raw 264.7 cells induced by A. actinomycetemcomitans. However, fenretinide had no significant effects on tumor necrosis factor alpha mRNA or protein levels. Furthermore, we showed that fenretinide inhibited the janus kinase-signal transducer and activator of transcription, phosphatidylinositol 3-kinase-Akt, protein kinase C, and nuclear factor-kappaB signaling pathways, whereas fenretinide up-regulated the mitogen-activated protein kinase signaling pathways after bacterial stimulation. This study emphasizes the de novo ceramide synthesis pathway in response to bacterial stimulation and demonstrates the anti-inflammatory role of fenretinide in the bacteria-induced immune response.

  19. Cytokine release and cytotoxicity in human keratinocytes and fibroblasts induced by phenols and sodium dodecyl sulfate.

    PubMed

    Newby, C S; Barr, R M; Greaves, M W; Mallet, A I

    2000-08-01

    Phenolic compounds used in pharmaceutical and industrial products can cause irritant contact dermatitis. We studied the effects of resorcinol, phenol, 3,5-xylenol, chloroxylenol, and 4-hexyl-resorcinol on normal human epidermal keratinocytes and dermal fibroblasts for cytotoxicity and cytokine release, determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide methodology and enzyme-linked immunosorbent assay, respectively. An inverse correlation between phenol concentrations causing a 50% reduction in keratinocyte and fibroblast viability at 24 h and their octanol water-partition coefficients (i.e., hydrophobicity) was observed. 3,5-xylenol, chloroxylenol, hexyl-resorcinol, and sodium dodecyl sulfate, but not resorcinol or phenol, induced release of interleukin-1alpha from keratinocytes at cytotoxic concentrations. Variable release of tumor necrosis factor-alpha and interleukin-8 from keratinocytes occurred only at toxic threshold concentrations of the phenols or sodium dodecyl sulfate. Subtoxic concentrations of phenols or sodium dodecyl sulfate did not induce cytokine release from keratinocytes. Neither the phenols nor sodium dodecyl sulfate induced release of the chemokines interleukin-8, growth-related oncogene-alpha or monocyte chemotactic protein-1 from fibroblasts. Conditioned media from keratinocytes treated with cytotoxic concentrations of 3,5-xylenol, chloroxylenol, hexyl-resorcinol, or sodium dodecyl sulfate stimulated further release of the chemokines from fibroblasts above that obtained with control media. Rabbit anti-interleukin-1alpha serum inhibited keratinocyte-conditioned media induction of chemokine release. We have shown a structure-cytotoxicity relationship for a series of phenols as well as an association of interleukin-1alpha release with a cytotoxic effect. We demonstrated a cytokine cascade amplification step by the actions of stimulated keratinocyte media on cultured dermal fibroblasts, identifying interleukin-1alpha as

  20. Type 1 and type 2 cytokine dysregulation in human infectious, neoplastic, and inflammatory diseases.

    PubMed Central

    Lucey, D R; Clerici, M; Shearer, G M

    1996-01-01

    In the mid-1980s, Mosmann, Coffman, and their colleagues discovered that murine CD4+ helper T-cell clones could be distinguished by the cytokines they synthesized. The isolation of human Th1 and Th2 clones by Romagnani and coworkers in the early 1990s has led to a large number of reports on the effects of Th1 and Th2 on the human immune system. More recently, cells other than CD4+ T cells, including CD8+ T cells, monocytes, NK cells, B cells, eosinophils, mast cells, basophils, and other cells, have been shown to be capable of producing "Th1" and "Th2" cytokines. In this review, we examine the literature on human diseases, using the nomenclature of type 1 (Th1-like) and type 2 (Th2-like) cytokines, which includes all cell types producing these cytokines rather than only CD4+ T cells. Type 1 cytokines include interleukin-2 (IL-2), gamma interferon, IL-12 and tumor necrosis factor beta, while type 2 cytokines include IL-4, IL-5, IL-6, IL-10, and IL-13. In general, type 1 cytokines favor the development of a strong cellular immune response whereas type 2 cytokines favor a strong humoral immune response. Some of these type 1 and type 2 cytokines are cross-regulatory. For example, gamma interferon and IL-12 decrease the levels of type 2 cytokines whereas IL-4 and IL-10 decrease the levels of type 1 cytokines. We use this cytokine perspective to examine human diseases including infections due to viruses, bacteria, parasites, and fungi, as well as selected neoplastic, atopic, rheumatologic, autoimmune, and idiopathic-inflammatory conditions. Clinically, type 1 cytokine-predominant responses should be suspected in any delayed-type hypersensitivity-like granulomatous reactions and in infections with intracellular pathogens, whereas conditions involving hypergammaglobulinemia, increased immunoglobulin E levels, and/or eosinophilia are suggestive of type 2 cytokine-predominant conditions. If this immunologic concept is relevant to human diseases, the potential exists for

  1. Comparative sequence analysis of cytokine genes from human and nonhuman primates

    SciTech Connect

    Villinger, F.; Brar, S.S.; Mayne, A.

    1995-10-15

    Two major issues severely limit the studies of human recombinant cytokines/growth factors in nonhuman primates. First, assays and reagents specific for the detection and quantitation of human cytokines do not all function when utilized to detect/quantitate the nonhuman primate cytokines. Second, although most of the human cytokines appear to induce similar, if not identical, biologic function when used with cells from nonhuman primates in vitro or in vivo, they invariably induce Ab responses in vivo, precluding their repeated and/or continued use in vivo. Our laboratory has thus initiated studies to clone, sequence, and prepare recombinant cytokines from nonhuman primates and to define assays and reagents for their detection and quantitation at the nucleic acid and protein level. The data that were derived from such studies show that the nonhuman primate cytokines IL-1{alpha}, IL-1{beta}, IL-2, IL-4, IL-5, IL-6, IL-8, IL-10, IL-12{alpha}, IL-12{beta}, IL-15, IFN-{alpha}, IFN-{gamma}, and TNF-{alpha} share 93 to 99% homology at the nucleic acid and protein level with the human equivalents. The most prominent differences between human and nonhuman primate cytokine sequences were noted for IL-1{alpha}/{beta}, IL-2, IL-8, IFN-{alpha}, IFN-{gamma}, and IL-12{beta}. The aligned sequences of cytokines for human and several nonhuman primate species are provided herein, and a phylogenetic analysis of the published sequences of select cytokines from other species, along with those of the nonhuman primates, are described. In addition, comparative analysis of the relative bioactivity of our immunoaffinity-purified recombinant rhesus macaque IL-4, IL-15, and IFN-{gamma} with commercially available human recombinant cytokines is described herein. 40 refs., 5 figs., 2 tabs.

  2. Cytokines directly induce degranulation and superoxide production from human eosinophils.

    PubMed

    Horie, S; Gleich, G J; Kita, H

    1996-08-01

    Cytokines are implicated in allergic diseases and can modulate effector functions of eosinophils stimulated by another agonist. However, little is known about the capacity of cytokines to directly trigger eosinophil degranulation. We attempted to determine whether cytokines can directly induce degranulation and superoxide production from eosinophils. Eosinophils from normal donors were incubated with various cytokines in albumin-coated tissue culture plates for 4 hours. To quantitate degranulation, the amounts of eosinophil-derived neurotoxin in supernatants were measured by radioimmunoassay. In addition, superoxide production was measured by superoxide dismutase-inhibitable reduction of cytochrome c. IL-5, IL-3, granulocyte-macrophage colony-stimulating factor, tumor necrosis factor- alpha, and RANTES all induced eosinophil degranulation. Granulocyte-macrophage colony-stimulating factor was the most potent and induced eosinophil-derived neurotoxin release comparable to that induced by secretory IgA beads, one of the most potent secretagogues for eosinophils. In addition, IL-5 and tumor necrosis factor- alpha were synergistic in their induction of eosinophil degranulation. In contrast, IL-1, IL-8, interferon- gamma, and macrophage inflammatory protein-1 alpha did not induce degranulation. Finally, IL-5, IL-3, granulocyte-macrophage colony-stimulating factor, and tumor necrosis factor- alpha, but not RANTES, also induced superoxide production from eosinophils. Certain cytokines directly induce eosinophil degranulation and superoxide production in vitro. Therefore these cytokines may be important in the release of toxic granule proteins from eosinophils in allergic diseases.

  3. St. John's wort extract and hyperforin protect rat and human pancreatic islets against cytokine toxicity.

    PubMed

    Novelli, Michela; Beffy, Pascale; Menegazzi, Marta; De Tata, Vincenzo; Martino, Luisa; Sgarbossa, Anna; Porozov, Svetlana; Pippa, Anna; Masini, Matilde; Marchetti, Piero; Masiello, Pellegrino

    2014-02-01

    The extract of Hypericum perforatum (St. John's wort, SJW) and its component hyperforin (HPF) were previously shown to inhibit cytokine-induced activation of signal transducer and activator of transcription-1 and nuclear factor κB and prevent apoptosis in a cultured β-cell line. Objective of this study was to assess the protection exerted by SJW and HPF on isolated rat and human islets exposed to cytokines in vitro. Functional, ultrastructural, biomolecular and cell death evaluation studies were performed. In both rat and human islets, SJW and HPF counteracted cytokine-induced functional impairment and down-regulated mRNA expression of pro-inflammatory target genes, such as iNOS, CXCL9, CXCL10, COX2. Cytokine-induced NO production from cultured islets, evaluated by nitrites measurement in the medium, was significantly reduced in the presence of the vegetal compounds. Noteworthy, the increase in apoptosis and necrosis following 48-h exposure to cytokines was fully prevented by SJW and partially by HPF. Ultrastructural morphometric analysis in human islets exposed to cytokines for 20 h showed that SJW or HPF avoided early β-cell damage (e.g., mitochondrial alterations and loss of insulin granules). In conclusion, SJW compounds protect rat and human islets against cytokine effects by counteracting key mechanisms of cytokine-mediated β-cell injury and represent promising pharmacological tools for prevention or limitation of β-cell dysfunction and loss in type 1 diabetes.

  4. Relation of ex vivo stimulated blood cytokine synthesis to post-traumatic sepsis.

    PubMed

    Flach, R; Majetschak, M; Heukamp, T; Jennissen, V; Flohé, S; Börgermann, J; Obertacke, U; Schade, F U

    1999-02-01

    The cytokine production in endotoxin stimulated blood of patients immediately after polytrauma with high risk for developing sepsis or multi organ failure was analysed. Forty patients sustaining traumatic injury with >/=317 pts according to the Injury Severity Score (ISS), 10 of whom developed severe sepsis (ACCP/SCCM conference 1992) were included in the study. Levels of interleukin 8 (IL-8), IL-6 and tumour necrosis factor (TNF) were measured by ELISA in endotoxin-stimulated whole blood and IL-10 and IL-6 in serum. The allotype for the bi-allelic Nco I restriction length polymorphism in the TNF locus was determined for each patient.Two to four hours after polytrauma endotoxin-stimulated synthesis of TNF and IL-6 was found to be reduced in whole blood from patients compared to healthy donors, whereas no such differences were found for IL-8 synthesis. At this time, however, the patients who developed sepsis at a later stage (day 4-6) showed significantly (P<0.05) enhanced IL-8 synthesis in endotoxin stimulated whole blood in comparison to healthy donors. The IL-6 and TNF production of their blood was significantly enhanced compared to patients with uncomplicated recovery. Ninety per cent of the patients developing sepsis were of the TNFB2/TNFB2 allotype, whereas this was the case for only 30% of the non-septic group. Assessment of endotoxin-stimulated cytokine synthesis may provide a prognostic indicator for patients at high risk for developing a sepsis syndrome. Copyright 1999 Academic Press.

  5. ROCKing cytokine secretion balance in human T cells.

    PubMed

    Zanin-Zhorov, Alexandra; Waksal, Samuel D

    2015-04-01

    Balanced regulation of cytokine secretion in T cells is critical for maintenance of immune homeostasis and prevention of autoimmunity. The Rho-associated kinase (ROCK) 2 signaling pathway was previously shown to be involved in controlling of cellular movement and shape. However, recent work from our group and others has demonstrated a new and important role of ROCK2 in regulating cytokine secretion in T cells. We found that ROCK2 promotes pro-inflammatory cytokines such as IL-17 and IL-21, whereas IL-2 and IL-10 secretion are negatively regulated by ROCK2 under Th17-skewing activation. Also, in disease, but not in steady state conditions, ROCK2 contributes to regulation of IFN-γ secretion in T cells from rheumatoid arthritis patients. Thus, ROCK2 signaling is a key pathway in modulation of T-cell mediated immune responses underscoring the therapeutic potential of targeted inhibition of ROCK2 in autoimmunity.

  6. T helper 2 cytokines differently regulate monocyte chemoattractant protein-1 production by human peripheral blood monocytes and alveolar macrophages.

    PubMed

    Yano, S; Yanagawa, H; Nishioka, Y; Mukaida, N; Matsushima, K; Sone, S

    1996-09-15

    Th2 cytokines, such as IL-4, IL-10, and IL-13, suppress proinflammatory cytokine production by monocytes/macrophages. Since monocyte chemoattractant protein-1 (MCP-1) is presumed to play an important role in monocyte recruitment and activation during inflammatory and immune responses, we examined here the effects of these Th2 cytokines on MCP-1 production by human blood monocytes and alveolar macrophages. Unstimulated, highly purified blood monocytes did not produce MCP-1 spontaneously, while LPS treatment induced the production of MCP-1 and its mRNA expression. All Th2 cytokines tested suppressed LPS-induced MCP-1 production and its mRNA expression, although the suppressive effect of IL-13 was weaker than that of IL-4 or IL-10. In contrast, IL-10, but neither IL-4 nor IL-13, induced unstimulated peripheral blood monocytes to produce biologically active MCP-1 protein within 4 h, reaching a maximal level at 12 h. IL-10-induced MCP-1 production was reduced by pretreatment of IL-10 with anti-IL-10 Ab, negating the involvement of contaminated endotoxin. Moreover, IL-10 induced MCP-1 mRNA expression in unstimulated monocytes, independent of de novo protein synthesis. Furthermore, human alveolar macrophages produced MCP-1 spontaneously, and the production was inhibited by IL-4 or IL-13, but was augmented by IL-10. These findings suggest that IL-10 regulates MCP-1 production by monocytes/macrophages in a different way from other Th2 cytokines, such as IL-4 and IL-13, and contributes to host defense responses.

  7. Immune-mediated side-effects of cytokines in humans.

    PubMed

    Vial, T; Descotes, J

    1995-12-20

    A large body of clinical experience on the adverse consequences of cytokine administration has accumulated since the last decade. Side-effects reported after the therapeutic use of cytokines has provided evidence that activation of the immune response may sometimes have deleterious consequences. Several effects appeared as a direct consequence of the immune activation induced by cytokines, e.g. flu-like reactions, vascular leak syndrome. Cytokine-induced exacerbation of underlying diseases or immune dysregulation were other complications of growing concern. Interferon-alpha (IFN-alpha) treatment has now been clearly linked with the exacerbation or the occurrence of several types of autoantibodies or autoimmune diseases (thyroiditis, systemic lupus erythematosus, hematologic disorders, insulin-dependent diabetes mellitus) or diseases involving altered cell-mediated immune functions (inflammatory dermatologic diseases, nephritis, pneumonitis, colitis). By contrast immunological side-effects of IFN-beta and IFN-gamma have been seldom reported. However, the extent of clinical experience with both of these cytokines is still very limited. Interleukin-2 (IL-2) has also been implicated in various conditions that may involve immunopathological processes (thyroid disorders, rheumatoid arthritis, dermatological diseases, interstitial nephritis). Growth factors have been more specifically linked with the development or the exacerbation of dermatological inflammatory diseases through neutrophils, monocytes/macrophages or eosinophils activation (e.g. cutaneous vasculitis and generalized cutaneous eruption, Sweet's syndrome, bullous eruption, psoriasis). Exacerbation of autoimmune thyroiditis was described with granulocyte-macrophage colony-stimulating factor (GM-CSF) only. The immunogenicity of cytokines is also of great relevance and the occurrence of antibodies binding IFN-alpha and IFN-beta, IL2 and GM-CSF have been reported. While the clinical significance of non

  8. Cytokine modulation of human blood viscosity from vivax malaria patients.

    PubMed

    Scherer, Edson Fredulin; Cantarini, Déborah Giovanna; Siqueira, Renan; Ribeiro, Elton Brito; Braga, Érika Martins; Honório-França, Adenilda Cristina; França, Eduardo Luzía

    2016-06-01

    Malaria is a major infectious disease in several countries and is caused by protozoa of the genus Plasmodium. In vivax malaria patients, inflammatory processes occur, as well as changes in cytokines and blood flow. The present study analyzed the cytokine modulation of blood viscosity from patients infected with Plasmodium vivax (P. vivax). Blood samples were collected from 42 non-infected individuals (control group) and 37 individuals infected with P. vivax. The IL-2, IL-4, IL-6, IL-10, TNFα, TGF-β and IL-17 cytokine concentrations in the serum were assessed, and the blood rheological properties were determined. The analysis of blood viscosity for shear rates revealed that the blood viscosity of the infected patients was significantly greater than that of the non-infected individuals. The viscosity of the blood was greater in the infected individuals than in the non-infected subjects. The serum from individuals with P. vivax infections exhibited higher IFN-γ and IL-17 concentrations and lower TGF-β levels. Incubation of the blood from infected individuals with IL-17 or IL-17 associated with IFN-γ reduced the viscosity to rates equivalent to the blood from non-infected individuals. Independently of cytokine modulation, no correlation was found between the parasitemia and blood viscosity of the infected patients. These data suggest that the alterations of blood viscosity are relevant as an auxiliary tool for the clinical diagnosis of disease. In malaria, erythrocytes are more sensitive to osmotic shock, and the reduction of viscosity by IL-17 may be related to a possible immunomodulator agent during infection.

  9. 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.

  10. Serum cytokine profiles of children with human enterovirus 71-associated hand, foot, and mouth disease.

    PubMed

    Han, Jun; Wang, Ying; Gan, Xing; Song, Juan; Sun, Peng; Dong, Xiao-Ping

    2014-08-01

    Cytokine profiles may impact the pathogenicity and severity of hand, foot, and mouth disease caused by human enterovirus (HEV) 71. In 91 severe or mild HEV 71-associated hand, foot, and mouth disease children, serum was collected between days 2 and 10 or day >10. Serum cytokines including Type 1 T helper (Th1) cytokines: interleukin (IL)-2, interferon-gamma (IFN-γ), IL-12, and IL-18, Type 1 T helper (Th2) cytokines: IL-4, IL-10, IL-13, proinflammatory cytokines: IL-1α, IL-1β, IL-6, IL-8, IL-17, and tumor necrosis factor alpha (TNF-α), were assessed during the early stage and recovery. In the patients with mild illness, the peaks of IL-8 and IL-10 were observed on day 6 and that of IL-18 was on day 4. In the patients with severe illness, all cytokines spiked on day 3 and peaked on day 11. All cytokines except IL-6, IL-8, IL-18, and TNF-α were significantly correlated with immunoglobulin M levels by the end of the disease course. Cytokine profile variations between the patients with mild and severe illness may indicate prognosis and strain virulence, useful in clinical treatment of patients.

  11. Specific activation of human interleukin-5 depends on de novo synthesis of an AP-1 complex.

    PubMed

    Schwenger, Gretchen T F; Kok, Chee Choy; Arthaningtyas, Estri; Thomas, Marc A; Sanderson, Colin J; Mordvinov, Viatcheslav A

    2002-12-06

    It is clear from the biology of eosinophilia that a specific regulatory mechanism must exist. Because interleukin-5 (IL5) is the key regulatory cytokine, it follows that a gene-specific control of IL5 expression must exist that differs even from closely related cytokines such as IL4. Two features of IL5 induction make it unique compared with other cytokines; first, induction by cyclic adenosine monophosphate (cAMP), which inhibits other T-cell-derived cytokines, and second, sensitivity to protein synthesis inhibitors, which have no effect on other cytokines. This study has utilized the activation of different transcription factors by different stimuli in a human T-cell line to study the role of conserved lymphokine element 0 (CLE0) in the specific induction of IL5. In unstimulated cells the ubiquitous Oct-1 binds to CLE0. Stimulation induces de novo synthesis of the AP-1 members JunD and Fra-2, which bind to CLE0. The amount of IL5 produced correlates with the production of the AP-1 complex, suggesting a key role in IL5 expression. The formation of the AP-1 complex is essential, but the rate-limiting step is the synthesis of AP-1, especially Fra-2. This provides an explanation for the sensitivity of IL5 to protein synthesis inhibitors and a mechanism for the specific induction of IL5 compared with other cytokines.

  12. A whole blood in vitro cytokine release assay with aqueous monoclonal antibody presentation for the prediction of therapeutic protein induced cytokine release syndrome in humans.

    PubMed

    Wolf, Babette; Morgan, Hannah; Krieg, Jennifer; Gani, Zaahira; Milicov, Adriana; Warncke, Max; Brennan, Frank; Jones, Stewart; Sims, Jennifer; Kiessling, Andrea

    2012-12-01

    The administration of several monoclonal antibodies (mAbs) to humans has been associated with acute adverse events characterized by clinically significant release of cytokines in the blood. The limited predictive value of toxicology species in this field has triggered intensive research to establish human in vitro assays using peripheral blood mononuclear cells or blood to predict cytokine release in humans. A thorough characterization of these assays is required to understand their predictive value for hazard identification and risk assessment in an optimal manner, and to highlight potential limitations of individual assay formats. We have characterized a whole human blood cytokine release assay with only minimal dilution by the test antibodies (95% v/v blood) in aqueous presentation format, an assay which has so far received less attention in the scientific world with respect to the evaluation of its suitability to predict cytokine release in humans. This format was compared with a human PBMC assay with immobilized mAbs presentation already well-characterized by others. Cytokine secretion into plasma or cell culture supernatants after 24h incubation with the test mAbs (anti-CD28 superagonist TGN1412-like material (TGN1412L), another anti-CD28 superagonistic mAb (ANC28.1), a T-cell depleting mAb (Orthoclone™), and a TGN1412 isotype-matched control (Tysabri™) not associated with clinically-relevant cytokine release) was detected by a multiplex assay based on electrochemiluminescent excitation. We provide proof that this whole blood assay is a suitable new method for hazard identification of safety-relevant cytokine release in the clinic based on its ability to detect the typical cytokine signatures found in humans for the tested mAbs and on a markedly lower assay background and cytokine release with the isotype-matched control mAb Tysabri™ - a clear advantage over the PBMC assay. Importantly, quantitative and qualitative differences in the relative cytokine

  13. Bacterial modulins: a novel class of virulence factors which cause host tissue pathology by inducing cytokine synthesis.

    PubMed Central

    Henderson, B; Poole, S; Wilson, M

    1996-01-01

    Cytokines are a diverse group of proteins and glycoproteins which have potent and wide-ranging effects on eukaryotic cell function and are now recognized as important mediators of tissue pathology in infectious diseases. It is increasingly recognized that for many bacterial species, cytokine induction is a major virulence mechanism. Until recent years, the only bacterial component known to stimulate cytokine synthesis was lipopolysaccharide (LPS). It is only within the past decade that it has been clearly shown that many components associated with the bacterial cell wall, including proteins, glycoproteins, lipoproteins, carbohydrates, and lipids, have the capacity to stimulate mammalian cells to produce a diverse array of cytokines. It has been established that many of these cytokine-inducing molecules act by mechanisms distinct from that of LPS, and thus their activities are not due to LPS contamination. Bacteria produce a wide range of virulence factors which cause host tissue pathology, and these diverse factors have been grouped into four families: adhesins, aggressins, impedins, and invasins. We suggest that the array of bacterial cytokine-inducing molecules represents a new class of bacterial virulence factor, and, by analogy with the known virulence families, we suggest the term "modulin" to describe these molecules, because the action of cytokines is to modulate eukaryotic cell behavior. This review summarizes our current understanding of cytokine biology in relation to tissue homeostasis and disease and concisely reviews the current literature on the cytokine-inducing molecules produced by gram-negative and gram-positive bacteria, with an emphasis on the cellular mechanisms responsible for cytokine induction. We propose that modulins, by controlling the host immune and inflammatory responses, maintain the large commensal flora that all multicellular organisms support. PMID:8801436

  14. Circadian disruption, Per3, and human cytokine secretion.

    PubMed

    Guess, Jaclyn; Burch, James B; Ogoussan, Kisito; Armstead, Cheryl A; Zhang, Hongmei; Wagner, Sara; Hebert, James R; Wood, Patricia; Youngstedt, Shawn D; Hofseth, Lorne J; Singh, Udai P; Xie, Dawen; Hrushesky, William J M

    2009-12-01

    Circadian disruption has been linked with inflammation, an established cancer risk factor. Per3 clock gene polymorphisms have also been associated with circadian disruption and with increased cancer risk. Patients completed a questionnaire and provided a blood sample prior to undergoing a colonoscopy (n = 70). Adjusted mean serum cytokine concentrations (IL-6, TNF-alpha, gamma-INF, IL-1ra, IL-1-beta, VEGF) were compared among patients with high and low scores for fatigue (Multidimensional Fatigue Inventory), depressive symptoms (Beck Depression Inventory II), or sleep disruption (Pittsburgh Sleep Quality Index), or among patients with different Per3 clock gene variants. Poor sleep was associated with elevated VEGF, and fatigue-related reduced activity was associated with elevated TNF-alpha concentrations. Participants with the 4/5 or 5/5 Per3 variable tandem repeat sequence had elevated IL-6 concentrations compared to those with the 4/4 genotype. Biological processes linking circadian disruption with cancer remain to be elucidated. Increased inflammatory cytokine secretion may play a role.

  15. Human cytokine responses induced by Gram-positive cell walls of normal intestinal microbiota

    PubMed Central

    Chen, T; Isomäki, P; Rimpiläinen, M; Toivanen, P

    1999-01-01

    The normal microbiota plays an important role in the health of the host, but little is known of how the human immune system recognizes and responds to Gram-positive indigenous bacteria. We have investigated cytokine responses of peripheral blood mononuclear cells (PBMC) to Gram-positive cell walls (CW) derived from four common intestinal indigenous bacteria, Eubacterium aerofaciens (Eu.a.), Eubacterium limosum(Eu.l.), Lactobacillus casei(L.c.), and Lactobacillus fermentum (L.f.). Our results indicate that Gram-positive CW of the normal intestinal microbiota can induce cytokine responses of the human PBMC. The profile, level and kinetics of these responses are similar to those induced by lipopolysaccharide (LPS) or CW derived from a pathogen, Streptococcus pyogenes (S.p.). Bacterial CW are capable of inducing production of a proinflammatory cytokine, tumour necrosis factor-alpha (TNF-α), and an anti-inflammatory cytokine, IL-10, but not that of IL-4 or interferon-gamma (IFN-γ). Monocytes are the main cell population in PBMC to produce TNF-α and IL-10. Induction of cytokine secretion is serum-dependent; both CD14-dependent and -independent pathways are involved. These findings suggest that the human cytokine responses induced by Gram-positive CW of the normal intestinal microbiota are similar to those induced by LPS or Gram-positive CW of the pathogens. PMID:10540188

  16. Infection and upregulation of proinflammatory cytokines in human brain vascular pericytes by human cytomegalovirus

    PubMed Central

    2012-01-01

    Background Congenital human cytomegalovirus (HCMV) infections can result in CNS abnormalities in newborn babies including vision loss, mental retardation, motor deficits, seizures, and hearing loss. Brain pericytes play an essential role in the development and function of the blood–brain barrier yet their unique role in HCMV dissemination and neuropathlogy has not been reported. Methods Primary human brain vascular pericytes were exposed to a primary clinical isolate of HCMV designated ‘SBCMV’. Infectivity was analyzed by microscopy, immunofluorescence, Western blot, and qRT-PCR. Microarrays were performed to identify proinflammatory cytokines upregulated after SBCMV exposure, and the results validated by real-time quantitative polymerase chain reaction (qPCR) methodology. In situ cytokine expression of pericytes after exposure to HCMV was examined by ELISA and in vivo evidence of HCMV infection of brain pericytes was shown by dual-labeled immunohistochemistry. Results HCMV-infected human brain vascular pericytes as evidenced by several markers. Using a clinical isolate of HCMV (SBCMV), microscopy of infected pericytes showed virion production and typical cytomegalic cytopathology. This finding was confirmed by the expression of major immediate early and late virion proteins and by the presence of HCMV mRNA. Brain pericytes were fully permissive for CMV lytic replication after 72 to 96 hours in culture compared to human astrocytes or human brain microvascular endothelial cells (BMVEC). However, temporal transcriptional expression of pp65 virion protein after SBCMV infection was lower than that seen with the HCMV Towne laboratory strain. Using RT-PCR and dual-labeled immunofluorescence, proinflammatory cytokines CXCL8/IL-8, CXCL11/ITAC, and CCL5/Rantes were upregulated in SBCMV-infected cells, as were tumor necrosis factor-alpha (TNF-alpha), interleukin-1 beta (IL-1beta), and interleukin-6 (IL-6). Pericytes exposed to SBCMV elicited higher levels of IL-6

  17. Faithful expression of the human 5q31 cytokine cluster intransgenic mice

    SciTech Connect

    Lacy, Dee A.; Wang, Zhi-En; Symula, Derek J.; McArthur, CliffordJ.; Rubin, Edward M.; Frazer, Kelly A.; Locksley, Richard M.

    1999-12-03

    ILs 4,5, and 13, cardinal cytokines produced by Th2 cells,are coordinately expressed and clustered in the 150-kb syntenic regions on mouse chromosome 11 and human chromosome 5q31. We analyzed two sets of human yeast artificial chromosome transgenic mice that contained the5931cytokines to assess whether conserved sequences required for their coordinate and cell-specific regulation are contained within the cytokine cluster itself. Human Il-4, IL-13, and Il-5 were expressed under Th2, but not Th1, conditions in vitro. Each of these cytokines was produced during infection with Nippostrongylus brasiliensis, a Th2 inducing stimulus, and human Il-4 was generated after activation of NK T cells in vivo.Consistently fewer cells produced the endogenous mouse cytokines in transgenic than in control mice, suggesting competition for stable expression between the mouse and human genes. These data imply the existence of both conserved trans-activating factors and cis-regulatory elements that underlie the coordinate expression and lineage specificity of the type 2 ctyokine genes in lymphocytes.

  18. Modulation of cytokines and chemokines expression by NAC in cadmium chloride treated human lung cells.

    PubMed

    Odewumi, Caroline O; Latinwo, Lekan M; Ruden, Michael L; Badisa, Veera L D; Fils-Aime, Sheila; Badisa, Ramesh B

    2016-11-01

    Cadmium (Cd), is one of the most hazardous metals found in the environment. Cd exposure through inhalation has been linked to various diseases in lungs. It was shown that Cd induces proinflammatory cytokines through oxidative stress mechanism. In this report, we studied the immunomodulatory effect of a well known antioxidant, N-acetylcysteine (NAC) on cadmium chloride (CdCl2 ) treated human lung A549 cells through human cytokine array 6. The lung cells were treated with 0 or 75 µM CdCl2 alone, 2.5 mM NAC alone, or co-treated with 2.5 mM NAC and 75 µM CdCl2 for 24 h. The viability of cells was measured by crystal violet dye. The array results were validated by human IL-1alpha enzyme- linked immunosorbent assay (ELISA) kit. The viability of the 75 µM CdCl2 alone treated cells was decreased to 44.5%, while the viability of the co-treated cells with 2.5 mM NAC was increased to 84.1% in comparison with untreated cells. In the cell lysate of CdCl2 alone treated cells, 19 and 8 cytokines were up and down-regulated, while in the medium 15 and 3 cytokines were up and downregulated in comparison with the untreated cells. In the co-treated cells, all these cytokines expression was modulated by the NAC treatment. The IL-1α ELISA result showed the same pattern of cytokine expression as the cytokine array. This study clearly showed the modulatory effect of NAC on cytokines and chemokines expression in CdCl2- treated cells and suggests the use of NAC as protective agent against cadmium toxicity. © 2015 Wiley Periodicals, Inc. Environ Toxicol 31: 1612-1619, 2016.

  19. Polyfunctional responses by human T cells result from sequential release of cytokines

    PubMed Central

    Han, Qing; Bagheri, Neda; Bradshaw, Elizabeth M.; Hafler, David A.; Lauffenburger, Douglas A.; Love, J. Christopher

    2012-01-01

    The release of cytokines by T cells defines a significant part of their functional activity in vivo, and their ability to produce multiple cytokines has been associated with beneficial immune responses. To date, time-integrated end-point measurements have obscured whether these polyfunctional states arise from the simultaneous or successive release of cytokines. Here, we used serial, time-dependent, single-cell analysis of primary human T cells to resolve the temporal dynamics of cytokine secretion from individual cells after activation ex vivo. We show that multifunctional, Th1-skewed cytokine responses (IFN-γ, IL-2, TNFα) are initiated asynchronously, but the ensuing dynamic trajectories of these responses evolve programmatically in a sequential manner. That is, cells predominantly release one of these cytokines at a time rather than maintain active secretion of multiple cytokines simultaneously. Furthermore, these dynamic trajectories are strongly associated with the various states of cell differentiation suggesting that transient programmatic activities of many individual T cells contribute to sustained, population-level responses. The trajectories of responses by single cells may also provide unique, time-dependent signatures for immune monitoring that are less compromised by the timing and duration of integrated measures. PMID:22160692

  20. Body mass index is not associated with cytokine induction during experimental human endotoxemia.

    PubMed

    van Eijk, Lucas T; van der Pluijm, Rob W; Ramakers, Bart Pc; Dorresteijn, Mirrin J; van der Hoeven, Johannes G; Kox, Matthijs; Pickkers, Peter

    2014-01-01

    A higher body mass index (BMI) appears to be associated with lower mortality in critically ill patients, possibly explained by an altered innate immune response. However, the precise relationship between BMI and the innate immune response in humans in vivo is unknown. We investigated the relationship between BMI and the systemic cytokine response during experimental human endotoxemia. Endotoxemia was induced in 112 healthy male volunteers by intravenous administration of 2 ng/kg Escherichia coli endotoxin. Plasma concentrations of TNF-α, IL-6, IL-10 and IL-1RA were serially determined. The relationship between BMI and the cytokine response, as well as body temperature, was investigated. The BMIs of the participants ranged from 18.3 to 33.6 kg/m(2), (median: 22.7 kg/m(2)). All participants showed a marked increase in plasma cytokine levels [median (interquartile range)] peak levels: TNF-α 509 (353-673) pg/ml; IL-6 757 (522-1098) pg/ml; IL-10 271 (159-401) pg/ml; IL-1RA 4882 (3927-6025) pg/ml; and an increase in body temperature [1.8(1.4-2.2)] during endotoxemia. No significant correlations were found between BMI and levels of any of the cytokines or body temperature. No relationship between BMI and the cytokine response was found in healthy volunteers subjected to experimental endotoxemia. These data question the relationship between BMI and cytokine responses in critical illness.

  1. Pro- and anti-inflammatory cytokines in human immunodeficiency virus infection and acquired immunodeficiency syndrome.

    PubMed

    Breen, Elizabeth Crabb

    2002-09-01

    In persons with human immunodeficiency virus (HIV) infection and/or acquired immunodeficiency syndrome (AIDS), the immune system becomes dysfunctional in many ways. There is both immunodeficiency due to the loss of CD4-positive T helper cells and hyperactivity as a result of B-cell activation. Likewise, both decreases and increases are seen in the production and/or activity of cytokines. Cytokine changes in HIV infection have been assessed by a variety of techniques, ranging from determination of cytokine gene expression at the mRNA level to secretion of cytokine proteins in vivo and in vitro. Changes in cytokine levels in HIV-infected persons can affect the function of the immune system, and have the potential to directly impact the course of HIV disease by enhancing or suppressing HIV replication. In particular, the balance between the pro-inflammatory cytokines interleukin (IL)-1, IL-6, and tumor necrosis factor-alpha, which up-regulate HIV expression, and IL-10, which can act both as an anti-inflammatory cytokine and a B-cell stimulatory factor, may play an important role in the progression to AIDS. In light of its ability to suppress the production of pro-inflammatory cytokines and, under some conditions, suppress HIV replication, increased IL-10 may be viewed as beneficial in slowing HIV disease progression. However, an association between increased IL-10 and the development of AIDS-associated B-cell lymphoma highlights the bifunctional nature of IL-10 as both an anti-inflammatory and B-cell-stimulatory cytokine that could have beneficial and detrimental effects on the course of HIV infection and AIDS.

  2. Cytokines and macrophage function in humans - role of stress

    NASA Technical Reports Server (NTRS)

    Sonnenfeld, Gerald (Principal Investigator)

    1996-01-01

    We have begun this study to commence the determination of the role of mild chronic stress in the effects of space flight on macrophage/monocyte function, a component of the immune response. Medical students undergoing regular periods of stress and relaxation have been shown to be an excellent model for determining the effects of stress on immune responses. We have begun using this model using the macrophage/monocyte as model leukocyte. The monocyte/macrophage plays a central role in immunoregulation. The studies to be included in this three year project are the effects of stress on: (1) interactions of monocytes with microbes, (2) monocyte production of cytokines, (3) monocyte phagocytosis and activity, and (4) monocyte expression of cell surface antigens important in immune responses. Stress hormone levels will also be carried out to determine if there is a correlation between stress effects on immune responses and hormonal levels. Psychological testing to insure subjects are actually stressed or relaxed at the time of testing will also be carried out. The results obtained from the proposed studies should be comparable with space flight studies with whole animals and isolated cell cultures. When complete this study should allow the commencement of the establishment of the role of stress as one compartment of the induction of immune alterations by space flight.

  3. The effect of garlic powder on human urinary cytokine excretion.

    PubMed

    Alma, Ergun; Eken, Alper; Ercil, Hakan; Yelsel, Kazim; Daglioglu, Nebile

    2014-03-04

    To evaluate the effects of orally administered dehydrated garlic powder on cytokine excretion in the urinary tract. A total of 60 healthy volunteers, randomized into 3 groups, were given a single oral dose of 1 g or 3 g of dehydrated garlic powder or placebo. Urine samples were obtained 6.0 and 24.0 h after garlic intake and assayed for interleukin-8 (IL-8), interleukin- 12 (IL-12), tumor necrosis factor-alpha (TNF-α), diallyl disulfide (DADS) and diallyl sulfide (DAS). Significant increases in IL-12 levels over baseline were noted in urine samples obtained after oral intake of 1 g and 3 g of garlic powder (P < .001). In the 1 g and 3 g garlic powder treatment groups, time-dependent variations in IL-12 levels over the study period were significantly different from the placebo group (P < .001). In both garlic treatment groups, urinary levels of IL-8 and TNF-α were not significantly different from baseline and placebo levels (P > .017). DADS and DAS were not detected in the urine samples at any time after garlic powder intake. Oral intake of doses of garlic traditionally used for daily supplementation increases urinary levels of IL-12, which is a potent stimulator of T helper cell 1 (Th-1) immune responses. This observation encourages further studies investigating the immunostimulatory role of garlic in the urinary tract.

  4. Cytokine synthesis in occupational allergy to caddisflies in hydroelectric plant workers.

    PubMed

    Warrington, R J; Whitman, C; McPhillips Warrington, S

    2003-10-01

    Workers in hydroelectric plants appear to be readily sensitized to caddisfly allergens. This sensitization probably occurs de novo from occupational exposure. In some workers, sensitization occurs on a non-atopic background. Cytokine synthesis of IFN-gamma, IL-5 and IL-13 in atopic and non-atopic caddisfly-allergic workers was examined to determine if responses were similar or different. Peripheral blood mononuclear cells were isolated from atopic caddisfly-allergic workers, non-atopic caddisfly-allergic workers and non-atopic caddisfly-exposed but non-allergic workers. Stimulation with caddisfly antigens was carried out and synthesis of IFN-gamma, IL-5 and IL-13 was determined by sandwich ELISA. Both caddisfly-allergic and non-allergic subjects responded to stimulation with caddisfly extract. The response in non-atopic caddisfly-non-allergic subjects was TH1 predominant, while that in atopic caddisfly-allergic subjects was TH2 predominant. The response in non-atopic caddisfly-allergic subjects was between that of the atopic caddisfly-allergic workers and the non-atopic caddisfly-non-allergic workers and the trend was to a TH2 response. Work-related symptoms were similarly intermediate between the atopic caddisfly-allergic and non-atopic caddisfly-non-allergic group. Differences were significant for IFN-gamma/IL-5 ratios but not IFN-gamma/IL-13 ratios for atopic and non-atopic caddisfly-allergic individuals, compared to non-atopic caddisfly-non-allergic workers. However, a linear relationship existed between IFN-gamma synthesis and IL-5 and IL-13 synthesis in non-atopic caddisfly-allergic workers but not in atopic caddisfly-allergic subjects. Caddisfly allergy in hydroelectric workers may be a useful model for the development of allergy to a previously unencountered allergen, and points to some interesting differences between atopic and non-atopic subjects who become sensitized to environmental allergens. Copyright 2003 S. Karger AG, Basel

  5. Yeast Modulation of Human Dendritic Cell Cytokine Secretion: An In Vitro Study

    PubMed Central

    Smith, Ida M.; Christensen, Jeffrey E.; Arneborg, Nils; Jespersen, Lene

    2014-01-01

    Probiotics are live microorganisms which when administered in adequate amounts confer a health benefit on the host. The concept of individual microorganisms influencing the makeup of T cell subsets via interactions with intestinal dendritic cells (DCs) appears to constitute the foundation for immunoregulatory effects of probiotics, and several studies have reported probiotic strains resulting in reduction of intestinal inflammation through modulation of DC function. Consequent to a focus on Saccharomyces boulardii as the fundamental probiotic yeast, very little is known about hundreds of non-Saccharomyces yeasts in terms of their interaction with the human gastrointestinal immune system. The aim of the present study was to evaluate 170 yeast strains representing 75 diverse species for modulation of inflammatory cytokine secretion by human DCs in vitro, as compared to cytokine responses induced by a S. boulardii reference strain with probiotic properties documented in clinical trials. Furthermore, we investigated whether cytokine inducing interactions between yeasts and human DCs are dependent upon yeast viability or rather a product of membrane interactions regardless of yeast metabolic function. We demonstrate high diversity in yeast induced cytokine profiles and employ multivariate data analysis to reveal distinct clustering of yeasts inducing similar cytokine profiles in DCs, highlighting clear species distinction within specific yeast genera. The observed differences in induced DC cytokine profiles add to the currently very limited knowledge of the cross-talk between yeasts and human immune cells and provide a foundation for selecting yeast strains for further characterization and development toward potentially novel yeast probiotics. Additionally, we present data to support a hypothesis that the interaction between yeasts and human DCs does not solely depend on yeast viability, a concept which may suggest a need for further classifications beyond the current

  6. Yeast modulation of human dendritic cell cytokine secretion: an in vitro study.

    PubMed

    Smith, Ida M; Christensen, Jeffrey E; Arneborg, Nils; Jespersen, Lene

    2014-01-01

    Probiotics are live microorganisms which when administered in adequate amounts confer a health benefit on the host. The concept of individual microorganisms influencing the makeup of T cell subsets via interactions with intestinal dendritic cells (DCs) appears to constitute the foundation for immunoregulatory effects of probiotics, and several studies have reported probiotic strains resulting in reduction of intestinal inflammation through modulation of DC function. Consequent to a focus on Saccharomyces boulardii as the fundamental probiotic yeast, very little is known about hundreds of non-Saccharomyces yeasts in terms of their interaction with the human gastrointestinal immune system. The aim of the present study was to evaluate 170 yeast strains representing 75 diverse species for modulation of inflammatory cytokine secretion by human DCs in vitro, as compared to cytokine responses induced by a S. boulardii reference strain with probiotic properties documented in clinical trials. Furthermore, we investigated whether cytokine inducing interactions between yeasts and human DCs are dependent upon yeast viability or rather a product of membrane interactions regardless of yeast metabolic function. We demonstrate high diversity in yeast induced cytokine profiles and employ multivariate data analysis to reveal distinct clustering of yeasts inducing similar cytokine profiles in DCs, highlighting clear species distinction within specific yeast genera. The observed differences in induced DC cytokine profiles add to the currently very limited knowledge of the cross-talk between yeasts and human immune cells and provide a foundation for selecting yeast strains for further characterization and development toward potentially novel yeast probiotics. Additionally, we present data to support a hypothesis that the interaction between yeasts and human DCs does not solely depend on yeast viability, a concept which may suggest a need for further classifications beyond the current

  7. Enhanced and efficient detection of virus-driven cytokine expression by human NK and T cells.

    PubMed

    Pattacini, Laura; Murnane, Pamela M; Fluharty, Tayler R; Katabira, Elly; De Rosa, Stephen C; Baeten, Jared M; Lund, Jennifer M

    2014-04-01

    Cutting edge immune monitoring techniques increasingly measure multiple functional outputs for various cell types, such as intracellular cytokine staining (ICS) assays that measure cytokines expressed by T cells. To date, however, there is no precise method to measure virus-specific cytokine production by both T cells as well as NK cells in the same well, which is important to a greater extent given recent identification of NK cells expressing a memory phenotype. This study describes an adaptable and efficient ICS assay platform that can be used to detect antigen-driven cytokine production by human T cells and NK cells, termed "viral ICS". Importantly, this assay uses limited amount of cryopreserved PBMCs along with autologous heat-inactivated serum, thereby allowing for this assay to be performed when sample is scarce as well as geographically distant from the laboratory. Compared to a standard ICS assay that detects antigen-specific T cell cytokine expression alone, the viral ICS assay is comparable in terms of both HIV-specific CD4 and CD8T cell cytokine response rates and magnitude of response, with the added advantage of ability to detect virus-specific NK cell responses.

  8. Cocoa procyanidins and human cytokine transcription and secretion.

    PubMed

    Mao, T; Van De Water, J; Keen, C L; Schmitz, H H; Gershwin, M E

    2000-08-01

    We examined whether cocoa, in its isolated procyanidin fractions (monomer through decamer), would modulate cytokine production at the levels of transcription and protein secretion in both resting and phytohemagglutinin (PHA)-stimulated peripheral blood mononuclear cells (PBMC). In resting cells, interleukin (IL)-1beta and IL-4 gene expression from cocoa-treated cells varied markedly among the subjects tested. However, at the protein level, the larger fractions (pentamer through decamer) stimulated a dramatic increase in IL-1beta concentration (up to ninefold) with increasing degree of polymerization. Similarly, these larger fractions augmented IL-4 concentration by as much as 2 pg/ml, whereas the control displayed levels nearly undetectable. In the presence of PHA, gene expression also seemed to be most affected by the larger procyanidin fractions. The pentameric through decameric fractions increased IL-1 beta expression by 7-19% compared with PHA control, whereas the hexameric through decameric fractions significantly inhibited PHA-induced IL-4 transcription in the range of 71-86%. This observation at the transcription level for IL-1 beta was reflected at the protein level in PHA-stimulated PBMC. Significant reductions in mitogen-induced IL-4 production were also seen at the protein level with the hexamer, heptamer and octamer. Individual oligomeric cocoa fractions were unstimulatory for IL-2 in resting PBMC. However, when induced with PHA, the pentamer, hexamer and heptamer fractions caused a 61-73% inhibition in IL-2 gene expression. This study offers additional data for the consideration of the health benefits of dietary polyphenols from a wide variety of foods, including those benefits associated specifically with cocoa and chocolate consumption.

  9. 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.

  10. A review of the influence of growth factors and cytokines in in vitro human keratinocyte migration.

    PubMed

    Peplow, Philip V; Chatterjee, Marissa P

    2013-04-01

    Keratinocyte migration from the wound edge is a crucial step in the reepithelization of cutaneous wounds. Growth factors and cytokines, released from cells that invade the wound matrix, play an important role, and several in vitro assays have been performed to elucidate this. The purposes of this study were to review in vitro human studies on keratinocyte migration to identify those growth factors or cytokines that stimulate keratinocyte migration and whether these assays might serve as a screening procedure prior to testing combinations of growth factors or cytokines to promote wound closure in vivo. Research papers investigating effect of growth factors and cytokines on human keratinocyte migration in vitro were retrieved from library sources, PubMed databases, reference lists of papers, and searches of relevant journals. Fourteen different growth factors and cytokines enhanced migration in scratch wound assay and HGF together with TGF-β, and IGF-1 with EGF, were more stimulatory than either growth factor alone. HGF with TGF-β1 had a greater chemokinetic effect than either growth factor alone in transmigration assay. TGF-β1, FGF-7, FGF-2 and AGF were chemotactic to keratinocytes. EGF, TGF-α, IL-1α, IGF and MGSA enhanced cell migration on ECM proteins. Many growth factors and cytokines enhanced migration of keratinocytes in vitro, and certain combinations of growth factors were more stimulatory than either alone. These and other combinations that stimulate keratinocyte migration in vitro should be tested for effect on wound closure and repair in vivo. The scratch wound assay provides a useful, inexpensive and easy-to-perform screening method for testing individual or combinations of growth factors or cytokines, or growth factors combined with other modalities such as laser irradiation, prior to performing wound healing studies with laboratory animals. Copyright © 2013 Elsevier Ltd. All rights reserved.

  11. Identification of Lactobacillus plantarum genes modulating the cytokine response of human peripheral blood mononuclear cells

    PubMed Central

    2010-01-01

    Background Modulation of the immune system is one of the most plausible mechanisms underlying the beneficial effects of probiotic bacteria on human health. Presently, the specific probiotic cell products responsible for immunomodulation are largely unknown. In this study, the genetic and phenotypic diversity of strains of the Lactobacillus plantarum species were investigated to identify genes of L. plantarum with the potential to influence the amounts of cytokines interleukin 10 (IL-10) and IL-12 and the ratio of IL-10/IL-12 produced by peripheral blood mononuclear cells (PBMCs). Results A total of 42 Lactobacillus plantarum strains isolated from diverse environmental and human sources were evaluated for their capacity to stimulate cytokine production in PBMCs. The L. plantarum strains induced the secretion of the anti-inflammatory cytokine IL-10 over an average 14-fold range and secretion of the pro-inflammatory cytokine IL-12 over an average 16-fold range. Comparisons of the strain-specific cytokine responses of PBMCs to comparative genome hybridization profiles obtained with L. plantarum WCFS1 DNA microarrays (also termed gene-trait matching) resulted in the identification of 6 candidate genetic loci with immunomodulatory capacities. These loci included genes encoding an N-acetyl-glucosamine/galactosamine phosphotransferase system, the LamBDCA quorum sensing system, and components of the plantaricin (bacteriocin) biosynthesis and transport pathway. Deletion of these genes in L. plantarum WCFS1 resulted in growth phase-dependent changes in the PBMC IL-10 and IL-12 cytokine profiles compared with wild-type cells. Conclusions The altered PBMC cytokine profiles obtained with the L. plantarum WCFS1 mutants were in good agreement with the predictions made by gene-trait matching for the 42 L. plantarum strains. This study therefore resulted in the identification of genes present in certain strains of L. plantarum which might be responsible for the stimulation of anti

  12. Complete artificial saliva alters expression of proinflammatory cytokines in human dermal fibroblasts.

    PubMed

    Malpass, Gloria E; Arimilli, Subhashini; Prasad, Gaddamanugu L; Howlett, Allyn C

    2013-07-01

    Complete artificial saliva (CAS) is a saliva substitute often used as a vehicle for test articles, including smokeless tobacco products. In the course of a study employing normal adult human dermal fibroblasts (HDFa) as a model in vitro, we discovered that CAS as a vehicle introduced a significant change in the expression of proinflammatory cytokines. To determine the effects of CAS on gene expression, real-time quantitative reverse-transcriptase PCR gene array analysis was used. Results indicate that robust changes in the expression of the proinflammatory cytokine interleukin 8 (IL8) and the vascular cell adhesion molecule 1 (VCAM1) occur within 5h of exposure to CAS. To determine whether CAS also alters cytokine release into the culture media, cytometric bead array assays for human inflammatory cytokines were performed. Analysis shows that CAS induced the release of IL8 and IL6. This study focused on determining which components in CAS were responsible for the proinflammatory response in HDFa. The following components were investigated: α-amylase, lysozyme, acid phosphatase, and urea. Results demonstrated that enzymatically active α-amylase induced gene expression for proinflammatory cytokines IL8, IL6, tumor necrosis factor-α, and IL1α and for VCAM1. Therefore, it is important to carefully evaluate the "vehicle effects" of CAS and its components in in vitro toxicology research.

  13. The expression pattern of two novel cytokines (IL-24 and IL-29) in human fetal membranes.

    PubMed

    Nace, Judith; Fortunato, Stephen J; Maul, Holger; Menon, Ramkumar

    2010-11-01

    interleukin (IL)-24 and -29 are novel cytokines, produced by immune cells in response to microbial antigens. The functions of these cytokines in the reproductive system are unknown. We examined the expression pattern of IL-24 and IL-29 in human fetal membranes from preterm and term births and in in vitro in response to microbial antigens. fetal membranes collected from cesarean sections at term (normal, not in labor) were placed in culture for 48 h. These membranes were then stimulated with bacterial lipopolysaccharide (LPS) or viral antigen poly-inosinic and cytidylic acid (polyIC) for an additional 24 h. Amniotic fluids (AF) and fetal membranes were also collected from preterm and term deliveries. IL-24 and IL-29 expressions were studied by RT-PCR. ELISA documented culture media and AF cytokine concentrations. IL-24 and IL-29 expressions were seen in cultured fetal membranes regardless of stimulation. Expressions were also found in preterm and term labor membranes, but not in non-labor tissues at term. IL-24 concentrations were higher after LPS stimulation whereas IL-29 concentrations were higher after polyIC-stimulation. AF analysis did not detect either of the cytokines either preterm or term. this is the first study to report IL-24 and IL-29 expressions in human fetal membranes. Higher concentrations of these cytokines in response to distinct infectious stimuli suggest different pathways for fetal immune response during infection.

  14. Enhanced Ca(2+) response and stimulation of prostaglandin release by the bradykinin B2 receptor in human retinal pigment epithelial cells primed with proinflammatory cytokines.

    PubMed

    Catalioto, Rose-Marie; Valenti, Claudio; Maggi, Carlo Alberto; Giuliani, Sandro

    2015-09-15

    Kallikrein, kininogen and kinin receptors are present in human ocular tissues including the retinal pigment epithelium (RPE), suggesting a possible role of bradykinin (BK) in physiological and/or pathological conditions. To test this hypothesis, kinin receptors expression and function was investigated for the first time in human fetal RPE cells, a model close to native RPE, in both control conditions and after treatment with proinflammatory cytokines. Results showed that BK evoked intracellular Ca(2+) transients in human RPE cells by activating the kinin B2 receptor. Pretreatment of the cells with TNF-α and/or IL-1β enhanced Ca(2+) response in a time- and concentration-dependent additive manner, whereas the potency of BK and that of the selective B2 receptor antagonist, fasitibant chloride, both in the nanomolar range, remained unaffected. Cytokines have no significant effect on cell number and viability and on the activity of other GPCRs such as the kinin B1, acetylcholine, ATP and thrombin receptors. Immunoblot analysis and immunofluorescence studies revealed that cytokines treatment was associated with an increase in both kinin B2 receptor and COX-2 expression and with the secretion of prostaglandin E1 and E2 into the extracellular medium. BK, through activation of the kinin B2 receptor, potentiated the COX-2 mediated prostaglandin release in cytokines-primed RPE cells while new protein synthesis and prostaglandin production contribute to the potentiating effect of cytokines on BK-induced Ca(2+) response. In conclusion, overall data revealed a cross-talk between the kinin B2 receptor and cytokines in human RPE in promoting inflammation, a key feature in retinal pathologies including diabetic retinopathy and macular edema. Copyright © 2015 Elsevier Inc. All rights reserved.

  15. Analysis of inflammatory cytokines in human blood, breath condensate, and urine using a multiplex immunoassay platform

    EPA Science Inventory

    A change in the expression of cytokines in human biological media indicates an inflammatory response to external stressors and reflects an early step along the adverse outcome pathway (AOP) for various health endpoints. To characterize and interpret this inflammatory response, m...

  16. Analysis of inflammatory cytokines in human blood, breath condensate, and urine using a multiplex immunoassay platform

    EPA Science Inventory

    A change in the expression of cytokines in human biological media indicates an inflammatory response to external stressors and reflects an early step along the adverse outcome pathway (AOP) for various health endpoints. To characterize and interpret this inflammatory response, m...

  17. The Insect Ortholog of the Human Orphan Cytokine Receptor CRLF3 Is a Neuroprotective Erythropoietin Receptor

    PubMed Central

    Hahn, Nina; Knorr, Debbra Y.; Liebig, Johannes; Wüstefeld, Liane; Peters, Karsten; Büscher, Marita; Bucher, Gregor; Ehrenreich, Hannelore; Heinrich, Ralf

    2017-01-01

    The cytokine erythropoietin (Epo) mediates various cell homeostatic responses to environmental challenges and pathological insults. While stimulation of vertebrate erythrocyte production is mediated by homodimeric “classical” Epo receptors, alternative receptors are involved in neuroprotection. However, their identity remains enigmatic due to complex cytokine ligand and receptor interactions and conflicting experimental results. Besides the classical Epo receptor, the family of type I cytokine receptors also includes the poorly characterized orphan cytokine receptor-like factor 3 (CRLF3) present in vertebrates including human and various insect species. By making use of the more simple genetic makeup of insect model systems, we studied whether CRLF3 is a neuroprotective Epo receptor in animals. We identified a single ortholog of CRLF3 in the beetle Tribolium castaneum, and established protocols for primary neuronal cell cultures from Tribolium brains and efficient in vitro RNA interference. Recombinant human Epo as well as the non-erythropoietic Epo splice variant EV-3 increased the survival of serum-deprived brain neurons, confirming the previously described neuroprotective effect of Epo in insects. Moreover, Epo completely prevented hypoxia-induced apoptotic cell death of primary neuronal cultures. Knockdown of CRLF3 expression by RNA interference with two different double stranded RNA (dsRNA) fragments abolished the neuroprotective effect of Epo, indicating that CRLF3 is a crucial component of the insect Epo-responsive receptor. This suggests that a common urbilaterian ancestor of the orphan human and insect cytokine receptor CRLF3 served as a neuroprotective receptor for an Epo-like cytokine. Our work also suggests that vertebrate CRLF3, like its insect ortholog, might represent a tissue protection-mediating receptor. PMID:28769759

  18. The Insect Ortholog of the Human Orphan Cytokine Receptor CRLF3 Is a Neuroprotective Erythropoietin Receptor.

    PubMed

    Hahn, Nina; Knorr, Debbra Y; Liebig, Johannes; Wüstefeld, Liane; Peters, Karsten; Büscher, Marita; Bucher, Gregor; Ehrenreich, Hannelore; Heinrich, Ralf

    2017-01-01

    The cytokine erythropoietin (Epo) mediates various cell homeostatic responses to environmental challenges and pathological insults. While stimulation of vertebrate erythrocyte production is mediated by homodimeric "classical" Epo receptors, alternative receptors are involved in neuroprotection. However, their identity remains enigmatic due to complex cytokine ligand and receptor interactions and conflicting experimental results. Besides the classical Epo receptor, the family of type I cytokine receptors also includes the poorly characterized orphan cytokine receptor-like factor 3 (CRLF3) present in vertebrates including human and various insect species. By making use of the more simple genetic makeup of insect model systems, we studied whether CRLF3 is a neuroprotective Epo receptor in animals. We identified a single ortholog of CRLF3 in the beetle Tribolium castaneum, and established protocols for primary neuronal cell cultures from Tribolium brains and efficient in vitro RNA interference. Recombinant human Epo as well as the non-erythropoietic Epo splice variant EV-3 increased the survival of serum-deprived brain neurons, confirming the previously described neuroprotective effect of Epo in insects. Moreover, Epo completely prevented hypoxia-induced apoptotic cell death of primary neuronal cultures. Knockdown of CRLF3 expression by RNA interference with two different double stranded RNA (dsRNA) fragments abolished the neuroprotective effect of Epo, indicating that CRLF3 is a crucial component of the insect Epo-responsive receptor. This suggests that a common urbilaterian ancestor of the orphan human and insect cytokine receptor CRLF3 served as a neuroprotective receptor for an Epo-like cytokine. Our work also suggests that vertebrate CRLF3, like its insect ortholog, might represent a tissue protection-mediating receptor.

  19. Role of the cytokine environment and cytokine receptor expression on the generation of functionally distinct dendritic cells from human monocytes.

    PubMed

    Conti, Lucia; Cardone, Marco; Varano, Barbara; Puddu, Patrizia; Belardelli, Filippo; Gessani, Sandra

    2008-03-01

    Myeloid dendritic cells (DC) and macrophages evolve from a common precursor. However, factors controlling monocyte differentiation toward DC or macrophages are poorly defined. We report that the surface density of the GM-CSF receptor (GM-CSFR) alpha subunit in human peripheral blood monocytes varies among donors. Although no correlation was found between the extent of GM-CSFR and monocyte differentiation into DC driven by GM-CSF and IL-4, GM-CSFR expression strongly influenced the generation of CD1a(+) dendritic-like cells in the absence of IL-4. CD1a(+) cells generated in the presence of GM-CSF express CD40, CD80, MHC class I and II, DC-SIGN, MR, CCR5, and partially retain CD14 expression. Interestingly, they spontaneously induce the expansion of CD4(+) and CD8(+) allogeneic T lymphocytes producing IFN-gamma, and migrate toward CCL4 and CCL19. Upon stimulation with TLR ligands, they acquire the phenotypic features of mature DC. In contrast, the allostimulatory capacity is not further increased upon LPS activation. However, by blocking LPS-induced IL-10, a higher T cell proliferative response and IL-12 production were observed. Interestingly, IL-23 secretion was not affected by endogenous IL-10. These results highlight the importance of GM-CSFR expression in monocytes for cytokine-induced DC generation and point to GM-CSF as a direct player in the generation of functionally distinct DC.

  20. Profiles of cytokine and chemokine gene expression in human pulmonary epithelial cells induced by human and avian influenza viruses

    PubMed Central

    2010-01-01

    Influenza pandemic remains a serious threat to human health. In this study, the repertoire of host cellular cytokine and chemokine responses to infections with highly pathogenic avian influenza H5N1, low pathogenicity avian influenza H9N2 and seasonal human influenza H1N1 were compared using an in vitro system based on human pulmonary epithelial cells. The results showed that H5N1 was more potent than H9N2 and H1N1 in inducing CXCL-10/IP-10, TNF-alpha and CCL-5/RANTES. The cytokine/chemokine profiles for H9N2, in general, resembled those of H1N1. Of interest, only H1N1, but none of the avian subtypes examined could induce a persistent elevation of the immune-regulatory cytokine - TGF-β2. The differential expression of cytokines/chemokines following infection with different influenza viruses could be a key determinant for clinical outcome. The potential of using these cytokines/chemokines as prognostic markers or targets of therapy is worth exploring. PMID:21108843

  1. Influence of extracellular matrix on cytokine stimulated pro-labour gene expression in human uterine myocytes.

    PubMed

    Engineer, Neelam; Sooranna, Suren R; Liang, Zhiqing; Bennett, Phillip R; Johnson, Mark R

    2008-11-01

    Cellular function is modulated by the interaction with the extracellular matrix within the myometrium. We formed the hypothesis that the cytokine-stimulated pro-labour gene expression by human uterine smooth muscle cells would be increased by growing the cells on collagen-coated plates. Primary cultures of human uterine smooth muscle cells grown on uncoated plates and on plates coated with collagen were exposed to the inflammatory cytokines (tumor necrosis factor-alpha, interleukin-1beta and interleukin-6) and assessed the messenger RNA expression of oxytocin receptor, interleukin-8, prostaglandin H synthase type-2 and prostaglandin F(2) alpha receptor. Basal pro-labour gene expression was unaffected by collagen coating and the response to the inflammatory cytokines was similar for oxytocin receptor and prostaglandin H synthase type-2, but appeared to be reduced for interleukin-8 and enhanced for FP. Collagen coating made no significant impact on basal integrin expression and interleukin-1beta induced phosphorylation of extracellular-regulated-kinase1/2 and RelA subunit of nuclear factor-kappa B (p65). We conclude that growing human uterine smooth muscle cells on collagen-coated plates may modulate the pro-labour gene response to the inflammatory cytokines.

  2. Acute exposure to methamphetamine alters TLR9-mediated cytokine expression in human macrophage

    PubMed Central

    Burns, Ariel; Ciborowski, Pawel

    2015-01-01

    Recent studies show that methamphetamine (Meth) use leads to higher susceptibility to and progression of infections, which suggests impairment of the immune system. The first line of defense against infections is the innate immune system and the macrophage is a key player in preventing and fighting infections. So we profiled cytokines over time in Meth treated THP-1 cells, as a human macrophage model, at a relevant concentration using high throughput screening to find a signaling target. We showed that after a single exposure, the effect of Meth on macrophage cytokine production was rapid and time dependent and shifted the balance of expression of cytokines to pro-inflammatory. Our results were analogous to previous reports in that Meth up-regulates TNF-α and IL-8 after two hours of exposure. However, global screening led to the novel identification of CXCL16, CXCL1 and many other up-regulated cytokines. We also showed CCL7 as the most down-regulated chemokine due to Meth exposure, which led us to hypothesize that Meth dysregulates the MyD88-dependent Toll-like receptor 9 (TLR9) signaling pathway. In conclusion, altered cytokine expression in macrophages suggests it could lead to a suppressed innate immunity in people who use Meth. PMID:26387832

  3. 12-Lipoxygenase Inhibitor Improves Functions of Cytokine-Treated Human Islets and Type 2 Diabetic Islets.

    PubMed

    Ma, Kaiwen; Xiao, An; Park, So Hyun; Glenn, Lindsey; Jackson, Laura; Barot, Tatvam; Weaver, Jessica R; Taylor-Fishwick, David A; Luci, Diane K; Maloney, David J; Mirmira, Raghavendra G; Imai, Yumi; Nadler, Jerry L

    2017-08-01

    The 12-lipoxygenase (12-LO) pathway produces proinflammatory metabolites, and its activation is implicated in islet inflammation associated with type 1 and type 2 diabetes (T2D). We aimed to test the efficacy of ML355, a highly selective, small molecule inhibitor of 12-LO, for the preservation of islet function. Human islets from nondiabetic donors were incubated with a mixture of tumor necrosis factor α , interluekin-1β, and interferon-γ to model islet inflammation. Cytokine-treated islets and human islets from T2D donors were incubated in the presence and absence of ML355. In vitro study. Human islets from organ donors aged >20 years of both sexes and any race were used. T2D status was defined from either medical history or most recent hemoglobin A1c value >6.5%. Glucose stimulation. Static and dynamic insulin secretion and oxygen consumption rate (OCR). ML355 prevented the reduction of insulin secretion and OCR in cytokine-treated human islets and improved both parameters in human islets from T2D donors. ML355 was efficacious in improving human islet function after cytokine treatment and in T2D islets in vitro. The study suggests that the blockade of the 12-LO pathway may serve as a target for both form of diabetes and provides the basis for further study of this small molecule inhibitor in vivo.

  4. Cytokine expression in human osteoblasts after antiseptic treatment: a comparative study between polyhexanide and chlorhexidine.

    PubMed

    Röhner, Eric; Hoff, Paula; Gaber, Timo; Lang, Annemarie; Vörös, Pauline; Buttgereit, Frank; Perka, Carsten; Windisch, Christoph; Matziolis, Georg

    2015-02-01

    Chlorhexidine and polyhexanide are frequently used antiseptics in clinical practice and have a broad antimicrobial range. Both antiseptics are helpful medical agents for septic wound treatment with a high potential for defeating joint infections. Their effect on human osteoblasts has, so far, not been sufficiently evaluated. The aim of this study was to investigate the activating potential of polyhexanide and chlorhexidine on inflammatory cytokines/chemokines in human osteoblasts in vitro. Human osteoblasts were isolated and cultivated in vitro and then treated separately with 0.1% and 2% chlorhexidine and 0.04% polyhexanide as commonly applied concentrations in clinical practice. Detection of cell structure and cell morphology was performed by light microscopic inspection. Cytokine and chemokine secretion was determined by using a multiplex suspension array. Cell shrinking, defective cell membrane, and the loss of cell adhesion indicated cell damage of human osteoblasts after treatment with both antiseptics was evaluated by using light microscopy. Polyhexanide, but not chlorhexidine, caused human osteoblasts to secrete various interleukins (1β, 6, and 7), interferon γ, tumor necrosis factor α, vascular endothelial growth factor, eotaxin, fibroblast growth factor basic, and granulocyte macrophage colony-stimulating factor as quantified by multiplex suspension array. Both antiseptics induced morphological cell damage at an optimum exposure between 1 and 10 min. But only polyhexanide mediated a pronounced secretion of inflammatory cytokines and chemokines in human osteoblasts. Therefore, we recommend a preferred usage of chlorhexidine in septic surgery to avoid the induction of an inflammatory reaction.

  5. 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.

  6. Human tear analysis with miniaturized multiplex cytokine assay on "wall-less" 96-well plate.

    PubMed

    Le Guezennec, Xavier; Quah, Joanne; Tong, Louis; Kim, Namyong

    2015-01-01

    Tears are a particularly limited body fluid and commonly used in the diagnosis of patients who have ocular diseases. A popular method for analysis of ocular inflammation in tears uses Luminex® bead multiplex technology to generate valuable multiple cytokine profile outputs with 25-50 µl tear sample volume. We propose a method for measuring tear cytokines with 5 μl tear sample volume and 80% reduced Luminex reagents compared to previous protocols. Using human tears pooled from 1,000 participants, the DA-Bead-based method running at 5-20 µl volume, using manual pipetting, in conjunction with a magnetic Luminex cytokine (four-plex) panel assay in a 96-well format was performed and validated for tumor necrosis factor (TNF)-α, interferon (IFN)-γ, interleukin (IL)-1β, and IL-6. Upon use of the DA-Bead method at the 5 μl volume with cytokine standards, the concentrations of each of the four cytokines were found to be linear over a range of 3.5-4 log pg/ml with an intra-assay coefficient of variation (CV) ≤5%, inter-assay %CV ≤10%, and accuracy within the 70-130% range. Upon use of a 5 µl healthy pooled tear sample, cytokine concentrations were detected with a precision intra-assay %CV ˂ 20% for IL-6, IFN-γ, or TNF-α or 30.37% with IL-1β. The inter-assay %CV with tears was ≤20.84% for all cytokines. Tear volumes run at 5 μl on DA-Bead produced a similar cytokine expression profile at a 1-month interval and were highly correlated with the larger 10 μl-based tear sample volume cytokine profile with R(2) = 0.98. DA-Bead assay is highly sensitive and reproducible and has a performance profile that is potentially suitable for use in standard clinical scenarios. Considering the use of as little as 5 µl of assay beads and 5 µl sample, this is also likely to reduce the assay cost significantly and ease diagnosis of patients with ocular diseases.

  7. Human tear analysis with miniaturized multiplex cytokine assay on “wall-less” 96-well plate

    PubMed Central

    Quah, Joanne; Tong, Louis; Kim, Namyong

    2015-01-01

    Purpose Tears are a particularly limited body fluid and commonly used in the diagnosis of patients who have ocular diseases. A popular method for analysis of ocular inflammation in tears uses Luminex® bead multiplex technology to generate valuable multiple cytokine profile outputs with 25–50 µl tear sample volume. We propose a method for measuring tear cytokines with 5 μl tear sample volume and 80% reduced Luminex reagents compared to previous protocols. Methods Using human tears pooled from 1,000 participants, the DA-Bead-based method running at 5–20 µl volume, using manual pipetting, in conjunction with a magnetic Luminex cytokine (four-plex) panel assay in a 96-well format was performed and validated for tumor necrosis factor (TNF)-α, interferon (IFN)-γ, interleukin (IL)-1β, and IL-6. Results Upon use of the DA-Bead method at the 5 μl volume with cytokine standards, the concentrations of each of the four cytokines were found to be linear over a range of 3.5–4 log pg/ml with an intra-assay coefficient of variation (CV) ≤5%, inter-assay %CV ≤10%, and accuracy within the 70–130% range. Upon use of a 5 µl healthy pooled tear sample, cytokine concentrations were detected with a precision intra-assay %CV ˂ 20% for IL-6, IFN-γ, or TNF-α or 30.37% with IL-1β. The inter-assay %CV with tears was ≤20.84% for all cytokines. Tear volumes run at 5 μl on DA-Bead produced a similar cytokine expression profile at a 1-month interval and were highly correlated with the larger 10 μl–based tear sample volume cytokine profile with R2 = 0.98. Conclusions DA-Bead assay is highly sensitive and reproducible and has a performance profile that is potentially suitable for use in standard clinical scenarios. Considering the use of as little as 5 µl of assay beads and 5 µl sample, this is also likely to reduce the assay cost significantly and ease diagnosis of patients with ocular diseases. PMID:26539027

  8. Profiling the human immune response to Mycobacterium tuberculosis by human cytokine array.

    PubMed

    Chen, Tao; Li, Zhenyan; Yu, Li; Li, Haicheng; Lin, Jinfei; Guo, Huixin; Wang, Wei; Chen, Liang; Zhang, Xianen; Wang, Yunxia; Chen, Yuhui; Liao, Qinghua; Tan, Yaoju; Shu, Yang; Huang, Wenyan; Cai, Changhui; Zhou, Zhongjing; Yu, Meiling; Li, Guozhou; Zhou, Lin; Zhong, Qiu; Bi, Lijun; Zhao, Meigui; Guo, Lina; Zhou, Jie

    2016-03-01

    Tuberculosis (TB) continues to be one of the most serious infectious diseases in the world, however, no effective biomarkers can be used for rapid screening of latent tuberculosis infection (LTBI) and active TB. In this study, serum cytokines were screened and tested as potential biomarker for TB diagnosis. Cytokine array was used to track the cytokine profile and its dynamic change after TB infection. The different expressions of cytokines were confirmed by ELISA assay. ROC curve analyses were used to evaluate the efficacy of a cytokine or cytokine combination for diagnosis. Eotaxin-2, ICAM-1, MCSF, IL-12p70, and IL-11 were significantly higher in the LTBI individuals. I-309, MIG, Eotaxin-2, IL-8, ICAM-1, IL-6sR, and Eotaxin were significantly higher in active TB patients. ROC curve analyses gave AUCs of 0.843, 0.898, and 0.888 for I-309, MIG, and IL-8, respectively, and 0.894 for the combination panel in active TB diagnosis. IFN-γ/IL-4 and IL-2/TNF-α ratios exhibit dynamic changes in the healthy control and LTBI to different stages of active TB. Serum cytokines, including I-309 and MIG, IL-8, Extoxin-2, ICAM-1 and combinations of cytokines, including IFN-γ/IL-4 and IL-2/TNF-α, can be used as serum biomarkers for LTBI and active TB screening, thus indicating prospective clinical applications. Copyright © 2016 Elsevier Ltd. All rights reserved.

  9. Efficacy of novel skin cream containing mixture of human growth factors and cytokines for skin rejuvenation.

    PubMed

    Gold, Michael H; Goldman, Mitchel P; Biron, Julie

    2007-02-01

    The crucial role of growth factors and cytokines in cutaneous wound healing is well described, but there has been little investigation into their use for skin rejuvenation. A novel skin cream containing a mixture of human growth factors and cytokines was recently marketed for skin rejuvenation. The mixture was obtained through a biotechnology process using cultured human fetal skin cells, which originate from a dedicated cell bank originally established for the development of products in wound healing. The cream significantly reduced periorbital and perioral wrinkles, as well as improved skin texture of the chin after one month of treatment, which confirms the beneficial use of growth factors and cytokines for skin rejuvenation reported in 2 earlier studies. After 60 days of twice-daily application, 83% of the subject showed an improved average wrinkle score in the eye area, while 50% showed an improved average wrinkle score in the mouth area. In order to exclude a placebo effect, the efficacy of this product should be confirmed with a double-blind, randomized, placebo-controlled study. Also, the difference between mixtures of growth factor and cytokines should be further elaborated.

  10. Tacaribe Virus but Not Junin Virus Infection Induces Cytokine Release from Primary Human Monocytes and Macrophages

    PubMed Central

    Groseth, Allison; Hoenen, Thomas; Weber, Michaela; Wolff, Svenja; Herwig, Astrid; Kaufmann, Andreas; Becker, Stephan

    2011-01-01

    The mechanisms underlying the development of disease during arenavirus infection are poorly understood. However, common to all hemorrhagic fever diseases is the involvement of macrophages as primary target cells, suggesting that the immune response in these cells may be of paramount importance during infection. Thus, in order to identify features of the immune response that contribute to arenavirus pathogenesis, we have examined the growth kinetics and cytokine profiles of two closely related New World arenaviruses, the apathogenic Tacaribe virus (TCRV) and the hemorrhagic fever-causing Junin virus (JUNV), in primary human monocytes and macrophages. Both viruses grew robustly in VeroE6 cells; however, TCRV titres were decreased by approximately 10 fold compared to JUNV in both monocytes and macrophages. Infection of both monocytes and macrophages with TCRV also resulted in the release of high levels of IL-6, IL-10 and TNF-α, while levels of IFN-α, IFN-β and IL-12 were not affected. However, we could show that the presence of these cytokines had no direct effect on growth of either TCRV of JUNV in macrophages. Further analysis also showed that while the production of IL-6 and IL-10 are dependent on viral replication, production of TNF-α also occurs after exposure to UV-inactivated TCRV particles and is thus independent of productive virus infection. Surprisingly, JUNV infection did not have an effect on any of the cytokines examined indicating that, in contrast to other viral hemorrhagic fever viruses, macrophage-derived cytokine production is unlikely to play an active role in contributing to the cytokine dysregulation observed in JUNV infected patients. Rather, these results suggest that an early, controlled immune response by infected macrophages may be critical for the successful control of infection of apathogenic viruses and prevention of subsequent disease, including systemic cytokine dysregulation. PMID:21572983

  11. Cytokine expression profile in human lungs undergoing normothermic ex-vivo lung perfusion.

    PubMed

    Sadaria, Miral R; Smith, Phillip D; Fullerton, David A; Justison, George A; Lee, Joon H; Puskas, Ferenc; Grover, Frederick L; Cleveland, Joseph C; Reece, T Brett; Weyant, Michael J

    2011-08-01

    A donor lung shortage prevents patients from receiving life-saving transplants. Ex-vivo lung perfusion (EVLP) is a viable means of expanding the donor pool by evaluating and potentially improving donor lung function. The metabolic and inflammatory effects of EVLP on human lung tissue are currently unknown. We sought to establish representative cytokine expression in human donor lungs meeting acceptable lung transplant criteria after prolonged normothermic EVLP. Seven single human lungs not meeting traditional transplantation criteria for various reasons underwent normothermic EVLP. Lungs were perfused with deoxygenated colloid, rewarmed, and ventilated per standard protocol. Lung function was evaluated every hour. Biopsies were taken at 1, 6, and 12 hours. Inflammatory cytokines were quantitatively measured using a human cytokine magnetic bead-based multiplex assay. All lungs met traditional transplant criteria after EVLP. The partial pressure of arterial oxygen and physiologic lung function significantly improved (p<0.05). No pulmonary edema was formed, and histology demonstrated no evidence of acute lung injury. Interleukin (IL)-6, IL-8, granulocyte colony-stimulating factor, and monocyte chemotactic protein-1 were upregulated, while granulocyte macrophage colony-stimulating factor was downregulated during EVLP (p<0.05). IL-1β, IL-4, IL-7, IL-12, interferon-γ, macrophage inflammatory protein-1β, and tumor necrosis factor-α were detectable and unchanged. Ex-vivo lung perfusion demonstrates the ability to improve oxygenation and physiologic lung function in donor lungs unacceptable for transplantation without injury to the lung. We establish here a cytokine expression profile in human lungs undergoing normothermic EVLP. These data can be used in the future to explore novel targeted therapies for ischemia-reperfusion injury. Copyright © 2011 The Society of Thoracic Surgeons. Published by Elsevier Inc. All rights reserved.

  12. A conserved motif in the promoters of several cytokines expressed by human Th2-type lymphocytes.

    PubMed

    Staynov, D Z; Cousins, D J; Lee, T H

    1995-01-01

    We have recently found a novel conserved motif in the promoters of several T-cell-expressed cytokines [human interleukin-2, -4, -5 and -13 and human and mouse granulocyte/macrophage-colony stimulating factor (GM-CSF)]. It contains a core sequence CTTGG ... CCAAG which is present as part of larger palindromic sequences in each gene. This suggest that they may interact with a new family of trans-acting factors. In transfection assays, the human GM-CSF element has a strong positive effect on the expression of a reporter gene by the human T cell line Jurkat J6 upon stimulation. In DNA mobility shift assays, this sequence can give either six different specific bands which are competed out by different parts of the sequence or one specific band which is competed out by each of the inverted repeats, depending on the reconstitution conditions. In different genes, the core sequences are separated by integer numbers of helical turns. Considering the strong positive regulatory effect of this element and its presence in several T-cell-expressed cytokine genes, it may be crucial to the coordinated expression of these cytokines in T helper cells.

  13. IL-10 is excluded from the functional cytokine memory of human CD4+ memory T lymphocytes.

    PubMed

    Dong, Jun; Ivascu, Claudia; Chang, Hyun-Dong; Wu, Peihua; Angeli, Roberta; Maggi, Laura; Eckhardt, Florian; Tykocinski, Lars; Haefliger, Carolina; Möwes, Beate; Sieper, Jochen; Radbruch, Andreas; Annunziato, Francesco; Thiel, Andreas

    2007-08-15

    Epigenetic modifications, including DNA methylation, profoundly influence gene expression of CD4(+) Th-specific cells thereby shaping memory Th cell function. We demonstrate here a correlation between a lacking fixed potential of human memory Th cells to re-express the immunoregulatory cytokine gene IL10 and its DNA methylation status. Memory Th cells secreting IL-10 or IFN-gamma were directly isolated ex vivo from peripheral blood of healthy volunteers, and the DNA methylation status of IL10 and IFNG was assessed. Limited difference in methylation was found for the IL10 gene locus in IL-10-secreting Th cells, as compared with Th cells not secreting IL-10 isolated directly ex vivo or from in vitro-established human Th1 and Th2 clones. In contrast, in IFN-gamma(+) memory Th cells the promoter of the IFNG gene was hypomethylated, as compared with IFN-gamma-nonsecreting memory Th cells. In accordance with the lack of epigenetic memory, almost 90% of ex vivo-isolated IL-10-secreting Th cells lacked a functional memory for IL-10 re-expression after restimulation. Our data indicate that IL10 does not become epigenetically marked in human memory Th cells unlike effector cytokine genes such as IFNG. The exclusion of IL-10, but not effector cytokines, from the functional memory of human CD4(+) T lymphocytes ex vivo may reflect the need for appropriate regulation of IL-10 secretion, due to its potent immunoregulatory potential.

  14. Cytokines derived from activated human mononuclear cells markedly stimulate transferrin secretion by cultured Sertoli cells.

    PubMed

    Hoeben, E; Van Damme, J; Put, W; Swinnen, J V; Verhoeven, G

    1996-02-01

    There is considerable evidence that Sertoli cell function is controlled not only by hormones, but also by locally produced growth factors and cytokines. To gain more insight into the nature and effects of cytokines potentially involved in the control of Sertoli cell function, we incubated rat Sertoli cells with media conditioned by activated human peripheral blood mononuclear cells. Such media (PBMC-CM) are known to be an extremely rich source of a variety of cytokines. It was demonstrated that PMBC-CM and protein fractions derived from them stimulate Sertoli cell transferrin secretion and messenger RNA production more potently then peritubular cell-conditioned medium or FIRT (a combination of FSH, insulin, retinol, and testosterone). Transferrin secretion expressed per mg cell DNA was stimulated approximately 5-fold by peritubular cell-conditioned medium or FIRT and nearly 20-fold by PBMC-CM. The effects of PBMC-CM were accompanied by a limited increase in cAMP and a noticeable rise in cGMP. Affinity chromatography on a column coated with an antiserum directed against interleukin-1 beta (IL-1 beta) showed that part of the activity in the PBMC-CM was related to IL-1 beta. The remainder of the activity was largely retained by an affinity column coated with an antiserum that recognizes IL-6 and a number of other known and unknown cytokines. Purified IL-1 beta provoked a 2- to 3-fold stimulation of Sertoli cell transferrin secretion. More limited stimulatory effects were observed with IL-6. Neither of these cytokines or their combination approached the degree of stimulation observed with crude PBMC-CM, suggesting that other cytokines are involved. It is concluded that the mixture of cytokines present in PBMC-CM is a more powerful stimulator of Sertoli cell transferrin secretion than any other agonist known at the present time. IL-1 and IL-6 may be responsible for part of the observed effects, but one or more other cytokines are probably involved.

  15. The production of immunoregulatory cytokines is localized to the extrafollicular area of human tonsils.

    PubMed

    Agren, K; Andersson, U; Litton, M; Funa, K; Nordlander, B; Andersson, J

    1996-05-01

    The localization and production at the single cell level of 19 different human cytokines, IL-1 alpha, IL-1 beta, IL-1ra, IL-2, IL-3, IL-4, IL-5, IL-6, IL-8, IL-10, IL-13, TNF alpha, TNF beta, IFN gamma, GM-CSF, G-CSF, and TGF beta 1-3, were studied in cryopreserved tonsillar tissue using immunohistochemical staining. The cytokine producing cells, with the exception of IL-1 expressing cells, had a characteristic morphology due to the accumulation of cytokine onto the Golgi organelle. The production of each cytokine was localized to specific compartments in tonsillar tissue sections from children with tonsillar hypertrophy or recurrent tonsillitis in the resting state. Immunoregulatory cytokines such as IL-2, IL-3, IL-4, G-CSF, GM-CSF and TGF beta were produced in the extrafollicular area and entrapped on the cell membranes as well as in pudels in the extracellular matrix surrounding the producer cells. The dominating cytokines both in tissues from recurrent tonsillitis and tonsillar hypertrophy were GM-CSF, G-CSF, and TGF beta 1-3 which were synthezised predominantly in the reticular crypt site. IL-1 alpha, beta and IL-1ra, on the other hand, were localized to the surface and crypt epithelium and to scattered regions in the extrafollicular area. IL-2, IL-6, IFN gamma and IL-10 were found much more often in sections obtained from recurrent tonsillitis tissue compared with those from tonsillar hypertrophy. Reversely, an excessive production of IL-4 was noted in tonsillar hypertrophy compared with that in recurrent tonsillitis. Thus, concomitant production of multiple cytokines was evident with similarities but also differences in cytokine pattern between the two groups studied. The data suggest that T-cell mediated B-cell activation and differentiation take place in the extrafollicular area. Children with recurrent tonsillitis had a higher amount of B-cells and monocytes compared with children with tonsillar hypertrophy. However, the number of CD3, CD4, CD8 or

  16. Lactobacillus acidophilus and L. reuteri modulate cytokine responses in gnotobiotic pigs infected with human rotavirus

    PubMed Central

    Azevedo, M. S. P.; Zhang, W.; Wen, K.; Gonzalez, A. M.; Saif, L. J.; Yousef, A. E.; Yuan, L.

    2012-01-01

    Probiotic lactic acid bacteria (LAB) have been shown to alleviate inflammation, enhance the immunogenicity of rotavirus vaccines, or reduce the severity of rotavirus diarrhoea. Although the mechanisms are not clear, the differential Th1/Th2/Th3-driving capacities and modulating effects on cytokine production of different LAB strains may be the key. Our goal was to delineate the influence of combining two probiotic strains L. acidophilus and L. reuteri on the development of cytokine responses in neonatal gnotobiotic pigs infected with human rotavirus (HRV). We demonstrated that HRV alone, or HRV plus LAB, but not LAB alone, initiated serum cytokine responses, as indicated by significantly higher concentrations of IFN-α, IFN-γ, IL-12, and IL-10 at post-inoculation day (PID) 2 in the HRV only and LAB+HRV+ pigs compared to LAB only and LAB-HRV- pigs. Peak cytokine responses coincided with the peak of HRV replication. LAB further enhanced the Th1 and Th2 cytokine responses to HRV infection as indicated by significantly higher concentrations of IL-12, IFN-γ, IL-4 and IL-10 in the LAB+HRV+ pigs compared to the LAB-HRV+ pigs. The LAB+HRV+ pigs maintained relatively constant concentrations of TGF-β compared to the HRV only group which had a significant increase at PID 2 and decrease at PID 7, suggesting a regulatory role of LAB in maintaining gut homeostasis. At PID 28, cytokine secreting cell (CSC) responses, measured by ELISpot, showed increased Th1 (IL-12, IFN-γ) CSC numbers in the LAB+HRV+ and LAB-HRV+ groups compared to LAB only and LAB-HRV- pigs, with significantly increased IL-12 CSCs in spleen and PBMCs and IFN-γ CSCs in spleen of the LAB+HRV+ group. Thus, HRV infection alone, but not LAB alone was effective in inducing cytokine responses but LAB significantly enhanced both Th1 and Th2 cytokines in HRV-infected pigs. LAB may also help to maintain immunological homeostasis during HRV infection by regulating TGF-β production. PMID:22348907

  17. Cross-Regulation of Proinflammatory Cytokines by Interleukin-10 and miR-155 in Orientia tsutsugamushi-Infected Human Macrophages Prevents Cytokine Storm.

    PubMed

    Tsai, Ming-Hsien; Chang, Chung-Hsing; Tsai, Rong-Kung; Hong, Yi-Ren; Chuang, Tsung-Hsien; Fan, Kan-Tang; Peng, Chi-Wen; Wu, Ching-Ying; Hsu, Wen-Li; Wang, Lih-Shinn; Chen, Li-Kuang; Yu, Hsin-Su

    2016-07-01

    Scrub typhus is caused by the obligate intracellular bacterium Orientia tsutsugamushi. Macrophages are host cells for its replication and clearance. Severe complications in patients are mainly caused by a cytokine storm resulting from overproduction of proinflammatory cytokines; nevertheless, the molecular mechanism for the occurrence remains obscure. Herein, we investigate the interactive regulation of cytokines and micro-RNA (miR) in human macrophages infected with low and high doses of O. tsutsugamushi. During low dose infection, macrophages produce high levels of IL-10 through extracellular signal-regulated kinase activation, which inhibits proinflammatory cytokine production and facilitates pathogen replication. Increasing levels of pathogen results in reduced levels of IL-10, and macrophages begin to generate high levels of proinflammatory cytokines through NF-κB activation. However, during a high dose infection, macrophages produce high levels of miR-155 to slow the proinflammatory response. The extracellular signal-regulated kinase/IL-10 axis suppresses the NF-κB/tumor necrosis factor alpha axis via activation of signal transducer and activator of transcription 3. Both IL-10 and miR-155 inhibit the NF-κB signaling pathway. Furthermore, IL-10 is a potent inhibitor of miR-155. Patients susceptible to a cytokine storm, peripheral blood mononuclear cells showed significantly lower IL-10 and miR-155 responses to O. tsutsugamushi challenge. Thus, IL-10 and miR-155 operate inhibitory mechanisms to achieve a proper defense mechanism and prevent a cytokine storm. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

  18. Gemifloxacin inhibits cytokine secretion by lipopolysaccharide stimulated human monocytes at the post-transcriptional level.

    PubMed

    Araujo, F; Slifer, T; Li, S; Kuver, A; Fong, L; Remington, J

    2004-03-01

    The fluroquinolone gemifloxacin was examined for its capacity to modulate secretion of cytokines by human monocytes stimulated with lipopolysaccharide (LPS). Monocytes from six male and two female healthy volunteers were stimulated with LPS, exposed to gemifloxacin and the amounts of secreted IL-1 alpha, IL-1 beta, IL-6, IL-10 and TNF-alpha measured at 3, 6 and 24 h. The results revealed that LPS alone increased secretion of each cytokine significantly. Treatment of the LPS-stimulated monocytes with gemifloxacin resulted in a significant inhibition (p < 0.01) of secretion of each of the cytokines from monocytes of the eight volunteers. Nuclear extracts of the human monocyte cell line, THP-1, were used in the electrophoretic mobility shift assay to determine whether gemifloxacin affects nuclear factor-kappa B (NF-kappa B) activation. In addition, RNA from THP-1 cells was used in Northern blots to determine whether inhibition of secretion of IL-1 beta and TNF-alpha by gemifloxacin occurred at the transcription or translation level. Whereas LPS induced a rapid increase in NF-kappa B activation, gemifloxacin alone did not. Gemifloxacin did not affect the kinetics or decrease the extent of activation. Northern blots indicated that the inhibitory activity of gemifloxacin occurred post-transcription. Thus, gemifloxacin may modulate the immune response by altering secretion of cytokines by human monocytes. Although the concentrations of gemifloxacin used were higher than those observed in the serum of human volunteers treated with the dose under clinical development, it should be taken into consideration that concentrations at tissue and intracellular levels may be considerably higher than serum concentrations.

  19. Recombinant human interleukin-1 receptor antagonist promotes M1 microglia biased cytokines and chemokines following human traumatic brain injury.

    PubMed

    Helmy, Adel; Guilfoyle, Mathew R; Carpenter, Keri Lh; Pickard, John D; Menon, David K; Hutchinson, Peter J

    2016-08-01

    Interleukin-1 receptor antagonist (IL1ra) has demonstrated efficacy in a wide range of animal models of neuronal injury. We have previously published a randomised controlled study of IL1ra in human severe TBI, with concomitant microdialysis and plasma sampling of 42 cytokines and chemokines. In this study, we have used partial least squares discriminant analysis to model the effects of drug administration and time following injury on the cytokine milieu within the injured brain. We demonstrate that treatment with rhIL1ra causes a brain-specific modification of the cytokine and chemokine response to injury, particularly in samples from the first 48 h following injury. The magnitude of this response is dependent on the concentration of IL1ra achieved in the brain extracellular space. Chemokines related to recruitment of macrophages from the plasma compartment (MCP-1) and biasing towards a M1 microglial phenotype (GM-CSF, IL1) are increased in patient samples in the rhIL1ra-treated patients. In control patients, cytokines and chemokines biased to a M2 microglia phenotype (IL4, IL10, MDC) are relatively increased. This pattern of response suggests that a simple classification of IL1ra as an 'anti-inflammatory' cytokine may not be appropriate and highlights the importance of the microglial response to injury.

  20. Analysis of inflammatory cytokines in human blood, breath condensate, and urine using a multiplex immunoassay platform.

    PubMed

    Stiegel, Matthew A; Pleil, Joachim D; Sobus, Jon R; Morgan, Marsha K; Madden, Michael C

    2015-02-01

    A change in the expression of cytokines in human biological media indicates an inflammatory response to external stressors and reflects an early step along the adverse outcome pathway (AOP) for various health endpoints. To characterize and interpret this inflammatory response, methodology was developed for measuring a suite of 10 different cytokines in human blood, exhaled breath condensate (EBC), and urine using an electrochemiluminescent multiplex Th1/Th2 cytokine immunoassay platform. Measurement distributions and correlations for eight interleukins (IL) (1β, 2, 4, 5, 8, 10, 12p70 and 13), interferon-γ (IFN-γ), and tumor necrosis factor-α (TNF-α) were evaluated using 90 blood plasma, 77 EBC, and 400 urine samples collected from nominally healthy adults subjects in North Carolina in 2008-2012. The in vivo results show that there is sufficient sensitivity for characterizing all 10 cytokines at levels of 0.05-0.10 ρg/ml with a dynamic range up to 100 ng/ml across all three of these biological media. The measured in vivo results also show that the duplicate analysis of blood, EBC and urine samples have average estimated fold ranges of 2.21, 3.49, and 2.50, respectively, which are similar to the mean estimated fold range (2.88) for the lowest concentration (0.610 ρg/ml) from a series of spiked control samples; the cytokine method can be used for all three biological media. Nine out of the 10 cytokines measured in EBC were highly correlated within one another with Spearman ρ coefficients ranging from 0.679 to 0.852, while the cytokines measured in blood had a mix of negative and positive correlations, ranging from -0.620 to 0.836. Almost all correlations between EBC and blood were positive. This work also represents the first successful within- and between-person evaluation of ultra trace-level inflammatory markers in blood, EBC, and urine.

  1. Distinct cytokine release profiles from human endothelial and THP-1 macrophage-like cells exposed to different amphotericin B formulations.

    PubMed

    Turtinen, Lloyd W; Bremer, Lindsay A; Prall, David N; Schwartzhoff, Jenifer; Hartsel, Scott C

    2005-01-01

    Amphotericin B(AmB) formulations, Fungizone, and Amphotec caused substantially greater proinflammatory cytokine release than AmBisome (L-AMB) and Abelcet in TPA differentiated THP-1 macrophages as determined by antibody based protein arrays. Lipopolysaccharide but not AmB induced significant pro-inflammatory cytokines in human endothelial cells.

  2. Nitric oxide promotes infectious bone resorption by enhancing cytokine-stimulated interstitial collagenase synthesis in osteoblasts.

    PubMed

    Lin, Sze-Kwan; Kok, Sang-Heng; Kuo, Mark Yen-Ping; Lee, Ming-Shu; Wang, Chih-Chiang; Lan, Wan-Hong; Hsiao, Michael; Goldring, Steven R; Hong, Chi-Yuan

    2003-01-01

    This experiment was undertaken to determine the role of macrophage-derived nitric oxide (NO) in mediating lipopolysaccharide (LPS)-induced bone resorption by using an in vitro co-culture system and an in vivo model of infectious bone resorption. Our results demonstrated that LPS stimulated the expression of inducible nitric oxide synthase (iNOS) and tumor necrosis factor (TNF)-a mRNAs and nitrite synthesis in the J774 mouse macrophage cell line but not in the UMR-106 (rat) and MC3T3-E1 (mouse) osteoblast cell lines. Conditioned media (CM) from LPS-stimulated J774 triggered only low to moderate levels of iNOS mRNAs in MC3T3-E1 and a trivial effect in UMR-106. On the other hand, CM induced matrix metalloproteinase-1 (MMP-1) gene expression in both osteoblast cell lines. The NOS inhibitor N(G)-monomethyl-L-arginine (L-NMMA) did not alter this effect in MC3T3-E1 and UMR-106, whereas TNF-a antibody diminished the CM-induced MMP-1 gene expression in both cell lines. Interestingly, SNAP, a NO donor, although by itself is not a MMP-1 stimulator for UMR-106, augmented the TNF-alpha-stimulated MMP-1 mRNA production in UMR-106. In a J774/UMR-106 co-culture system, LPS stimulated significant MMP-1 gene expression in UMR-106, and this upregulation was abolished by L-NMMA and TNF-alpha antibodies. Immunohistochemical analysis in a rat model of infectious bone resorption (periapical lesion) showed co-distributions of iNOS+ macrophages and MMP-1+ osteoblasts around the osteolytic areas. Administration of L-NMMA markedly reduced the extent of bone loss and the percentage of MMP-1-synthesizing osteoblasts. These data suggest that NO derived from macrophages after LPS stimulation may enhance bone loss by augmenting the cytokine-induced MMP-1 production in osteoblasts.

  3. A High-Dimensional Atlas of Human T Cell Diversity Reveals Tissue-Specific Trafficking and Cytokine Signatures.

    PubMed

    Wong, Michael Thomas; Ong, David Eng Hui; Lim, Frances Sheau Huei; Teng, Karen Wei Weng; McGovern, Naomi; Narayanan, Sriram; Ho, Wen Qi; Cerny, Daniela; Tan, Henry Kun Kiaang; Anicete, Rosslyn; Tan, Bien Keem; Lim, Tony Kiat Hon; Chan, Chung Yip; Cheow, Peng Chung; Lee, Ser Yee; Takano, Angela; Tan, Eng-Huat; Tam, John Kit Chung; Tan, Ern Yu; Chan, Jerry Kok Yen; Fink, Katja; Bertoletti, Antonio; Ginhoux, Florent; Curotto de Lafaille, Maria Alicia; Newell, Evan William

    2016-08-16

    Depending on the tissue microenvironment, T cells can differentiate into highly diverse subsets expressing unique trafficking receptors and cytokines. Studies of human lymphocytes have primarily focused on a limited number of parameters in blood, representing an incomplete view of the human immune system. Here, we have utilized mass cytometry to simultaneously analyze T cell trafficking and functional markers across eight different human tissues, including blood, lymphoid, and non-lymphoid tissues. These data have revealed that combinatorial expression of trafficking receptors and cytokines better defines tissue specificity. Notably, we identified numerous T helper cell subsets with overlapping cytokine expression, but only specific cytokine combinations are secreted regardless of tissue type. This indicates that T cell lineages defined in mouse models cannot be clearly distinguished in humans. Overall, our data uncover a plethora of tissue immune signatures and provide a systemic map of how T cell phenotypes are altered throughout the human body.

  4. [Modulation of the oxidative burst of human neutrophils by pro- and anti-inflammatory cytokines].

    PubMed

    Gougerot-Podicalo, M A; Elbim, C; Chollet-Martin, S

    1996-01-01

    Human polymorphonuclear neutrophils (PMN) and cytokines play a critical role in host defences against invading microorganisms. In response to a variety of stimuli, PMN are a major source of reactive oxygen species (ROS) which are essential for bacterial killing and may induce oxidative stress in tissue environment. A precise regulation of the oxidase activity is therefore necessary. Cytokines such as TNF alpha, GM-CSF, IL-8, IL-6, IL-1 alpha and IL-1 beta produced during the immune and inflammatory responses to pathogens have been reported to interact with PMN activities. However, contradictory results have been reported on their direct and priming effects on the PMN release of ROS (oxidative burst). We have used a flow cytometry method to study the effects of these cytokines on the oxidative burst of PMN in whole blood, in order to avoid PMN activation related to isolation procedures. None of the cytokines tested directly activated the PMN oxidative burst, but they did have differential priming effects on the oxidative burst in response to N-formyl peptides. TNF, GM-CSF and IL-8 strongly primed a subpopulation of PMN to produce H2O2 in response to fMLP, while IL-1 alpha, IL-1 beta and IL-6 failed to do so. Furthermore, the addition of TNF, GM-CSF or IL-8 to whole blood increased the capacity of a subpopulation of PMN to bind N-formyl peptides, a phenomenon that could account for the strong H2O2 production in response to fMLP following priming by the cytokines. These results show that, among the various cytokines tested, TNF, GM-CSF and IL-8 strongly prime the PMN oxidative burst in response to bacterial peptides in whole blood and suggest that these cytokines may play a critical role in bacterial killing in vivo and also in the surrounding tissue injury secondary to pathological inflammatory reactions. In particular, TNF and IL-8 plasma levels as well as LPS-induced monocytic production of these cytokines ex vivo have been correlated with the production of ROS by

  5. Selected scorpion toxin exposures induce cytokine release in human peripheral blood mononuclear cells.

    PubMed

    Corzo, Gerardo; Espino-Solis, Gerardo Pavel

    2017-03-01

    A cytokine screening on human peripheral blood mononuclear cells (PBMCs) stimulated with selected scorpion toxins (ScTx's) was performed in order to evaluate their effect on human immune cells. The ScTx's chosen for this report were three typical buthid scorpion venom peptides, one with lethal effects on mammals Centruroides suffussus suffusus toxin II (CssII), another, with lethal effects on insects and crustaceans Centruroides noxius toxin 5 (Cn5), and one more without lethal effects Tityus discrepans toxin (Discrepin). A Luminex multiplex analysis was performed in order to determine the amounts chemokines and cytokines IL-1β, IL-2, IL-4, IL-5, IL-6, IL-10, IL-12-p40, IL-13, interferon alpha (IFN-α), interferon gamma (IFN-γ), tumor necrosis factor alpha TNF-α, and interferon-inducible protein-10 (IP-10) secreted from human PBMCs exposed to these toxins. Although, the ScTx Cn5 is not lethal for mammals, it was able to induce the secretion of cytokines IL-1β, IL-6, and TNF-α, IL-10 and IP-10 in comparison to the lethal CssII, which was able to induce only IP-10 secretion. Discrepin also was able to induce only IP-10. Interestingly, only low amounts of interferons α and β were induced in the presence of the ScTx's assayed. In a synergic experiment, the combination of Discrepin and Cn5 displayed considerable reverse effects on induction of IL-1β, IL-6, IL-10 and TNF-α, but they had a slight synergic effect on IP-10 cytokine production in comparison with the single effect obtained with the Cn5 alone. Thus, the results obtained suggest that the profile of secreted cytokines promoted by ScTx Cn5 is highly related with a cytokine storm event, and also it suggests that the mammalian lethal neurotoxins are not solely responsible of the scorpion envenomation symptomatology. Copyright © 2017 Elsevier Ltd. All rights reserved.

  6. The human carotid body releases acetylcholine, ATP and cytokines during hypoxia.

    PubMed

    Kåhlin, Jessica; Mkrtchian, Souren; Ebberyd, Anette; Hammarstedt-Nordenvall, Lalle; Nordlander, Britt; Yoshitake, Takashi; Kehr, Jan; Prabhakar, Nanduri; Poellinger, Lorenz; Fagerlund, Malin Jonsson; Eriksson, Lars I

    2014-08-01

    Studies on experimental animals established that the carotid bodies are sensory organs for detecting arterial blood O2 levels and that the ensuing chemosensory reflex is a major regulator of cardiorespiratory functions during hypoxia. However, little information is available on the human carotid body responses to hypoxia. The present study was performed on human carotid bodies obtained from surgical patients undergoing elective head and neck cancer surgery. Our results show that exposing carotid body slices to hypoxia for a period as brief as 5 min markedly facilitates the release of ACh and ATP. Furthermore, prolonged hypoxia for 1 h induces an increased release of interleukin (IL)-1β, IL-4, IL-6, IL-8 and IL-10. Immunohistochemical analysis revealed that type 1 cells of the human carotid body express an array of cytokine receptors as well as hypoxia-inducible factor-1α and hypoxia-inducible factor-2α. Taken together, these results demonstrate that ACh and ATP are released from the human carotid body in response to hypoxia, suggesting that these neurotransmitters, as in several experimental animal models, play a role in hypoxic signalling also in the human carotid body. The finding that the human carotid body releases cytokines in response to hypoxia adds to the growing body of information suggesting that the carotid body may play a role in detecting inflammation, providing a link between the immune system and the nervous system.

  7. Chemically Modified N-Acylated Hyaluronan Fragments Modulate Proinflammatory Cytokine Production by Stimulated Human Macrophages*

    PubMed Central

    Babasola, Oladunni; Rees-Milton, Karen J.; Bebe, Siziwe; Wang, Jiaxi; Anastassiades, Tassos P.

    2014-01-01

    Low molecular mass hyaluronans are known to induce inflammation. To determine the role of the acetyl groups of low molecular mass hyaluronan in stimulating the production of proinflammatory cytokines, partial N-deacetylation was carried out by hydrazinolysis. This resulted in 19.7 ± 3.5% free NH2 functional groups, which were then acylated by reacting with an acyl anhydride, including acetic anhydride. Hydrazinolysis resulted in bond cleavage of the hyaluronan chain causing a reduction of the molecular mass to 30–214 kDa. The total NH2 and N-acetyl moieties in the reacetylated hyaluronan were 0% and 98.7 ± 1.5% respectively, whereas for butyrylated hyaluronan, the total NH2, N-acetyl, and N-butyryl moieties were 0, 82.2 ± 4.6, and 22.7 ± 3.8%, respectively, based on 1H NMR. We studied the effect of these polymers on cytokine production by cultured human macrophages (THP-1 cells). The reacetylated hyaluronan stimulated proinflammatory cytokine production to levels similar to LPS, whereas partially deacetylated hyaluronan had no stimulatory effect, indicating the critical role of the N-acetyl groups in the stimulation of proinflammatory cytokine production. Butyrylated hyaluronan significantly reduced the stimulatory effect on cytokine production by the reacetylated hyaluronan or LPS but had no stimulatory effect of its own. The other partially N-acylated hyaluronan derivatives tested showed smaller stimulatory effects than reacetylated hyaluronan. Antibody and antagonist experiments suggest that the acetylated and partially butyrylated lower molecular mass hyaluronans exert their effects through the TLR-4 receptor system. Selectively N-butyrylated lower molecular mass hyaluronan shows promise as an example of a novel semisynthetic anti-inflammatory molecule. PMID:25053413

  8. Antitumor actions of cytokines on new human papillary thyroid carcinoma cell lines.

    PubMed

    Ohta, K; Pang, X P; Berg, L; Hershman, J M

    1996-07-01

    To investigate the in vitro effects of cytokines on the growth of human papillary thyroid carcinoma (PTC) cells, we established six new PTC cell lines, designated BHP 5, 14, 15, 17, 18, and 19, from different patients. We studied the antiproliferative actions of cytokines by using BHP cells, NP cells (PTC cell line), and ARO cells (anaplastic thyroid carcinoma cell line). These cells were treated with various concentrations of tumor necrosis factor-alpha (TNF alpha), interferon-gamma (IFN gamma), interleukin-1 beta (IL-1 beta), and transforming growth factor-beta 1 (TGF beta 1), alone and in combination. Cell proliferation was assessed by [3H]thymidine incorporation and cell number measurement. In BHP cell lines, IFN gamma, IL-1 beta, and TGF beta 1 inhibited [3H]thymidine incorporation and decreased cell number, but TNF alpha stimulated [3H]thymidine incorporation. In NP cells, treatment with each cytokine decreased [3H]thymidine incorporation and cell number. In contrast, the proliferation of ARO cells was either stimulated by or resistant to TNF alpha, IL-1 beta, and TGF beta 1. The effects of these cytokines on [3H]thymidine incorporation were additive in these cell lines. The results suggest that IL-1 beta and TGF beta 1 play a pivotal role in growth inhibition of PTC cells, and the escape from negative control of IL-1 beta and TGF beta 1 may be a step toward anaplastic changes. The additive effects of these cytokines suggest that they act through different pathways.

  9. Cytokine-mediated inhibition of fibrillar amyloid-β peptide degradation by human mononuclear phagocytes1

    PubMed Central

    Yamamoto, Masaru; Kiyota, Tomomi; Walsh, Shannon M.; Liu, Jianuo; Kipnis, Jonathan; Ikezu, Tsuneya

    2008-01-01

    Vaccination therapy of AD animal models and patients strongly suggests an active role of brain mononuclear phagocytes in immune-mediated clearance of amyloid-β peptides (Aβ) in brain. Although Aβ uptake by macrophages can be regulated by pro- and anti-inflammatory cytokines, their effects on macrophage-mediated Aβ degradation are poorly understood. To better understand this mechanism of degradation, we examined whether pro- and anti-inflammatory cytokines affect the degradation of Aβ using primary cultured human monocyte-derived macrophages (MDM) and microglia using pulse-chase analysis of fibrillar and oligomer 125I-Aβ40 and Aβ42. Initial uptake of fibrillar Aβ40 and Aβ42 was 40% and its degradation was saturated by 120 hrs in both MDM and microglia, compared to an initial uptake of oligomeric Aβ less than 0.5% and saturation of degradation within 24 hrs. Interferon-γ (IFN-γ) increased the intracellular retention of fibrillar Aβ40 and Aβ42 by inhibiting degradation, whereas interleukin-4 (IL-4), IL-10, and transforming growth factor-β1 (TGF-β1), but not IL-13 and IL-27, enhanced degradation. Fibrillar Aβ degradation in MDM is sensitive to lysosomal and insulin degrading enzyme (IDE) inhibitors but insensitive to proteasomal and neprilysin inhibitors. IFN-γ and TNF-α directly reduced the expression of IDE and chaperone molecules (Hsp70 and Hsc70), which are involved in refolding of aggregated proteins. Co-culture of MDM with activated, but not naïve T cells, suppressed Aβ degradation in MDM, which was partially blocked by a combination of neutralizing antibodies against pro-inflammatory cytokines. These data suggest that pro-inflammatory cytokines suppress Aβ degradation in MDM, whereas select anti-inflammatory and regulatory cytokines antagonize these effects. PMID:18768842

  10. Involvement of cytokines in human immunodeficiency virus-1 protein Tat and methamphetamine interactions in the striatum.

    PubMed

    Theodore, Shaji; Cass, Wayne A; Maragos, William F

    2006-06-01

    Human immunodeficiency virus-1 (HIV-1) infection of the brain causes elevation in pro-inflammatory cytokines and inflammatory changes in the striatum. HIV-1-infected individuals who also abuse drugs including the psychostimulant methamphetamine (MA) develop more severe encephalitis and neuronal damage compared to HIV-1-infected patients who do not abuse drugs. In previous studies, we demonstrated that the HIV-1 protein Tat and MA interacted to cause enhanced loss of dopamine in the rat striatum via the destruction of dopaminergic terminals. Since both Tat and MA activate glia and induce cytokine production, we investigated the role of cytokines in the synergistic neurotoxicity induced by Tat and MA using cytokine arrays. Significant increases in monocyte chemotactic protein (MCP-1), interleukin-1 alpha (IL-1alpha) and tissue inhibitor of metalloproteinase-1 (TIMP-1) levels were noted 4 h following Tat + MA treatment compared to saline, Tat or MA. MCP-1 and TIMP-1 levels remained elevated 16 h after Tat + MA compared to saline or MA but were not different from the Tat-treated group at this time point. Weak, but significant elevations in cytokine-induced neutrophil chemoattractant-3 (CINC-3), ciliary neurotrophic factor (CNTF) and macrophage inflammatory protein-3 alpha (MIP-3alpha) were also noted with Tat + MA. The interaction of Tat and MA was prevented in mice genetically deficient in MCP-1 with a consequent attenuation of Tat + MA neurotoxicity. Our findings suggest that HIV-1 infection with concurrent drug abuse might profoundly increase chemokine levels in the striatum resulting in enhanced damage to the dopaminergic system.

  11. Inflammatory cytokines are suppressed by light-emitting diode irradiation of P. gingivalis LPS-treated human gingival fibroblasts: inflammatory cytokine changes by LED irradiation.

    PubMed

    Choi, HongRan; Lim, WonBong; Kim, InAe; Kim, JiSun; Ko, YoungJong; Kwon, Hyukil; Kim, SangWoo; Kabir, K M Ahsan; Li, Xiaojie; Kim, Oksu; Lee, YoungJoon; Kim, SeoYune; Kim, OkJoon

    2012-03-01

    Human gingival fibroblasts (hGFs) play an important role in the inflammatory reaction to lipopolysaccharide (LPS) from P. gingivalis, which infects periodontal connective tissue. In addition, although light-emitting diode (LED) irradiation has been reported to have biostimulatory effects, including anti-inflammatory activity, the pathological mechanisms of these effects are unclear. This study examined the effects of 635-nm irradiation of P. gingivalis LPS-treated human gingival fibroblasts on inflammatory cytokine profiles and the mitogen-activated protein kinase (MAPK) pathway, which is involved in cytokine production. Gingival fibroblasts treated or not treated with P. gingivalis LPS were irradiated with 635-nm LED light, and cytokine profiles in the supernatant were assessed using a human inflammation antibody array. Expression of cyclooxyginase-2 (COX-2) protein and phosphorylation of extracellular signal-regulated kinase (ERK 1/2), p38, and c-Jun-N-terminal kinase (JNK) were assessed by Western-blot analysis to determine the effects on the MAPK pathway, and prostaglandin E(2) (PGE(2)) in the supernatant was measured using an enzyme-linked immunoassay. COX-2 protein expression and PGE(2) production were significantly increased in the LPS-treated group and decreased by LED irradiation. LPS treatment of gingival fibroblasts led to the increased release of the pro-inflammatory-related cytokines interleukin-6 (IL-6) and IL-8, whereas LED irradiation inhibited their release. Analysis of MAPK signal transduction revealed a considerable decrease in p38 phosphorylation in response to 635-nm radiation either in the presence or absence of LPS. In addition, 635-nm LED irradiation significantly promoted JNK phosphorylation in the presence of LPS. LED irradiation can inhibit activation of pro-inflammatory cytokines, mediate the MAPK signaling pathway, and may be clinically useful as an anti-inflammatory tool.

  12. Mitogen-activated protein kinases and phosphatidylinositol 3-kinase are involved in Prevotella intermedia-induced proinflammatory cytokines expression in human periodontal ligament cells.

    PubMed

    Guan, Su-Min; Zhang, Ming; He, Jian-Jun; Wu, Jun-Zheng

    2009-08-28

    Chronic periodontitis is an inflammatory disease affecting periodontal connective tissues and alveolar bone. Proinflammatory mediators induced by periodontal pathogens play vital roles in the initiation and progression of the disease. In this study, we examined whether Prevotella intermedia induces proinflammatory cytokines expression in human periodontal ligament cells (hPDLs). The mRNA expression and protein production were determined by reverse transcription-polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbant assay (ELISA) respectively. P. intermedia treatment dose- and time-dependently increased IL-6, IL-8 and M-CSF, but not IL-1beta and TNF-alpha mRNA expression and protein secretion. Preincubation of hPDLs with extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), p38 kinase and phosphatidylinositol 3-kinase (PI3K) inhibitors PD98059, SP600125, SB203580 and LY294002 resulted in significant reduction in P. intermedia-induced IL-6, IL-8 and M-CSF expression. Blocking the synthesis of prostaglandin E(2) (PGE(2)) by indomethacin also abolished the stimulatory effects of P. intermedia on cytokines expression. Our results indicate that P. intermedia induces proinflammatory cytokines through MAPKs and PI3K signaling pathways, and PGE(2) is involved in the P. intermedia-induced proinflammatory cytokines upregulation.

  13. 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

  14. Caries induced cytokine network in the odontoblast layer of human teeth

    PubMed Central

    2011-01-01

    Background Immunologic responses of the tooth to caries begin with odontoblasts recognizing carious bacteria. Inflammatory propagation eventually leads to tooth pulp necrosis and danger to health. The present study aims to determine cytokine gene expression profiles generated within human teeth in response to dental caries in vivo and to build a mechanistic model of these responses and the downstream signaling network. Results We demonstrate profound differential up-regulation of inflammatory genes in the odontoblast layer (ODL) in human teeth with caries in vivo, while the pulp remains largely unchanged. Interleukins, chemokines, and all tested receptors thereof were differentially up-regulated in ODL of carious teeth, well over one hundred-fold for 35 of 84 genes. By interrogating reconstructed protein interaction networks corresponding to the differentially up-regulated genes, we develop the hypothesis that pro-inflammatory cytokines highly expressed in ODL of carious teeth, IL-1β, IL-1α, and TNF-α, carry the converged inflammatory signal. We show that IL1β amplifies antimicrobial peptide production in odontoblasts in vitro 100-fold more than lipopolysaccharide, in a manner matching subsequent in vivo measurements. Conclusions Our data suggest that ODL amplifies bacterial signals dramatically by self-feedback cytokine-chemokine signal-receptor cycling, and signal convergence through IL1R1 and possibly others, to increase defensive capacity including antimicrobial peptide production to protect the tooth and contain the battle against carious bacteria within the dentin. PMID:21261944

  15. Rebamipide suppresses PolyI:C-stimulated cytokine production in human conjunctival epithelial cells.

    PubMed

    Ueta, Mayumi; Sotozono, Chie; Yokoi, Norihiko; Kinoshita, Shigeru

    2013-09-01

    We previously documented that ocular surface epithelial cells could regulate ocular surface inflammation and suggested that, while Toll-like receptor 3 upregulates, EP3, one of the prostaglandin E2 receptors, downregulates ocular surface inflammation. Others reported that rebamipide, a gastroprotective drug, could not only increase the gastric mucus production, but also suppressed gastric mucosal inflammation and that it was dominantly distributed in mucosal tissues. The eyedrop form of rebamipide, approved in Japan for use in the treatment of dry eye diseases, upregulates mucin secretion and production, thereby suppressing superficial punctate keratopathy on the ocular surface of patients with this disease. In the current study, we investigated whether rebamipide has anti- inflammatory effects on the ocular surface. To examine the effects of rebamipide on polyI:C-induced cytokine expression by primary human conjunctival epithelial cells, we used enzyme-linked immunosorbent assay and quantitative reverse transcription-polymerase chain reaction assay. We studied the effects of rebamipide on ocular surface inflammation in our murine experimental allergic conjunctivitis (EAC) model. Rebamipide could suppress polyI:C-induced cytokine production and the expression of mRNAs for CXCL10, CXCL11, RANTES, MCP-1, and IL-6 in human conjunctival epithelial cells. In our EAC model, the topical administration of rebamipide suppressed conjunctival allergic eosinophil infiltration. The topical application of rebamipide on the ocular surface might suppress ocular surface inflammation by suppressing the production of cytokines by ocular surface epithelial cells.

  16. Human dendritic cell maturation and cytokine secretion upon stimulation with Bordetella pertussis filamentous haemagglutinin.

    PubMed

    Dirix, Violette; Mielcarek, Nathalie; Debrie, Anne-Sophie; Willery, Eve; Alonso, Sylvie; Versheure, Virginie; Mascart, Françoise; Locht, Camille

    2014-07-01

    In addition to antibodies, Th1-type T cell responses are also important for long-lasting protection against pertussis. However, upon immunization with the current acellular vaccines, many children fail to induce Th1-type responses, potentially due to immunomodulatory effects of some vaccine antigens, such as filamentous haemagglutinin (FHA). We therefore analysed the ability of FHA to modulate immune functions of human monocyte-derived dendritic cells (MDDC). FHA was purified from pertussis toxin (PTX)-deficient or from PTX- and adenylate cyclase-deficient Bordetella pertussis strains, and residual endotoxin was neutralized with polymyxin B. FHA from both strains induced phenotypic maturation of human MDDC and cytokine secretion (IL-10, IL-12p40, IL-12p70, IL-23 and IL-6). To identify the FHA domains responsible for MDDC immunomodulation, MDDC were stimulated with FHA containing a Gly→Ala substitution at its RGD site (FHA-RAD) or with an 80-kDa N-terminal moiety of FHA (Fha44), containing its heparin-binding site. Whereas FHA-RAD induced maturation and cytokine production comparable to those of FHA, Fha44 did not induce IL-10 production, but maturated MDDC at least partially. Nevertheless, Fha44 induced the secretion of IL-12p40, IL-12p70, IL-23 and IL-6 by MDDC, albeit at lower levels than FHA. Thus, FHA can modulate MDDC responses in multiple ways, and IL-10 induction can be dissociated from the induction of other cytokines.

  17. 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

  18. Increased hydrophobicity in Malassezia species correlates with increased proinflammatory cytokine expression in human keratinocytes.

    PubMed

    Akaza, Narifumi; Akamatsu, Hirohiko; Takeoka, Shiori; Mizutani, Hiroshi; Nakata, Satoru; Matsunaga, Kayoko

    2012-11-01

    Malassezia cells stimulate cytokine production by keratinocytes, although this ability differs among Malassezia species for unknown reasons. The aim of this study was to clarify the factors determining the ability to induce cytokine production by human keratinocytes in response to Malassezia species. M. furfur NBRC 0656, M. sympodialis CBS 7222, M. dermatis JCM 11348, M. globosa CBS 7966, M. restricta CBS 7877, and three strains each of M. globosa, M. restricta, M. dermatis, M. sympodialis, and M. furfur maintained under various culture conditions were used. Normal human epidermal keratinocytes (NHEKs) (1 × 10(5) cells) and the Malassezia species (1 × 10(6) cells) were co-cultured, and IL-1α, IL-6, and IL-8 mRNA levels were determined. Moreover, the hydrophobicity and β-1,3-glucan expression at the surface of Malassezia cells were analyzed. The ability of Malassezia cells to trigger the mRNA expression of proinflammatory cytokines in NHEKs differed with the species and conditions and was dependent upon the hydrophobicity of Malassezia cells not β-1,3-glucan expression.

  19. Human TNF cytokine neutralization with a vNAR from Heterodontus francisci shark

    PubMed Central

    Camacho-Villegas, Tanya; Mata-Gonzalez, Teresa; Paniagua-Solis, Jorge; Sanchez, Edna; Licea, Alexei

    2013-01-01

    The therapeutic use of single domain antibodies (sdAbs) is a promising new approach because these small antibodies maintain antigen recognition and neutralization capacity, have thermal and chemical stability and have good solubility. In this study, using phage display technology, we isolated a variable domain of a IgNAR (vNAR) from a Heterodontus francisci shark immunized against the recombinant human cytokine TNFα (rhTNFα). One clone T43, which expresses the vNAR protein in the periplasmic space, was isolated from the fourth round of panning. T43 had the capacity to recognize rhTNF and neutralize it in vitro, indicating that T43 has potential as a therapeutic that can be used for diseases in which this pro-inflammatory cytokine needs to be controlled. PMID:23221782

  20. Echinacea purpurea (L.) Moench modulates human T-cell cytokine response.

    PubMed

    Fonseca, Fabiana N; Papanicolaou, Genovefa; Lin, Hong; Lau, Clara B S; Kennelly, Edward J; Cassileth, Barrie R; Cunningham-Rundles, Susanna

    2014-03-01

    The study objective was to evaluate the composition of a neutral and weakly acidic water-soluble extract from Echinacea purpurea (L.) Moench (EchNWA) previously shown to modify murine influenza infection, and to assess immunomodulatory effects on human T-cells. EchNWA extract from fresh aerial parts was extracted with water, ethanolic precipitation, and size-exclusion chromatography. The chemical profile of EchNWA was characterized by chromatography (size-exclusion, HPLC, GC-MS), and small molecule fingerprint analysis performed by HPLC-PDA. Jurkat T-cells at high and low cell density were pretreated or not with doses of EchNWA, followed by activation with phorbol 12-myristate 13-acetate plus ionomycin (PMA+I). Interleukin-2 (IL-2) and interferon gamma (IFNg) cytokine secretions were measured by multi-cytokine luminex technology. Results showed that EchNWA contains 80% polysaccharides, predominantly a 10kDa entity; phenolic compounds, cynarin, cichoric and caftaric acids, but no detectable alkylamides. Cytokine production required stimulation and was lower after PMA+I activation in high-density compared to low-density conditions. EchNWA mediated a strong dose-dependent enhancement of high-density T-cell production of IL-2 and IFNg response to PMA+I. EchNWA alone did not stimulate T-cells. EchNWA enhanced mean fluorescence intensity of IL-2 in Jurkat T-cells activated by PMA+1 or ionomycin alone. Conversely EchNWA mediated modest but significant suppression of IFNg response and reduced the percentage of CD25+ T-cells under low-density conditions. Conclusions are that EchNWA polysaccharides, but not phenolic compounds have dose-related adjuvant effects on human T-cell cytokine responses characterized by enhancing and suppressive effects that are regulated by T-cell density.

  1. Regulation of the syncytin-1 promoter in human astrocytes by multiple sclerosis-related cytokines

    SciTech Connect

    Mameli, Giuseppe . E-mail: viross@uniss.it; Astone, Vito; Khalili, Kamel; Serra, Caterina; Sawaya, Bassel E.; Dolei, Antonina

    2007-05-25

    Syncytin-1 has a physiological role during early pregnancy, as mediator of trophoblast fusion into the syncytiotrophoblast layer, hence allowing embryo implantation. In addition, its expression in nerve tissue has been proposed to contribute to the pathogenesis of multiple sclerosis (MS). Syncytin-1 is the env glycoprotein of the ERVWE1 component of the W family of human endogenous retroviruses (HERV), located on chromosome 7q21-22, in a candidate region for genetic susceptibility to MS. The mechanisms of ERVWE1 regulation in nerve tissue remain to be identified. Since there are correlations between some cytokines and MS outcome, we examined the regulation of the syncytin-1 promoter by MS-related cytokines in human U-87MG astrocytic cells. Using transient transfection assays, we observed that the MS-detrimental cytokines TNF{alpha}, interferon-{gamma}, interleukin-6, and interleukin-1 activate the ERVWE1 promoter, while the MS-protective interferon-{beta} is inhibitory. The effects of cytokines are reduced by the deletion of the cellular enhancer domain of the promoter that contains binding sites for several transcription factors. In particular, we found that TNF{alpha} had the ability to activate the ERVWE1 promoter through an NF-{kappa}B-responsive element located within the enhancer domain of the promoter. Electrophoretic mobility shift and ChIP assays showed that TNF{alpha} enhances the binding of the p65 subunit of NF-{kappa}B, to its cognate site within the promoter. The effect of TNF{alpha} is abolished by siRNA directed against p65. Taken together, these results illustrate a role for p65 in regulating the ERVWE1 promoter and in TNF{alpha}-mediated induction of syncytin-1 in multiple sclerosis.

  2. [Cytokine profile in human hydatidosis: possible role in the immunosurveillance of patients infected with Echinococcus granulosus].

    PubMed

    Mezioug, D; Touil-Boukoffa, C

    2009-03-01

    Hydatidosis is a severe parasitic disease caused by infection with metacestode of the tapeworms Echinococcus granulosus. In human, the larval forms develop into large cysts especially in the liver, lung and brain. Our aim in this study was to investigate Th1 and Th2 cytokines production in hydatid disease in order to evaluate implication of Th1/Th2 ratio in the evolution of pathology according to the cystic localization, clinical stage and clinical evolution. Interferon-gamma (IFN-gamma), interleukine-12 (Il-12), interleukine-16 (Il-16), interleukine-18 (Il-18), interleukine-4 (Il-4), interleukine-5 (Il-5), interleukine-10 (Il-10) and interleukine-13 (Il-13) production is determined in sera from hydatid Algerian patients (n = 177) with liver, lung, liver and lung associated, spleen, kidney, osseo, heart and multiples hydatid cyst and in sera from patients with clinical complications (calcified liver cysts; infected lung cysts; vomique lung cysts and patients who relapsed) and according clinical stage (before surgical extirpation of the cyst and after surgical extirpation of the cyst). Cytokines are evaluated by enzyme-linked immunosorbent Kits (ELISA Immunotech). The coexistence of elevated activities of IFN-gamma, Il-12, Il-16, Il-18, Il-4, Il-5, Il-10, and Il-13 is observed in most of sera from hydatid patients. In contrast, healthy controls showed minor levels. These results support Th1 and Th2 cell subsets activation in human hydatidosis. The comparison of Th1/Th2 production shows that the induction of these mediators is related to the cystic localization, the clinical stage and clinical evolution. Collectively, our data indicates that Th1 cytokines are related to the protective immunity, in contrast Th2 cytokines are responsible to the susceptibility to disease and associated with chronicle stage, clinical complications and secondary locations.

  3. Echinacea purpurea (L.) Moench modulates human T-cell cytokine response☆

    PubMed Central

    Fonseca, Fabiana N.; Papanicolaou, Genovefa; Lin, Hong; Lau, Clara B.S.; Kennelly, Edward J.; Cassileth, Barrie R.; Cunningham-Rundles, Susanna

    2014-01-01

    The study objective was to evaluate the composition of a neutral and weakly acidic water-soluble extract from Echinacea purpurea (L.) Moench (EchNWA) previously shown to modify murine influenza infection, and to assess immunomodulatory effects on human T-cells. EchNWA extract from fresh aerial parts was extracted with water, ethanolic precipitation, and size-exclusion chromatography. The chemical profile of EchNWA was characterized by chromatography (size-exclusion, HPLC, GC–MS), and small molecule finger-print analysis performed by HPLC–PDA. Jurkat T-cells at high and low cell density were pretreated or not with doses of EchNWA, followed by activation with phorbol 12-myristate 13-acetate plus ionomycin (PMA+I). Interleukin-2 (IL-2) and interferon gamma (IFNg) cytokine secretions were measured by multi-cytokine luminex technology. Results showed that EchNWA contains 80% polysaccharides, predominantly a 10 kDa entity; phenolic compounds, cynarin, cichoric and caftaric acids, but no detectable alkylamides. Cytokine production required stimulation and was lower after PMA+I activation in high-density compared to low-density conditions. EchNWA mediated a strong dose-dependent enhancement of high-density T-cell production of IL-2 and IFNg response to PMA+I. EchNWA alone did not stimulate T-cells. EchNWA enhanced mean fluorescence intensity of IL-2 in Jurkat T-cells activated by PMA+1 or ionomycin alone. Conversely EchNWA mediated modest but significant suppression of IFNg response and reduced the percentage of CD25+ T-cells under low-density conditions. Conclusions are that EchNWA polysaccharides, but not phenolic compounds have dose-related adjuvant effects on human T-cell cytokine responses characterized by enhancing and suppressive effects that are regulated by T-cell density. PMID:24434371

  4. Transcription and translation of human F11R gene are required for an initial step of atherogenesis induced by inflammatory cytokines

    PubMed Central

    2011-01-01

    Background - The F11 Receptor (F11R; aka JAM-A, JAM-1) is a cell adhesion protein present constitutively on the membrane surface of circulating platelets and within tight junctions of endothelial cells (ECs). Previous reports demonstrated that exposure of ECs to pro-inflammatory cytokines causes insertion of F11R molecules into the luminal surface of ECs, ensuing with homologous interactions between F11R molecules of platelets and ECs, and a resultant adhesion of platelets to the inflamed ECs. The main new finding of the present report is that the first step in this chain of events is the de-novo transcription and translation of F11R molecules, induced in ECs by exposure to inflammatory cytokines. Methods - The experimental approach utilized isolated, washed human platelet suspensions and cultured human venous endothelial cells (HUVEC) and human arterial endothelial cells (HAEC) exposed to the proinflammatory cytokines TNF-alpha and/or IFN-gamma, for examination of the ability of human platelets to adhere to the inflamed ECs thru the F11R. Our strategy was based on testing the effects of the following inhibitors on this activity: general mRNA synthesis inhibitors, inhibitors of the NF-kappaB and JAK/STAT pathways, and small interfering F11R-mRNA (siRNAs) to specifically silence the F11R gene. Results - Treatment of inflamed ECs with the inhibitors actinomycin, parthenolide or with AG-480 resulted in complete blockade of F11R- mRNA expression, indicating the involvement of NF-kappaB and JAK/STAT pathways in this induction. Transfection of ECs with F11R siRNAs caused complete inhibition of the cytokine-induced upregulation of F11R mRNA and inhibition of detection of the newly- translated F11R molecules in cytokine-inflamed ECs. The functional consequence of the inhibition of F11R transcription and translation was the significant blockade of the adhesion of human platelets to inflamed ECs. Conclusion - These results prove that de novo synthesis of F11R in ECs is

  5. Immunomodulatory action of Copaifera spp oleoresins on cytokine production by human monocytes.

    PubMed

    Santiago, Karina Basso; Conti, Bruno José; Murbach Teles Andrade, Bruna Fernanda; Mangabeira da Silva, Jonas Joaquim; Rogez, Hervé Louis Ghislain; Crevelin, Eduardo José; Beraldo de Moraes, Luiz Alberto; Veneziani, Rodrigo; Ambrósio, Sérgio Ricardo; Bastos, Jairo Kenupp; Sforcin, José Maurício

    2015-03-01

    Copaifera spp oleoresins have been used in folk medicine for centuries; nevertheless, its immunomodulatory action has not been investigated. Thus, the goal of this study was to characterize different oleoresins and to verify their action on human monocytes regarding pro- and anti-inflammatory cytokine production (TNF-α and IL-10, respectively). The chemical composition of Brazilian Copaifera reticulata, Copaifera duckey and Copaifera multijuga oleoresins was analyzed by HPLC-MS. Cell viability was assessed by MTT method after incubation of cells with Copaifera spp. Noncytotoxic concentrations of oleoresins were incubated with human monocytes from healthy donors, and cytokine production was determined by ELISA. HPLC-MS analysis for terpenes allowed the identification of six diterpene acids and one sesquiterpene acid. Oleoresins exerted no cytotoxic effects on human monocytes. All oleoresins had a similar profile: LPS-induced TNF-α production was maintained by oleoresins, while a significant inhibitory action on IL-10 production was seen. Copaifera oleoresins seemed to exert an activator profile on human monocytes without affecting cell viability. Such effect may be due to the presence of either diterpene or sesquiterpene acids; however, further studies are necessary to determine the involvement of such compounds in Copaifera immunomodulatory effects. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

  6. Glucose-dependent insulinotropic polypeptide induces cytokine expression, lipolysis, and insulin resistance in human adipocytes.

    PubMed

    Timper, Katharina; Grisouard, Jean; Sauter, Nadine S; Herzog-Radimerski, Tanja; Dembinski, Kaethi; Peterli, Ralph; Frey, Daniel M; Zulewski, Henryk; Keller, Ulrich; Müller, Beat; Christ-Crain, Mirjam

    2013-01-01

    Obesity-related insulin resistance is linked to a chronic state of systemic and adipose tissue-derived inflammation. Glucose-dependent insulinotropic polypeptide (GIP) is an incretin hormone also acting on adipocytes. We investigated whether GIP affects inflammation, lipolysis, and insulin resistance in human adipocytes. Human subcutaneous preadipocyte-derived adipocytes, differentiated in vitro, were treated with human GIP to analyze mRNA expression and protein secretion of cytokines, glycerol, and free fatty acid release and insulin-induced glucose uptake. GIP induced mRNA expression of IL-6, IL-1β, and the IL-1 receptor antagonist IL-1Ra, whereas TNFα, IL-8, and monocyte chemotactic protein (MCP)-1 remained unchanged. Cytokine induction involved PKA and the NF-κB pathway as well as an autocrine IL-1 effect. Furthermore, GIP potentiated IL-6 and IL-1Ra secretion in the presence of LPS, IL-1β, and TNFα. GIP induced lipolysis via activation of hormone-sensitive lipase and was linked to NF-κB activation. Finally, chronic GIP treatment impaired insulin-induced glucose uptake possibly due to the observed impaired translocation of glucose transporter GLUT4. In conclusion, GIP induces an inflammatory and prolipolytic response via the PKA -NF-κB-IL-1 pathway and impairs insulin sensitivity of glucose uptake in human adipocytes.

  7. Erythropoietin stimulation of human adipose tissue for therapeutic refilling releases protective cytokines

    PubMed Central

    Sabbatini, Maurizio; Bosetti, Michela; Borrone, Alessia; Moalem, Liah; Taveggia, Antonio; Verna, Giovanni; Cannas, Mario

    2016-01-01

    Apoptosis and inflammatory processes may be at the basis of reducing graft survival. Erythropoietin is a tissue-protective hormone with pleiotropic potential, and it interferes with the activities of pro-inflammatory cytokines and stimulates healing following injury, preventing destruction of tissue surrounding the injury site. It may represent a useful tool to increase the autograft integration. Through the use of multipanel kit cytokine analysis we have detected the cytokines secreted by human tissue adipose mass seeded in culture following withdrawal by Coleman’s modified technique in three groups: control, after lipopolysaccharides stimulation and after erythropoietin stimulation. In the control group, we have observed expression of factors that may have a role in protecting the tissue homeostatic mechanism. But the same factors were secreted following stimulation with lipopolysaccharides combined with others factors that delineated the inflammatory state. Instead through erythropoietin stimulation, the factors known to exert tissue-protective action were secreted. Therefore, the use of a trophic factors such as erythropoietin may help to inhibit the potential inflammatory process development and stimulate the activation of reparative/regenerative process in the tissue graft. PMID:27738510

  8. T-lymphocyte and cytokine expression in human inflammatory periapical lesions.

    PubMed

    de Brito, Luciana Carla Neves; Teles, Flávia Rocha Fonseca; Teles, Ricardo Palmier; Totola, Antônio Helvécio; Vieira, Leda Quércia; Sobrinho, Antônio Paulino Ribeiro

    2012-04-01

    Lymphocytes, among many cells, express different sets of cytokines, chemokines, and receptors, which are considered important mediators of periapical immune response to infection. The aim of this study was to evaluate the mRNA expression of CD4(+)CD28(+) and CD8(+) T genes and the gene expression of interferon-γ, tumor necrosis factor-α, interleukin (IL)-1β, IL-17A, IL-10, CCL2/MCP-1, CCL4, CCL5, CXCR4, CCR5, and receptor activator for nuclear factor kappa B ligand (RANKL) in periapical interstitial fluid from human root canal infections. The samples were collected immediately after root canal cleaning and 7 days later (restrained root canal bacterial load) to characterize those gene expressions. Real-time polymerase chain reaction demonstrated significantly higher levels of CD4(+)CD28(+) and CD8(+) T-cell markers in the former root canal condition and an increase of IL-10 and CXCR4, followed by a decrease of proinflammatory cytokines such as RANKL, interferon-γ, IL-1β, and CCL5. Analyses of T-lymphocyte and cytokine expression in periapical area were able to show that distinct root canal conditions might play regulatory roles in controlling local immune/inflammatory processes. Copyright © 2012 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

  9. Cytokine response of human mononuclear cells induced by intestinal Clostridium species.

    PubMed

    Tuovinen, Elina; Keto, Joni; Nikkilä, Janne; Mättö, Jaana; Lähteenmäki, Kaarina

    2013-02-01

    Altered composition of intestinal microbiota has been associated with various immunological disorders such as inflammatory bowel disease. Although Clostridium species are major inhabitants of the intestinal tract, their interaction with the host immunological system is yet poorly characterized. In this study, cytokine responses of human monocytic cell line THP-1 and peripheral blood mononuclear cells (PBMC) to six type strains representing common intestinal clostridial species were determined. The strains induced diverse cytokine responses in both THP-1 cells and PBMC. Clostridium perfringens was the most potent inducer of both tumour necrosis factor alpha (TNF-alpha) and interleukin-10 (IL-10), as compared to Clostridium histolyticum, Clostridium clostridioforme, Clostridium leptum, Clostridium sporosphaeroides and Blautia coccoides. Interleukin-8 (IL-8) production in PBMC was most efficiently stimulated by C. sporosphaeroides. The same PBMC preparations that responded strongly to Escherichia coli lipopolysaccharide (LPS) also responded strongly to bacterial stimulation. This indicates that the level of responsiveness is an individual feature of mononuclear cell preparations, and that the overall cytokine response is composed by a combination of host factors and microbial structures affecting them. This work supports the idea that the composition of the intestinal clostridial population influences immune responses and is likely to play an important role in intestinal homeostasis.

  10. Modulation of the cytokine response in human monocytes by mycobacterium leprae phenolic glycolipid-1.

    PubMed

    Manca, Claudia; Peixoto, Blas; Malaga, Wladimir; Guilhot, Christophe; Kaplan, Gilla

    2012-01-01

    Leprosy is a chronic but treatable infectious disease caused by the intracellular pathogen Mycobacterium leprae. M. leprae cell wall is characterized by a unique phenolic glycolipid-1 (PGL-1) reported to have several immune functions. We have examined the role of PGL-1 in the modulation of monocyte cytokine/chemokine production in naive human monocytes. PGL-1 in its purified form or expressed in a recombinant Mycobacterium bovis Bacillus Colmette-Guérin (BCG) background (rBCG-PGL-1) was tested. We found that PGL-1 selectively modulated the induction of specific monocyte cytokines and chemokines and, when used as prestimulus, exerted priming and/or inhibitory effects on the induction of selected cytokines/chemokines in response to a second stimulus. Taken together, the results of this study support a modulatory role for PGL-1 in the innate immune response to M. leprae. Thus, PGL-1 may play an important role in the development of the anergic clinical forms of disease and in tissue damage seen in lepromatous patients and during the reactional states of leprosy.

  11. 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

  12. Estrogens interfere with leflunomide modulation of cytokine production by human activated monocytes.

    PubMed

    Montagna, Paola; Soldano, Stefano; Brizzolara, Renata; Villaggio, Barbara; Triolo, Pierfranco; Clerico, Paolo; Meroni, Marianna; Sulli, Alberto; Seriolo, Bruno; Molfetta, Luigi; Cutolo, Maurizio

    2010-04-01

    Rheumatoid arthritis (RA) prevalence is greater in females than in males, supporting estrogens as modulators of immune response. Leflunomide (LEF) is employed in the RA treatment. We studied the combinatory effects of LEF active metabolite A77 1726 (LEF-M) and 17beta-estradiol (E2) on inflammatory cytokine production by cultured macrophages obtained from activated human monocytes (THP-1 cells). Macrophages were cultured with LEF-M alone and in combination with E2. IL-6, TNF-alpha, and TGF-beta were evaluated by immunocytochemistry (ICC), Western blot (WB), and reverse transcriptase-polymerase chain reaction (RT-PCR). ICC, as well as WB and RT-PCR, showed that LEF-M, in respect to untreated cells, significantly downregulated the cytokine production (IL-6 P < 0.01, TNF-alphaP < 0.001, TGF-betaP < 0.01). On the contrary, E2 increased the cytokine production, a result that was significantly reversed when LEF-M was subsequently added (IL-6, TNF-alpha, TGF-betaP < 0.001 vs. E2). E2 seems to contrast the LEF-M activity. These results might support a more efficient therapeutical effect of LEF in male with respect to female RA patients.

  13. Significance of human cytomegalovirus and Epstein-Barr virus in inducing cytokine expression in periapical lesions.

    PubMed

    Sabeti, Mohammad; Kermani, Vali; Sabeti, Sara; Simon, James H

    2012-01-01

    Because herpesviruses might be etiologically involved in periapical pathosis of endodontic origin, this study aimed to determine the occurrence of human cytomegalovirus (HCMV), Epstein-Barr virus (EBV), and the expression of mRNA transcripts of tumor necrosis factor (TNF)-α, γ-interferon (IFN), interleukin (IL)-1β, IL-6, IL-12, and IL-10 in periapical granulomatous lesions collected in conjunction with apicoectomy. A total of 9 symptomatic and 6 asymptomatic teeth with periapical lesions were studied. Periapical samples were collected in conjunction with apicoectomy, which was being performed because of radiographic evidence of incomplete periapical healing after conventional root canal therapy. By using established polymerase chain reaction primers and procedures, polymerase chain reaction assays were used to identify herpesvirus and cytokine gene expression. The difference in occurrence of HCMV, EBV, and cytokines between symptomatic and asymptomatic periapical lesions was statistically significant: HCMV (P = .048), EBV (P = .002), IFN (P = .001), IL-1 (P = .012), IL-6 (P = .026), IL-10 (P = .026), IL-12 (P = .012), and TNF (P < .001) (Mann-Whitney U test). There was a significant correlation between EBV, HCMV, and TNF, γ-IFN, IL-1, and IL-12 in symptomatic periapical lesions (Spearman test). The present findings provide evidence of a putative role of HCMV and EBV in the pathogenesis of symptomatic periapical pathosis. The release of tissue-destructive cytokines might be of pathogenetic significance. Copyright © 2012 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

  14. Maraviroc reduces cytokine expression and secretion in human adipose cells without altering adipogenic differentiation.

    PubMed

    Díaz-Delfín, Julieta; Domingo, Pere; Giralt, Marta; Villarroya, Francesc

    2013-03-01

    Maraviroc (MVC) is a drug approved for use as part of HAART in treatment-experienced HIV-1 patients with CCR5-tropic virus. Despite the current concerns on the alterations in adipose tissue that frequently appear in HIV-infected patients under HAART, there is no information available on the effects of MVC on adipose tissue. Here we studied the effects of MVC during and after the differentiation of human adipocytes in culture, and compared the results with the effects of efavirenz (EFV). We measured the acquisition of adipocyte morphology; the gene expression levels of markers for mitochondrial toxicity, adipogenesis and inflammation; and the release of adipokines and cytokines to the medium. Additionally, we determined the effects of MVC on lipopolysaccharide (LPS)-induced pro-inflammatory cytokine expression in adipocytes. Unlike EFV-treated pre-adipocytes, MVC-treated pre-adipocytes showed no alterations in the capacity to differentiate into adipocytes and accumulated lipids normally. Consistent with this, there were no changes in the mRNA levels of PPARγ or SREBP-1c, two master regulators of adipogenesis. In addition, MVC caused a significant decrease in the gene expression and release of pro-inflammatory cytokines, whereas EFV had the opposite effect. Moreover, MVC lowered inflammation-related gene expression and inhibited the LPS-induced expression of pro-inflammatory genes in differentiated adipocytes. We conclude that MVC does not alter adipocyte differentiation but rather shows anti-inflammatory properties by inhibiting the expression and secretion of pro-inflammatory cytokines. Collectively, our results suggest that MVC may minimize adverse effects on adipose tissue development, metabolism, and inflammation, and thus could be a potentially beneficial component of antiretroviral therapy.

  15. 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

  16. Elevated serum cytokines during human anaphylaxis: Identification of potential mediators of acute allergic reactions.

    PubMed

    Stone, Shelley F; Cotterell, Claire; Isbister, Geoffrey K; Holdgate, Anna; Brown, Simon G A

    2009-10-01

    Anaphylaxis is generally unanticipated and requires emergency management. Therefore, the biological mediators in human beings have been difficult to define. Our aim was to identify cytokines and chemokines whose concentrations increase during anaphylaxis in human beings and to determine how each correlates with severity. We measured the concentrations of potential mediators, including cytokines, chemokines, mast cell tryptase (MCT), and histamine, over 3 time points in 76 patients presenting to emergency departments with anaphylaxis and correlated these with a global severity scale, hypotension, and hypoxia. IL-2, IL-6, IL-10, TNF receptor I, MCT, and histamine were significantly elevated in patients with severe reactions (n = 36) compared with moderate reactions (n = 40) and healthy controls. Histamine levels peaked at emergency department arrival, whereas other mediators peaked later. IL-4, IL-5, IL-13, IFN-gamma, and TNF-alpha were marginally elevated in severe reactions compared with healthy controls but did not correlate with reaction severity. Severe reactions tended to be either hypotensive (n = 19) or hypoxemic (n = 12). Levels of IL-6, IL-10, TNF receptor I, MCT, and histamine correlated with hypotension. No mediator correlated with hypoxemia or other respiratory features. This study confirms that the concentrations of a number of cytokines are elevated in blood during anaphylaxis in human beings and that some correlate with the presence of hypotension. Others were only marginally elevated within a concentration range that available assays do not reliably detect. During respiratory reactions, mediators may be largely confined to the airways so that blood concentrations do not reflect activity.

  17. Cytokine profile in human eyes: contribution of a new cytokine combination for differential diagnosis between intraocular lymphoma or uveitis.

    PubMed

    Fisson, Sylvain; Ouakrim, Hanane; Touitou, Valérie; Baudet, Sylvie; Ben Abdelwahed, Rym; Donnou, Sabrina; Miloudi, Amine; Galand, Claire; Bodaghi, Bahram; Lehoang, Phuc; Brissard, Martine; Le Garff-Tavernier, Magali; Fridman, Wolf Herman; Sautès-Fridman, Catherine; Cassoux, Nathalie; Merle-Béral, Hélène

    2013-01-01

    Primary intraocular lymphoma (PIOL), also called primary vitreoretinal lymphomas, often masquerades as uveitis. This misdiagnosis can result in subsequent brain involvement and oculocerebral lymphoma (OCL). In this study, we sought to characterize the helper T-cell type 1 (Th1)/Th2 cytokine profile in vitreous samples from patients with PIOL, OCL, uveitis and controls with non-inflammatory disease. Vitreous and aqueous humor samples from 87 patients with PIOL (n = 30), OCL (n = 12), uveitis (n = 34), and retinal detachment (RD) without hemorrhage (n = 11) were analyzed and their concentrations of interleukin (IL)-2, IL-4, IL-6, IL-10, interferon (IFN)-γ, and tumor necrosis factor (TNF)-α were determined by flow cytometric bead arrays (CBA). The IL-10 levels determined by CBA were compared with those by ELISA. IL-10 concentrations measured by CBA and ELISA were highly correlated. IL-2, IL-4, and TNFα were not detected in any sample. The only cytokine detected at a significant level in samples from RD vitreous was IL-6. The IL-10/IL-6 ratio, as previously reported, was slightly higher in PIOL than in uveitis samples, but not for all patients. Cytokine profiles from PIOL and OCL samples did not differ. The combination of the IL-10/IL-6 and IL-10/IFNγ ratios was highly informative for discriminating PIOL/OCL from uveitis samples and for therapeutic follow up of PIOL. This strategy might be very helpful as an initial screening to rule out PIOL in patients thought to have uveitis.

  18. Induction of proinflammatory cytokines in human lung epithelial cells during Rhodococcus equi infection.

    PubMed

    Remuzgo-Martínez, Sara; Pilares-Ortega, Lilian; Alvarez-Rodríguez, Lorena; Aranzamendi-Zaldunbide, Maitane; Padilla, Daniel; Icardo, Jose Manuel; Ramos-Vivas, Jose

    2013-08-01

    Rhodococcus equi is an opportunistic human pathogen associated with immunosuppressed people. While the interaction of R. equi with macrophages has been comprehensively studied, little is known about its interactions with non-phagocytic cells. Here, we characterized the entry process of this bacterium into human lung epithelial cells. The invasion is inhibited by nocodazole and wortmannin, suggesting that the phosphatidylinositol 3-kinase pathway and microtubule cytoskeleton are important for invasion. Pre-incubation of R. equi with a rabbit anti-R. equi polyclonal antiserum resulted in a dramatic reduction in invasion. Also, the invasion process as studied by immunofluorescence and scanning electron microscopy indicates that R. equi make initial contact with the microvilli of the A549 cells, and at the structural level, the entry process was observed to occur via a zipper-like mechanism. Infected lung epithelial cells upregulate the expression of cytokines IL-8 and IL-6 upon infection. The production of these pro-inflammatory cytokines was significantly enhanced in culture supernatants from cells infected with non-mucoid plasmid-less strains when compared with cells infected with mucoid strains. These results demonstrate that human airway epithelial cells produce pro-inflammatory mediators against R. equi isolates.

  19. Distinct differences in the responses of the human pancreatic β-cell line EndoC-βH1 and human islets to proinflammatory cytokines

    PubMed Central

    Oleson, Bryndon J.; McGraw, Jennifer A.; Broniowska, Katarzyna A.; Annamalai, Mani; Chen, Jing; Bushkofsky, Justin R.; Davis, Dawn B.; Mathews, Clayton E.

    2015-01-01

    While insulinoma cells have been developed and proven to be extremely useful in studies focused on mechanisms controlling β-cell function and viability, translating findings to human β-cells has proven difficult because of the limited access to human islets and the absence of suitable insulinoma cell lines of human origin. Recently, a human β-cell line, EndoC-βH1, has been derived from human fetal pancreatic buds. The purpose of this study was to determine whether human EndoC-βH1 cells respond to cytokines in a fashion comparable to human islets. Unlike most rodent-derived insulinoma cell lines that respond to cytokines in a manner consistent with rodent islets, EndoC-βH1 cells fail to respond to a combination of cytokines (IL-1, IFN-γ, and TNF) in a manner consistent with human islets. Nitric oxide, produced following inducible nitric oxide synthase (iNOS) expression, is a major mediator of cytokine-induced human islet cell damage. We show that EndoC-βH1 cells fail to express iNOS or produce nitric oxide in response to this combination of cytokines. Inhibitors of iNOS prevent cytokine-induced loss of human islet cell viability; however, they do not prevent cytokine-induced EndoC-βH1 cell death. Stressed human islets or human islets expressing heat shock protein 70 (HSP70) are resistant to cytokines, and, much like stressed human islets, EndoC-βH1 cells express HSP70 under basal conditions. Elevated basal expression of HSP70 in EndoC-βH1 cells is consistent with the lack of iNOS expression in response to cytokine treatment. While expressing HSP70, EndoC-βH1 cells fail to respond to endoplasmic reticulum stress activators, such as thapsigargin. These findings indicate that EndoC-βH1 cells do not faithfully recapitulate the response of human islets to cytokines. Therefore, caution should be exercised when making conclusions regarding the actions of cytokines on human islets when using this human-derived insulinoma cell line. PMID:26084699

  20. Distinct differences in the responses of the human pancreatic β-cell line EndoC-βH1 and human islets to proinflammatory cytokines.

    PubMed

    Oleson, Bryndon J; McGraw, Jennifer A; Broniowska, Katarzyna A; Annamalai, Mani; Chen, Jing; Bushkofsky, Justin R; Davis, Dawn B; Corbett, John A; Mathews, Clayton E

    2015-09-01

    While insulinoma cells have been developed and proven to be extremely useful in studies focused on mechanisms controlling β-cell function and viability, translating findings to human β-cells has proven difficult because of the limited access to human islets and the absence of suitable insulinoma cell lines of human origin. Recently, a human β-cell line, EndoC-βH1, has been derived from human fetal pancreatic buds. The purpose of this study was to determine whether human EndoC-βH1 cells respond to cytokines in a fashion comparable to human islets. Unlike most rodent-derived insulinoma cell lines that respond to cytokines in a manner consistent with rodent islets, EndoC-βH1 cells fail to respond to a combination of cytokines (IL-1, IFN-γ, and TNF) in a manner consistent with human islets. Nitric oxide, produced following inducible nitric oxide synthase (iNOS) expression, is a major mediator of cytokine-induced human islet cell damage. We show that EndoC-βH1 cells fail to express iNOS or produce nitric oxide in response to this combination of cytokines. Inhibitors of iNOS prevent cytokine-induced loss of human islet cell viability; however, they do not prevent cytokine-induced EndoC-βH1 cell death. Stressed human islets or human islets expressing heat shock protein 70 (HSP70) are resistant to cytokines, and, much like stressed human islets, EndoC-βH1 cells express HSP70 under basal conditions. Elevated basal expression of HSP70 in EndoC-βH1 cells is consistent with the lack of iNOS expression in response to cytokine treatment. While expressing HSP70, EndoC-βH1 cells fail to respond to endoplasmic reticulum stress activators, such as thapsigargin. These findings indicate that EndoC-βH1 cells do not faithfully recapitulate the response of human islets to cytokines. Therefore, caution should be exercised when making conclusions regarding the actions of cytokines on human islets when using this human-derived insulinoma cell line.

  1. Chemoprotective effect of monoisoamyl 2, 3-dimercaptosuccinate (MiADMS) on cytokines expression in cadmium chloride treated human lung cells.

    PubMed

    Odewumi, Caroline O; Fils-Aime, Shiela; Badisa, Veera L D; Latinwo, Lekan M; Ruden, Michael L; Ikediobi, Christopher; Badisa, Ramesh B

    2015-01-01

    Cadmium is commercially profitable element, but it causes toxicity in humans and animals leading to diseases in various organs. The main route of cadmium exposure to humans is through inhalation. Lungs respond to insult through secretion of cytokines. In this study, the chemoprotective effect of monoisoamyl 2, 3-dimercaptosuccinate (MiADMS) was evaluated on viability and cytokines expression in CdCl2 treated human lung A549 cells by cytokine array. Cells were treated with 0, 50, 75, and 100 µM CdCl2 alone, 300 µM MiADMS alone, and co-treated with 300 µM MiADMS and 75 µM CdCl2 for 24 h. The viability was measured by crystal violet dye. The level of cytokines in the cells' lysate and cell culture medium was measured using Ray Biotech's Human Cytokine Array 6 in control cells, 75 µM CdCl2 alone and MiADMS co-treated cells. Array results were validated by ELISA kit. The CdCl2 caused a dose dependent decrease in cell viability, while MiADMS co-treatment resulted in a significant increase in viability of CdCl2 treated cells. Morphology of the cells treated with CdCl2 was destroyed, while MiADMS restored the lost shape in CdCl2 treated cells. In addition, the cells co-treated with MiADMS and CdCl2 showed modulation of cytokines expression in comparison to the CdCl2 alone treated cells. The ELISA results showed the similar pattern of cytokine expression as Human Cytokine Array and validated the array results. These results clearly show the chemoprotective effect of MiADMS and suggest that MiADMS can be used as antidote at moderate dose against CdCl2 toxicity. © 2014 Wiley Periodicals, Inc.

  2. Longitudinal Study of Cytokine Expression, Lipid Profile and Neuronal Growth Factors in Human Breast Milk from Term and Preterm Deliveries.

    PubMed

    Collado, Maria Carmen; Santaella, Marina; Mira-Pascual, Laia; Martínez-Arias, Elena; Khodayar-Pardo, Parisá; Ros, Gaspar; Martínez-Costa, Cecilia

    2015-10-19

    Breast milk (BM) is considered as a reference for infant nutrition. The role of bioactive components, such as cytokines, hormones, growth factors (GFs) and fatty acids (FAs) is poorly known, but they might be implicated in immune response development. The aim of this study was to identify the lipid profile and the spectrum of cytokines and neuronal GF in BM samples and analyse the influence of gestational age and lactation time on these components. This study used a longitudinal prospective method for the characterization of cytokines, FAs and GFs global profiles in 120 BM samples from 40 healthy mothers (20 preterm and 20 term) collected as colostrum, transitional and mature milk. The cytokines were analysed by protein array (Ray Bio® Human Cytokine Array G6. Ray Biotech, Inc. Norcross, GA, USA) and the FAs were analysed by gas chromatography. The FA profile was similar between the term and the preterm BM samples. Omega-3-α-linoleic and docosahexaenoic acid (DHA) and omega-6-linoleic acid were the most abundant in the term and preterm samples during lactation. Omega-3 ETA and omega-3 EPA we observed exclusively in the preterm samples. The cytokine profile showed a different trend based on gestational age. A significantly higher expression of neurotrophic factors was found in the mature preterm milk samples as compared to the mature term samples. Our study is the first to identify the influence and interactions of perinatal factors on cytokine, GFs and FAs in human milk.

  3. Longitudinal Study of Cytokine Expression, Lipid Profile and Neuronal Growth Factors in Human Breast Milk from Term and Preterm Deliveries

    PubMed Central

    Collado, Maria Carmen; Santaella, Marina; Mira-Pascual, Laia; Martínez-Arias, Elena; Khodayar-Pardo, Parisá; Ros, Gaspar; Martínez-Costa, Cecilia

    2015-01-01

    Breast milk (BM) is considered as a reference for infant nutrition. The role of bioactive components, such as cytokines, hormones, growth factors (GFs) and fatty acids (FAs) is poorly known, but they might be implicated in immune response development. The aim of this study was to identify the lipid profile and the spectrum of cytokines and neuronal GF in BM samples and analyse the influence of gestational age and lactation time on these components. This study used a longitudinal prospective method for the characterization of cytokines, FAs and GFs global profiles in 120 BM samples from 40 healthy mothers (20 preterm and 20 term) collected as colostrum, transitional and mature milk. The cytokines were analysed by protein array (Ray Bio® Human Cytokine Array G6. Ray Biotech, Inc. Norcross, GA, USA) and the FAs were analysed by gas chromatography. The FA profile was similar between the term and the preterm BM samples. Omega-3-α-linoleic and docosahexaenoic acid (DHA) and omega-6-linoleic acid were the most abundant in the term and preterm samples during lactation. Omega-3 ETA and omega-3 EPA we observed exclusively in the preterm samples. The cytokine profile showed a different trend based on gestational age. A significantly higher expression of neurotrophic factors was found in the mature preterm milk samples as compared to the mature term samples. Our study is the first to identify the influence and interactions of perinatal factors on cytokine, GFs and FAs in human milk. PMID:26492267

  4. Leukocyte Lysis and Cytokine Induction by the Human Sexually Transmitted Parasite Trichomonas vaginalis

    PubMed Central

    Mercer, Frances; Diala, Fitz Gerald I.; Chen, Yi-Pei; Molgora, Brenda M.; Ng, Shek Hang; Johnson, Patricia J.

    2016-01-01

    Trichomonas vaginalis (Tv) is an extracellular protozoan parasite that causes the most common non-viral sexually transmitted infection: trichomoniasis. While acute symptoms in women may include vaginitis, infections are often asymptomatic, but can persist and are associated with medical complications including increased HIV susceptibility, infertility, pre-term labor, and higher incidence of cervical cancer. Heightened inflammation resulting from Tv infection could account for these complications. Effective cellular immune responses to Tv have not been characterized, and re-infection is common, suggesting a dysfunctional adaptive immune response. Using primary human leukocyte components, we have established an in vitro co-culture system to assess the interaction between Tv and the cells of the human immune system. We determined that in vitro, Tv is able to lyse T-cells and B-cells, showing a preference for B-cells. We also found that Tv lysis of lymphocytes was mediated by contact-dependent and soluble factors. Tv lysis of monocytes is far less efficient, and almost entirely contact-dependent. Interestingly, a common symbiont of Tv, Mycoplasma hominis, did not affect cytolytic activity of the parasite, but had a major impact on cytokine responses. M. hominis enabled more diverse inflammatory cytokine secretion in response to Tv and, of the cytokines tested, Tv strains cleared of M. hominis induced only IL-8 secretion from monocytes. The quality of the adaptive immune response to Tv is therefore likely influenced by Tv symbionts, commensals, and concomitant infections, and may be further complicated by direct parasite lysis of effector immune cells. PMID:27529696

  5. Leukotriene B4 potentiates CpG signaling for enhanced cytokine secretion by human leukocytes.

    PubMed

    Gaudreault, Eric; Gosselin, Jean

    2009-08-15

    TLRs are known to be important in innate host defense against a variety of microbial infections. In particular, TLR9 has been associated with immune defense against different foreign organisms by recognition of unmethylated DNA sequences. In this report, we provide evidence that leukotriene B(4) (LTB(4)) has the capacity to modulate TLR9 expression on human neutrophils. The effect of LTB(4) was found to be specific, because related leukotrienes such as LTC(4) and LTD(4) or neutrophil agonists IL-8 and C5a failed to modulate TLR9 expression in neutrophils. Using fluorochrome-tagged CpG DNA, we observed that LTB(4) treatment also increased TLR9 ligand binding in neutrophils. Moreover, LTB(4) stimulation potentiates CpG-mediated signaling via an endosome-independent mechanism in human neutrophils, leading to enhanced secretion of proinflammatory cytokines. The increase in cytokine secretion by LTB(4) following CpG stimulation of neutrophils was associated with the activation of TGF-beta-activated kinase (TAK-1) as well as p38 and c-Jun (JNK) kinases. In contrast, in PBMC LTB(4) leads to an increase in cytokine secretion following CpG stimulation but via a MyD88- and endosome-dependent mechanism. As observed in neutrophils, PBMC stimulation with LTB(4) in the presence of CpG also results in enhanced TAK-1, p38, and JNK phosphorylation/activation. These data provide new evidence underlying the immunomodulatory properties of LTB(4) leading to antimicrobial defense.

  6. Injury Induces Localized Airway Increases in Pro-Inflammatory Cytokines in Humans and Mice

    PubMed Central

    Jonker, Mark A.; Hermsen, Joshua L.; Gomez, F. Enrique; Sano, Yoshifumi

    2011-01-01

    Abstract Background Secretory immunoglobulin A (sIgA) increases in the airways of humans and mice after injury to protect against infection. The pro-inflammatory cytokines tumor necrosis factor (TNF)-α, interleukin (IL)-1β, and IL-6 are linked molecularly to sIgA production and secretion and are required for sIgA increases in the airway after injury in a mouse model. We investigated the injury effect on airway and serum concentrations to determine the source of the cytokines involved in the airway IgA response. Methods In the first experiment, TNF-α, IL-1β, and IL-6 concentrations in bronchoalveolar lavage (BAL) fluid and serum obtained from 11 ventilated trauma patients within 30 h of admission were compared with those in eight elective surgical patients. In the second experiment, male ICR mice received no injury (n = 7) or injury with sham celiotomy and neck incisions (n = 8) with sacrifice of all animals at 8 h for BAL fluid and serum cytokine measurements by enzyme-linked immunosorbent assay. Results Injured patients had significantly higher BAL fluid and serum TNF-α, IL-1β, and IL-6 concentrations, with greater increases in the BAL fluid than in the serum. Injured mice had significantly increased BAL fluid concentrations of TNF-α, IL-1β, and IL-6 without significant changes in serum TNF-α or IL-1β. Serum IL-6 increased significantly. Conclusions Injury significantly increases human and mouse airway TNF-α, IL-1β, and IL-6. Increases are greater in the airway than in serum, implying a local rather than a systemic stress response to injury. PMID:21166596

  7. 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

  8. Leukocyte Lysis and Cytokine Induction by the Human Sexually Transmitted Parasite Trichomonas vaginalis.

    PubMed

    Mercer, Frances; Diala, Fitz Gerald I; Chen, Yi-Pei; Molgora, Brenda M; Ng, Shek Hang; Johnson, Patricia J

    2016-08-01

    Trichomonas vaginalis (Tv) is an extracellular protozoan parasite that causes the most common non-viral sexually transmitted infection: trichomoniasis. While acute symptoms in women may include vaginitis, infections are often asymptomatic, but can persist and are associated with medical complications including increased HIV susceptibility, infertility, pre-term labor, and higher incidence of cervical cancer. Heightened inflammation resulting from Tv infection could account for these complications. Effective cellular immune responses to Tv have not been characterized, and re-infection is common, suggesting a dysfunctional adaptive immune response. Using primary human leukocyte components, we have established an in vitro co-culture system to assess the interaction between Tv and the cells of the human immune system. We determined that in vitro, Tv is able to lyse T-cells and B-cells, showing a preference for B-cells. We also found that Tv lysis of lymphocytes was mediated by contact-dependent and soluble factors. Tv lysis of monocytes is far less efficient, and almost entirely contact-dependent. Interestingly, a common symbiont of Tv, Mycoplasma hominis, did not affect cytolytic activity of the parasite, but had a major impact on cytokine responses. M. hominis enabled more diverse inflammatory cytokine secretion in response to Tv and, of the cytokines tested, Tv strains cleared of M. hominis induced only IL-8 secretion from monocytes. The quality of the adaptive immune response to Tv is therefore likely influenced by Tv symbionts, commensals, and concomitant infections, and may be further complicated by direct parasite lysis of effector immune cells.

  9. Stimulatory effect of Aggregatibacter actinomycetemcomitans DNA on proinflammatory cytokine expression by human gingival fibroblasts.

    PubMed

    Soto-Barreras, Uriel; Cortés-Sandoval, Gabriela; Dominguez-Perez, Ruben; Loyola-Leyva, Alejandra; Martinez-Rodriguez, Panfilo-Raymundo; Loyola-Rodriguez, Juan Pablo

    2017-10-01

    While different virulence factors have been reported of Aggregatibacter actinomycetemcomitans (Aa), there is little information about the stimulatory effect of its DNA. The main purpose of this study was to assess the inflammatory response of human gingival fibroblasts (HGFs) stimulated with A. actinomycetemcomitans DNA. Cytokine levels of IL-6, IL-1α and TNF-α were measured on the supernatant of HGFs activated with 10, 25, 50 and 100μg/ml DNA of Aa during 24h. Primary cultures of HGFs were infected with Aa and its DNA at different times and concentrations to compare its cytotoxic effect. Cell damage and adhesion of Aa to HGFs were evaluated under light microscopy and Scanning electron microscopy respectively. There was a statistical difference (p<0.05) in cytokine expression in HGFs activated by bacterial DNA with a dose dependent on IL-6 expression and a significantly elevated expression of IL-1α and TNF-α compared to Human DNA negative control. Substantial morphological alterations were observed after infection of A. actinomycetemcomitans in HGFs but not with bDNA exposure. Aggregatibacter actinomycetemcomitans showed a high rate of adhesion and cell damage to HGFs after 30min. Genomic DNA of A. actinomycetemcomitans could be a factor in the pathogenesis of periodontitis that might play a major role in the inflammatory response. Copyright © 2017 Elsevier Ltd. All rights reserved.

  10. Concurrent detection of secreted products from human lymphocytes by microengraving: cytokines and antigen-reactive antibodies

    PubMed Central

    Bradshaw, Elizabeth M.; Kent, Sally C.; Tripuraneni, Vinay; Orban, Tihamer; Ploegh, Hidde L.; Hafler, David A.; Love, J. Christopher

    2008-01-01

    Cell surface determinants, cytokines and antibodies secreted by hematopoietic cells are used to classify their lineage and function. Currently available techniques are unable to elucidate multiple secreted proteins while also assigning phenotypic surface-displayed markers to the individual living cells. Here, a soft lithographic method, microengraving, was adapted for the multiplexed interrogation of populations of individual human peripheral blood mononuclear cells for secreted cytokines (IFN-γ and IL-6), antigen-specific antibodies, and lineage-specific surface-expressed markers. Application of the method to a clinical sample from a recent onset Type 1 diabetic subject with a positive titer of anti-insulin antibodies showed that ~0.58% of circulating CD19+ B cells secreted proinsulin-reactive antibodies of the IgG isotype and 2–3% of circulating cells secreted IL-6. These data demonstrate the utility of microengraving for interrogating multiple phenotypes of single human cells concurrently and for detecting rare populations of cells by their secreted products. PMID:18675591

  11. B7 expression and antigen presentation by human brain endothelial cells: requirement for proinflammatory cytokines.

    PubMed

    Prat, A; Biernacki, K; Becher, B; Antel, J P

    2000-02-01

    Interaction between systemic immune cells with cells of the blood-brain barrier is a central step in development of CNS-directed immune responses. Endothelial cells are the first cells of the blood-brain barrier encountered by migrating lymphocytes. To investigate the antigen-presenting capacity of human adult brain endothelial cells (HBECs), we used HBECs derived from surgically resected temporal lobe tissue, cocultured with allogeneic peripheral blood derived CD4+ T lymphocytes. HBECs in response to IFN-gamma, but not under basal culture conditions, expressed HLA-DR, B7.1 and B7.2 antigens. Despite such up-regulation, these IFN-gamma-treated HBECs, in contrast to human microglia and PB monocytes, did not sustain allogeneic CD4+ cell proliferation, supported only low levels of IL-2 and IFN-gamma production, and did not stimulate IL-2 receptor expression. CD4+ T cell proliferation and increased IL-2 receptor expression could be obtained by addition of IL-2. Our data suggests that, although HBECs cannot alone support T cell proliferation and cytokine production, HBECs acting in concert with cytokines derived from a proinflammatory environment could support such a response.

  12. 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.

  13. 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.

  14. Disruption of actin cytoskeleton enhanced cytokine synthesis of splenocytes stimulated with beta-glucan from the cauliflower medicinal mushroom, Sparassis crispa Wulf.:Fr. (higher Basidiomycetes) in vitro.

    PubMed

    Shibata, Asami; Hida, Toshie H; Ishibashi, Ken-Ichi; Miura, Noriko N; Adachi, Yoshiyuki; Ohno, Naohito

    2012-01-01

    Beta-glucan (BG) is a representative pathogen-associated molecular pattern (PAMP) produced by pathogenic fungi. SCG is a BG obtained from Sparassis crispa, which stimulates splenocytes in DBA/2 mice to produce cytokines, such as GM-CSF, IFN-γ, and TNF-α. In the present study, we analyzed the molecular mechanism of SCG-mediated cytokine synthesis using cytocharasin D (CytD), an inhibitor of actin polymerization. It was found that GM-CSF and TNF-α synthesis of splenocytes stimulated with SCG, but not with lipopolysaccharide, was significantly enhanced in the presence of CytD. CRDO, partially hydrolyzed linear 1,3-BG curdlan, stimulated splenocytes of DBA/2 mice slightly to produce cytokines. CRDO, acting as an antagonist in the presence of SCG, changed to a strong agonist in the presence of CytD. CytD also enhanced cytokine synthesis of bone marrow-derived dendritic cells. Taken together, cytokine productivity of BG was significantly dependent on molecular weight, and CytD treatment is useful to enhance the sensitivity for analyzing the immunostimulating activity of BG in vitro.

  15. Anti-Inflammatory Cytokines Predominate in Acute Human Plasmodium knowlesi Infections

    PubMed Central

    Cox-Singh, Janet; Singh, Balbir; Daneshvar, Cyrus; Planche, Timothy; Parker-Williams, John; Krishna, Sanjeev

    2011-01-01

    Plasmodium knowlesi has entered the human population of Southeast Asia. Naturally acquired knowlesi malaria is newly described with relatively little available data, including data on the host response to infection. Therefore pre-treatment cytokine and chemokine profiles were determined for 94 P. knowlesi, and for comparison, 20, P. vivax and 22 P. falciparum, patients recruited in Malaysian Borneo. Nine, five and one patient with P. knowlesi, P. falciparum and P. vivax respectively had complicated malaria as defined by World Health Organisation. Patients with uncomplicated P. knowlesi had lower levels of the pro-inflammatory cytokines IL-8 and TNFα than those with complicated disease (both p<0.05, Dunn's post test, DPT). The anti-inflammatory cytokines IL-1ra and IL-10 were detected in all patients in the study. IL-1ra, the most abundant cytokine measured, correlated with parasitaemia in P. knowlesi (rs = 0.47, p =  <0.0001), P. vivax (rs = 0.61, p = 0.0042) and P. falciparum (rs = 0.57,p = 0.0054) malaria. IL-10 correlated with parasitaemia in both P. knowlesi (rs = 0.54, p =  <0.0001) and P. vivax (rs = 0.78, p =  <0.0001) infections. There were between group differences in soluble markers of macrophage activation (MIP-1β and MCP-1). P. knowlesi patients had significantly lower levels of MIP-1β than P. falciparum (DPT, p =  <0.01). Uncomplicated P. knowlesi patients had significantly lower levels of MCP-1 than uncomplicated P. falciparum patients (DPT, p =  <0.001). There was no significant difference between complicated and uncomplicated P. knowlesi infections. MCP-1, MIP-1β, IL-8 and TNFα increased in complicated P. knowlesi but decreased in complicated P. falciparum infections. Descriptions of human knowlesi malaria provide a comparative means to discover mediators of pathophysiology in severe P. knowlesi as well as P. falciparum malaria. Crucially, P. knowlesi may be the disease and experimental primate

  16. Differential subnetwork of chemokines/cytokines in human, mouse, and rat brain cells after oxygen-glucose deprivation.

    PubMed

    Du, Yang; Deng, Wenjun; Wang, Zixing; Ning, MingMing; Zhang, Wei; Zhou, Yiming; Lo, Eng H; Xing, Changhong

    2016-01-01

    Mice and rats are the most commonly used animals for preclinical stroke studies, but it is unclear whether targets and mechanisms are always the same across different species. Here, we mapped the baseline expression of a chemokine/cytokine subnetwork and compared responses after oxygen-glucose deprivation in primary neurons, astrocytes, and microglia from mouse, rat, and human. Baseline profiles of chemokines (CX3CL1, CXCL12, CCL2, CCL3, and CXCL10) and cytokines (IL-1α, IL-1β, IL-6, IL-10, and TNFα) showed significant differences between human and rodents. The response of chemokines/cytokines to oxygen-glucose deprivation was also significantly different between species. After 4 h oxygen-glucose deprivation and 4 h reoxygenation, human and rat neurons showed similar changes with a downregulation in many chemokines, whereas mouse neurons showed a mixed response with up- and down-regulated genes. For astrocytes, subnetwork response patterns were more similar in rats and mice compared to humans. For microglia, rat cells showed an upregulation in all chemokines/cytokines, mouse cells had many down-regulated genes, and human cells showed a mixed response with up- and down-regulated genes. This study provides proof-of-concept that species differences exist in chemokine/cytokine subnetworks in brain cells that may be relevant to stroke pathophysiology. Further investigation of differential gene pathways across species is warranted.

  17. Optimising the quantification of cytokines present at low concentrations in small human mucosal tissue samples using Luminex assays☆

    PubMed Central

    Staples, Emily; Ingram, Richard James Michael; Atherton, John Christopher; Robinson, Karen

    2013-01-01

    Sensitive measurement of multiple cytokine profiles from small mucosal tissue biopsies, for example human gastric biopsies obtained through an endoscope, is technically challenging. Multiplex methods such as Luminex assays offer an attractive solution but standard protocols are not available for tissue samples. We assessed the utility of three commercial Luminex kits (VersaMAP, Bio-Plex and MILLIPLEX) to measure interleukin-17A (IL-17) and interferon-gamma (IFNγ) concentrations in human gastric biopsies and we optimised preparation of mucosal samples for this application. First, we assessed the technical performance, limits of sensitivity and linear dynamic ranges for each kit. Next we spiked human gastric biopsies with recombinant IL-17 and IFNγ at a range of concentrations (1.5 to 1000 pg/mL) and assessed kit accuracy for spiked cytokine recovery and intra-assay precision. We also evaluated the impact of different tissue processing methods and extraction buffers on our results. Finally we assessed recovery of endogenous cytokines in unspiked samples. In terms of sensitivity, all of the kits performed well within the manufacturers' recommended standard curve ranges but the MILLIPLEX kit provided most consistent sensitivity for low cytokine concentrations. In the spiking experiments, the MILLIPLEX kit performed most consistently over the widest range of concentrations. For tissue processing, manual disruption provided significantly improved cytokine recovery over automated methods. Our selected kit and optimised protocol were further validated by measurement of relative cytokine levels in inflamed and uninflamed gastric mucosa using Luminex and real-time polymerase chain reaction. In summary, with proper optimisation Luminex kits (and for IL-17 and IFNγ the MILLIPLEX kit in particular) can be used for the sensitive detection of cytokines in mucosal biopsies. Our results should help other researchers seeking to quantify multiple low concentration cytokines in

  18. Modulation of mdr1 expression by cytokines in human colon carcinoma cells: an approach for reversal of multidrug resistance.

    PubMed Central

    Stein, U.; Walther, W.; Shoemaker, R. H.

    1996-01-01

    Reversal of multidrug resistance (MDR) may offer a means of increasing the effectiveness of tumour chemotherapy. A variety of recent evidence indicates that cytokines may be particularly useful in this endeavour. To investigate the molecular mechanism by which cytokines may sensitise multidrug-resistant colon carcinoma cells, HCT15 and HCT116, to treatment with MDR-related drugs, we evaluated the effects of the human cytokines tumour necrosis factor alpha (TNF alpha), interleukin 2 (IL-2) and interferon gamma (IFN gamma) on mdr1 gene expression at the mRNA level by reverse transcription-polymerase chain reaction (RT-PCR) and at the protein level with monoclonal antibodies by immuno flow cytometry. P-glycoprotein function was examined after accumulation of the fluorescent drug, doxorubicin, by flow cytometry. Chemosensitivity to doxorubicin and vincristine was analysed using the XTT assay. All three cytokines were found to modulate the MDR characteristics on mdr1 expression levels, P-glycoprotein function and measured chemosensitivity to MDR-associated anti-cancer drugs. This cytokine-induced reversal of MDR was strongly time dependent, with maximal effects after 48 and 72 h of cytokine treatment. If similar modulation of MDR phenotype can be obtained in in vivo models, it may be possible to verify the time course for modulation by cytokine treatment and to design appropriate clinical trials of this strategy for MDR reversal. Images Figure 1 PMID:8912533

  19. Human Brain Chemokine and Cytokine Expression in Sepsis: A Report of Three Cases.

    PubMed

    Warford, Jordan; Lamport, Anna-Claire; Kennedy, Barry; Easton, Alexander S

    2017-01-01

    Sepsis is a systemic response to infection that can affect brain function by inducing resident cells (including astrocytes and microglia) to generate brain chemokines and cytokines. However, there are few studies on the human brain. Since this information may shed further light on pathogenesis, our study objective was to measure the expression of 36 chemokines and cytokines in autopsied brain from 3 cases of sepsis and 10 controls, and to relate this to astrocyte and microglial activation. The right frontal pole was removed at autopsy and chemokine and cytokine expression measured by multiplexed enzyme-linked immunosorbent assay and real-time quantitative polymerase chain reaction (qPCR). Immunohistochemistry and image analysis were carried out to determine the expression of glial fibrillary acidic protein (GFAP), a marker of activated astrocytes, and CD68 and CD45, markers of activated microglial cells. Concentrations of the chemokines CXCL8, CXCL10, CXCL12, CCL13 and CCL22 were increased in pooled data from the three cases of sepsis (p<0.05); however, their messenger RNA (mRNA) expression was unaltered. CXCL13, CXCL1, CXCL2, CCL1, CCL2, CCL8, CCL20, (interleukin) IL-16, IL-1β and (tumour necrosis factor) TNF concentrations showed increases in two of three sepsis cases. Additionally, individual sepsis cases showed increases in mRNA expression for HDAC (histone deacetylase) 6 and EIF (eukaryotic translation initiation factor) 4A2. Brain GFAP expression was significantly increased (p<0.05) in pooled data from the three sepsis cases. Individual sepsis cases showed increases in CD68 or CD45 expression. These expression patterns add to our understanding of the pathogenesis of sepsis and its effects on the brain.

  20. Dynamic interactions between plasma IL-1 family cytokines and central endogenous opioid neurotransmitter function in humans.

    PubMed

    Prossin, Alan R; Zalcman, Steven S; Heitzeg, Mary M; Koch, Alisa E; Campbell, Phillip L; Phan, K Luan; Stohler, Christian S; Zubieta, Jon-Kar

    2015-02-01

    Evidence in animal models suggests IL-1 family cytokines interact with central endogenous opioid neurotransmitter systems, inducing or perpetuating pathological states such as persistent pain syndromes, depression, substance use disorders, and their comorbidity. Understanding these interactions in humans is particularly relevant to understanding pathological states wherein this neurotransmitter system is implicated (ie, persistent pain, mood disorders, substance use disorders, etc). Here, we examined relationships between IL-1β, IL-1ra, and functional measures of the endogenous opioid system in 34 healthy volunteers, in the absence and presence of a standardized sustained muscular pain challenge, a psychophysical challenge with emotionally and physically stressful components. Mu-opioid receptor availability in vivo was examined with [(11)C]carfentanil positron emission tomography (PET) scanning. Sex and neuroticism impacted IL-1 family cytokines; higher baseline IL-1β and IL-1ra was identified in females with lower neuroticism. Higher baseline IL-1β was also associated with reduced μ-opioid receptor availability (amygdala) and greater pain sensitivity. The pain challenge increased IL-1β in females with high neuroticism. Strong associations between IL-1ra (an anti-nociceptive cytokine) and μ-opioid receptor activation (VP/NAcc) were identified during the pain challenge and the resulting analgesic effect of μ-opioid receptor activation was moderated by changes in IL-1β whereby volunteers with greater pain induced increase in IL-1β experienced less endogenous opioid analgesia. This study demonstrates the presence of relationships between inflammatory factors and a specific central neurotransmitter system and circuitry, of relevance to understanding interindividual variations in regulation of responses to pain and other physical and emotional stressors.

  1. Effects of Rilpivirine on Human Adipocyte Differentiation, Gene Expression, and Release of Adipokines and Cytokines

    PubMed Central

    Díaz-Delfín, Julieta; Domingo, Pere; Mateo, Maria Gracia; Gutierrez, Maria del Mar; Domingo, Joan Carles; Giralt, Marta

    2012-01-01

    Rilpivirine is a nonnucleoside reverse transcriptase inhibitor (NNRTI) recently developed as a drug of choice for initial antiretroviral treatment of HIV-1 infection. Disturbances in lipid metabolism and, ultimately, in adipose tissue distribution and function are common concerns as secondary effects of antiretroviral treatment. Efavirenz, the most commonly used NNRTI, causes mild dyslipidemic effects in patients and strongly impaired adipocyte differentiation in vitro. In this study, we provide the first demonstration of the effects of rilpivirine on human adipocyte differentiation, gene expression, and release of regulatory proteins (adipokines and cytokines) and compare them with those caused by efavirenz. Rilpivirine caused a repression of adipocyte differentiation that was associated with impaired expression of the master adipogenesis regulators peroxisome proliferator-activated receptor gamma (PPARγ), CCAAT enhancer binding protein alpha (C/EBPα), and sterol regulatory element binding transcription factor 1 (SREBP-1) and their target genes encoding lipoprotein lipase and the adipokines leptin and adiponectin. Rilpivirine also repressed adiponectin release by adipocytes, but only at high concentrations, and did not alter leptin release. Rilpivirine induced the release of proinflammatory cytokines (interleukin-6 and -8, monocyte chemoattractant protein 1 [MCP-1], plasminogen activator inhibitor type 1 [PAI-1]) only at very high concentrations (10 μM). A comparison of the effects of rilpivirine and efavirenz at the same concentration (4 μM) or even at lower concentrations of efavirenz (2 μM) showed that rilpivirine-induced impairment of adipogenesis and induction of proinflammatory cytokine expression and release were systematically milder than those of efavirenz. It is concluded that rilpivirine causes an antiadipogenic and proinflammatory response pattern, but only at high concentrations, whereas efavirenz causes similar effects at lower concentrations

  2. Dynamic Interactions Between Plasma IL-1 Family Cytokines and Central Endogenous Opioid Neurotransmitter Function in Humans

    PubMed Central

    Prossin, Alan R; Zalcman, Steven S; Heitzeg, Mary M; Koch, Alisa E; Campbell, Phillip L; Phan, K Luan; Stohler, Christian S; Zubieta, Jon-Kar

    2015-01-01

    Evidence in animal models suggests IL-1 family cytokines interact with central endogenous opioid neurotransmitter systems, inducing or perpetuating pathological states such as persistent pain syndromes, depression, substance use disorders, and their comorbidity. Understanding these interactions in humans is particularly relevant to understanding pathological states wherein this neurotransmitter system is implicated (ie, persistent pain, mood disorders, substance use disorders, etc). Here, we examined relationships between IL-1β, IL-1ra, and functional measures of the endogenous opioid system in 34 healthy volunteers, in the absence and presence of a standardized sustained muscular pain challenge, a psychophysical challenge with emotionally and physically stressful components. Mu-opioid receptor availability in vivo was examined with [11C]carfentanil positron emission tomography (PET) scanning. Sex and neuroticism impacted IL-1 family cytokines; higher baseline IL-1β and IL-1ra was identified in females with lower neuroticism. Higher baseline IL-1β was also associated with reduced μ-opioid receptor availability (amygdala) and greater pain sensitivity. The pain challenge increased IL-1β in females with high neuroticism. Strong associations between IL-1ra (an anti-nociceptive cytokine) and μ-opioid receptor activation (VP/NAcc) were identified during the pain challenge and the resulting analgesic effect of μ-opioid receptor activation was moderated by changes in IL-1β whereby volunteers with greater pain induced increase in IL-1β experienced less endogenous opioid analgesia. This study demonstrates the presence of relationships between inflammatory factors and a specific central neurotransmitter system and circuitry, of relevance to understanding interindividual variations in regulation of responses to pain and other physical and emotional stressors. PMID:25139063

  3. TGF-β2, a protective intestinal cytokine, is abundant in maternal human milk and human-derived fortifiers but not in donor human milk.

    PubMed

    Reeves, Aaron A; Johnson, Marney C; Vasquez, Margarita M; Maheshwari, Akhil; Blanco, Cynthia L

    2013-12-01

    This study compared cytokines (in particular transforming growth factor [TGF]-β2) and lactoferrin in maternal human milk (MHM), human-derived milk fortifier (HDMF), and donor human milk (DHM). MHM was randomly collected from breastfeeding mothers who had no infectious illness at the time of milk expression. HDMF and DHM were products derived from human milk processed by Holder pasteurization. MHM samples were collected at different times (early/late) and gestations (preterm/term). Lactoferrin was analyzed by western blotting, and cytokines were quantified using commercial enzyme-linked immunosorbent assays. Significance was determined using analysis of variance. In the 164 samples analyzed, TGF-β2 concentrations in HDMF and preterm MHM (at all collection times) were fivefold higher than in DHM (p<0.05). Early preterm MHM had levels of interleukin (IL)-10 and IL-18, 11-fold higher than DHM (p<0.05). IL-6 in DHM was 0.3% of the content found in MHM. IL-18 was fourfold higher in early MHM versus late MHM regardless of gestational age (p<0.05). Lactoferrin concentration in DHM was 6% of that found in MHM. Pasteurization decreases concentrations of most cytokines and lactoferrin in DHM. TGF-β2, a protective intestinal cytokine, has comparable concentrations in HDMF to MHM despite pasteurization.

  4. 17beta-Estradiol inhibits cytokine induction of the human E-selectin promoter.

    PubMed

    Tyree, Curtis M; Zou, Aihua; Allegretto, Elizabeth A

    2002-03-01

    Estradiol has been shown to decrease levels of the cell adhesion molecule E-selectin in cultured cells and in women on hormone replacement therapy. We set out to determine if the mechanism of estradiol action on E-selectin is at the level of its promoter. It was found that estradiol repressed the cytokine-stimulated induction of luciferase activity driven by the human E-selectin promoter in a reporter plasmid (hE-sel-LUC) in co-transfected human hepatoma cells (Hep G2) and human umbilical cord endothelial cells (ECV-304). Repression by estradiol was dependent on the presence of transfected estrogen receptor (ER) alpha or beta expression vectors. The ER antagonist ICI-182,780 blocked the repression by estradiol, confirming the receptor-dependence of the effect. The intact DNA-binding domain of ERalpha was required for estradiol repression of the cytokine-induced stimulation of the promoter in each cell line as demonstrated by the inability of an ER construct with two point mutations in the DNA-binding domain to inhibit reporter activity. Mutation of the NFK-B site at -94 to -85 within the E-selectin promoter led to less stimulation of hE-sel-LUC by interleukin one beta (IL-1beta). Estradiol did not inhibit this IL-1beta stimulated luciferase activity, indicating that the NFK-B site is necessary for ER-mediated inhibition of this promoter. Mutation of the AP-1 site at -500 to -494 within the E-selectin promoter had no effect on the ability of IL-1beta to stimulate its transcription, and estradiol repressed this activation in an ER-dependent manner with identical efficacy and potency in comparison with the wild-type promoter. Therefore, the E-selectin promoter is down-regulated by estradiol working through either ERalpha or ERbeta and requires the NFK-B site at -94 to -85 within the promoter.

  5. Control of pro-inflammatory cytokine release from human monocytes with the use of an interleukin-10 monoclonal antibody.

    PubMed

    Patel, Hardik; Davidson, Dennis

    2014-01-01

    The monocytes (MONOs) can be considered as "double-edge swords"; they have both important pro-inflammatory and anti-inflammatory functions manifested in part by cytokine production and release. Although MONOs are circulating cells, they are the major precursors of a variety of tissue-specific immune cells such as the alveolar macrophage, dendritic cells, microglial cells, and Kupffer cells. Unlike the polymorphonuclear leukocyte, which produces no or very little interleukin-10 (IL-10), the monocyte can produce this potent anti-inflammatory cytokine to control inflammation. IL-10, on an equimolar basis, is a more potent inhibitor of pro-inflammatory cytokines produced by monocytes than many anti-inflammatory glucocorticoids which are used clinically. This chapter describes how to isolate monocytes from human blood and the use of IL-10 monoclonal antibody to determine the effect and timing of endogenous IL-10 release on the production and release of pro-inflammatory cytokines.

  6. Viability, apoptosis, proliferation, activation, and cytokine secretion of human keratoconus keratocytes after cross-linking.

    PubMed

    Song, Xuefei; Stachon, Tanja; Wang, Jiong; Langenbucher, Achim; Seitz, Berthold; Szentmáry, Nóra

    2015-01-01

    The purpose of this study was to determine the impact of cross-linking (CXL) on viability, apoptosis, proliferation, activation, and cytokine secretion of human keratoconus (KC) keratocytes, in vitro. Primary KC keratocytes were cultured in DMEM/Ham's F12 medium supplemented with 10% FCS and underwent UVA illumination (370 nm, 2 J/cm(2)) during exposure to 0.1% riboflavin and 20% Dextran in PBS. Twenty-four hours after CXL, viability was assessed using Alamar blue assay; apoptosis using APO-DIRECT Kit; proliferation using ELISA-BrdU kit; and CD34 and alpha-smooth muscle actin (α-SMA) expression using flow cytometry. Five and 24 hours after CXL, FGFb, HGF, TGFβ1, VEGF, KGF, IL-1β, IL-6, and IL-8 secretion was measured using enzyme-linked-immunoabsorbent assay (ELISA). Following CXL, cell viability and proliferation decreased (P < 0.05; P = 0.009), the percentage of apoptotic keratocytes increased (P < 0.05) significantly, and CD34 and α-SMA expression remained unchanged (P > 0.06). Five hours after CXL, FGFb secretion increased significantly (P = 0.037); however no other cytokine secretion differed significantly from controls after 5 or 24 hours (P > 0.12). Cross-linking decreases viability, triggers apoptosis, and inhibits proliferation, without an impact on multipotent hematopoietic stem cell transformation and myofibroblastic transformation of KC keratocytes. CXL triggers FGFb secretion of KC keratocytes transiently (5 hours), normalizing after 24 hours.

  7. Thimerosal induces TH2 responses via influencing cytokine secretion by human dendritic cells.

    PubMed

    Agrawal, Anshu; Kaushal, Poonam; Agrawal, Sudhanshu; Gollapudi, Sastry; Gupta, Sudhir

    2007-02-01

    Thimerosal is an organic mercury compound that is used as a preservative in vaccines and pharmaceutical products. Recent studies have shown a TH2-skewing effect of mercury, although the underlying mechanisms have not been identified. In this study, we investigated whether thimerosal can exercise a TH2-promoting effect through modulation of functions of dendritic cells (DC). Thimerosal, in a concentration-dependent manner, inhibited the secretion of LPS-induced proinflammatory cytokines TNF-alpha, IL-6, and IL-12p70 from human monocyte-derived DC. However, the secretion of IL-10 from DC was not affected. These thimerosal-exposed DC induced increased TH2 (IL-5 and IL-13) and decreased TH1 (IFN-gamma) cytokine secretion from the T cells in the absence of additional thimerosal added to the coculture. Thimerosal exposure of DC led to the depletion of intracellular glutathione (GSH), and addition of exogenous GSH to DC abolished the TH2-promoting effect of thimerosal-treated DC, restoring secretion of TNF-alpha, IL-6, and IL-12p70 by DC and IFN-gamma secretion by T cells. These data suggest that modulation of TH2 responses by mercury and thimerosal, in particular, is through depletion of GSH in DC.

  8. Interleukin-13 as an important cytokine: A review on its roles in some human diseases.

    PubMed

    Seyfizadeh, Narges; Seyfizadeh, Nayer; Gharibi, Tohid; Babaloo, Zohreh

    2015-12-01

    Interleukin-13 (IL-13) as a pleiotropic cytokine acts through the IL-13Ra1/IL-4Ra complex to induce activation responses which contribute to the inflammatory diseases. Genetic polymorphisms in IL-13 and its receptor components have been proved to be associated with higher disease prevalence rates. Animal models such as in IL-13 deficient mice and transgenic animals also have been confirmed the critical role of this cytokine in the immune responses, mostly by IL-13 neutralization and IL-13/IL-4 dual neutralization strategies. This review highlights IL-13 structure as well as its pivotal roles in the normal physiologic and pathologic states. It is followed by a section on the recent findings on IL-13 receptors and signalling mechanisms to briefly summarize its functions in the immune systems. IL-13 roles in the human diseases such as asthma, systematic sclerosis, and some inflammatory diseases are described concisely. Finally some of the ongoing therapeutic applications are presented to comprehensively review IL-13 mediator roles.

  9. Combined effects of proinflammatory cytokines and intermittent cyclic mechanical strain in inhibiting osteogenicity in human periodontal ligament cells.

    PubMed

    Sun, Chaofan; Chen, Lijiao; Shi, Xinlian; Cao, Zhensheng; Hu, Bibo; Yu, Wenbin; Ren, Manman; Hu, Rongdang; Deng, Hui

    2016-09-01

    Mechanical strain plays an important role in bone formation and resorption during orthodontic tooth movement. The mechanism has not been fully studied, and the process becomes complex with increased amounts of periodontal patients seeking orthodontic care. Our aims were to elucidate the combined effects of proinflammatory cytokines and intermittent cyclic strain (ICS) on the osteogenic capacity of human periodontal ligament cells. Cultured human periodontal ligament cells were exposed to proinflammatory cytokines (interleukin-1β 5 ng/mL and tumor necrosis factor-α 10 ng/mL) for 1 and 5 days, and ICS (0.5 Hz, 12% elongation) was applied for 4 h per day. The autocrine of inflammatory cytokines was measured by enzyme-linked immunosorbent assay. The expression of osteoblast markers runt-related transcription factor 2 and rabbit collagen type I was determined using real-time polymerase chain reaction and Western blot. The osteogenic capacity was also detected by alkaline phosphatase (ALP) staining, ALP activity, and alizarin red staining. We demonstrated that ICS impaired the osteogenic capacity of human periodontal ligament cells when incubated with proinflammatory cytokines, as evidenced by the low expression of ALP staining, low ALP activity, reduced alizarin red staining, and reduced osteoblast markers. These data, for the first time, suggest that ICS has a negative effect on the inductive inhibition of osteogenicity in human PDL cells mediated by proinflammatory cytokines. © 2016 International Federation for Cell Biology.

  10. Peripheral CD4+ T cell cytokine responses following human challenge and re-challenge with Campylobacter jejuni.

    PubMed

    Fimlaid, Kelly A; Lindow, Janet C; Tribble, David R; Bunn, Janice Y; Maue, Alexander C; Kirkpatrick, Beth D

    2014-01-01

    Campylobacter jejuni is a leading cause of human gastroenteritis worldwide; however, our understanding of the human immune response to C. jejuni infection is limited. A previous human challenge model has shown that C. jejuni elicits IFNγ production by peripheral blood mononuclear cells, a response associated with protection from clinical disease following re-infection. In this study, we investigate T lymphocyte profiles associated with campylobacteriosis using specimens from a new human challenge model in which C. jejuni-naïve subjects were challenged and re-challenged with C. jejuni CG8421. Multiparameter flow cytometry was used to investigate T lymphocytes as a source of cytokines, including IFNγ, and to identify cytokine patterns associated with either campylobacteriosis or protection from disease. Unexpectedly, all but one subject evaluated re-experienced campylobacteriosis after re-challenge. We show that CD4+ T cells make IFNγ and other pro-inflammatory cytokines in response to infection; however, multifunctional cytokine response patterns were not found. Cytokine production from peripheral CD4+ T cells was not enhanced following re-challenge, which may suggest deletion or tolerance. Evaluation of alternative paradigms or models is needed to better understand the immune components of protection from campylobacteriosis.

  11. Effect of cadmium on the expression levels of interleukin-1α and interleukin-10 cytokines in human lung cells

    PubMed Central

    ODEWUMI, CAROLINE; LATINWO, LEKAN M.; SINCLAIR, ANDRE; BADISA, VEERA L.D.; ABDULLAH, AHKINYALA; BADISA, RAMESH B.

    2015-01-01

    Cadmium is an environmentally hazardous metal, which causes toxicity in humans. Inhalation of cigarette smoke and industrial fumes containing cadmium are sources of cadmium exposure. It is responsible for the malfunction of various organs, leading to disease particularly in the lungs, liver and kidneys. In the present study, the effect of cadmium chloride (CdCl2) on cell viability, and the expression levels of interleukin (IL)-1α and IL-10 cytokines at various concentrations and incubation durations were assessed in MRC-9 human normal lung and A549 human lung cancer cells to elucidate the mechanism of cadmium toxicity. Cell viability was measured using a crystal violet dye binding assay. The expression levels of the cytokines were measured by cytokine specific enzyme-linked immunosorbent assay kits. The viability assay results revealed higher sensitivity of the A549 lung cancer cells to CdCl2 compared with the normal MRC-9 lung cells. In the normal MRC-9 lung cells, higher expression levels of the cytokines were observed at the lowest CdCl2 concentration at a shorter exposure time compared with the lung cancer cells. Higher levels of the cytokines were observed in the A549 lung cancer cells at all other times and concentrations compared with the MRC-9 cells, indicating higher levels of inflammation. The cytokine levels were reduced at higher CdCl2 concentrations and longer exposure durations, demonstrating the toxic effect of cadmium. The results indicated that CdCl2 affected the expression levels of the cytokines and led to cytotoxicity in human lung cells, and suggested that compounds which reduce inflammation may prevent cadmium toxicity. PMID:26397147

  12. Staphylococcus aureus Biofilm and Planktonic cultures differentially impact gene expression, mapk phosphorylation, and cytokine production in human keratinocytes.

    PubMed

    Secor, Patrick R; James, Garth A; Fleckman, Philip; Olerud, John E; McInnerney, Kate; Stewart, Philip S

    2011-06-21

    Many chronic diseases, such as non-healing wounds are characterized by prolonged inflammation and respond poorly to conventional treatment. Bacterial biofilms are a major impediment to wound healing. Persistent infection of the skin allows the formation of complex bacterial communities termed biofilm. Bacteria living in biofilms are phenotypically distinct from their planktonic counterparts and are orders of magnitude more resistant to antibiotics, host immune response, and environmental stress. Staphylococcus aureus is prevalent in cutaneous infections such as chronic wounds and is an important human pathogen. The impact of S. aureus soluble products in biofilm-conditioned medium (BCM) or in planktonic-conditioned medium (PCM) on human keratinocytes was investigated. Proteomic analysis of BCM and PCM revealed differential protein compositions with PCM containing several enzymes involved in glycolysis. Global gene expression of keratinocytes exposed to biofilm and planktonic S. aureus was analyzed after four hours of exposure. Gene ontology terms associated with responses to bacteria, inflammation, apoptosis, chemotaxis, and signal transduction were enriched in BCM treated keratinocytes. Several transcripts encoding cytokines were also upregulated by BCM after four hours. ELISA analysis of cytokines confirmed microarray results at four hours and revealed that after 24 hours of exposure, S. aureus biofilm induced sustained low level cytokine production compared to near exponential increases of cytokines in planktonic treated keratinocytes. The reduction in cytokines produced by keratinocytes exposed to biofilm was accompanied by suppressed phosphorylation of MAPKs. Chemical inhibition of MAPKs did not drastically reduce cytokine production in BCM-treated keratinocytes suggesting that the majority of cytokine production is mediated through MAPK-independent mechanisms. Collectively the results indicate that S. aureus biofilms induce a distinct inflammatory response

  13. The influence of hypothalamic cytokine PRP on protein synthesis in brain subcellular compartments in crush syndrome.

    PubMed

    Guevorkian, Artashes G; Kanayan, Alexander S; Chailian, Gor G; Danielyan, Kristine E; Hayrapetyan, Hripsime L; Barsegyan, Karine A; Khachatryan, Hranush F; Galoyan, Armen A; Kevorkian, Guevork A

    2011-09-01

    Crush-syndrom (CS) was characterized by Bywaters E.G.L. in 1941 after London blitz. The soft tissues is followed by acute hemodynamic shock, myoglobinuria, acute renal insufficiency, and lethal endotoxicity. Data of CS pathogenesis study has shown that the largest changes in Crush occur during decompression and are accompanied by acute alteration of brain protein synthesis and strong morphological changes of brain structures. The period of decompression might be characterized by the proteolytic breakdown of the myoglobine and formation of toxic peptides. In our current work we have identified four newly formed peptides in the brain of the animals subjected to the experimental muscle tissue injury. Our investigations related with the CS experimental model have demonstrated that during the 2-hours compression protein synthesis was decreased in cytosol (32,7%) and mitochondria (49%), after 5-h compression there were registered non-significant changes in the level of protein synthesis. Intraperitoneal administration of Proline-rich peptide, ((PRP), 1 mcg/100g weight of rats), originating from proteolysis of C-terminal glycoprotein a neurophysin II along with vasopressin and oxytocin and transferring from the hypothalamus to the neurohypophysis by axonal transport, initiates activation of the protein synthesis in all studied cellular subcomponents of brain cells. The positive effect of the peptide is conditioned, most probably, by activation of the immune system and adaptation mechanisms, including mobilization of endogen-protective mechanisms of the organism.

  14. Benexate hydrochloride betadex modulates nitric oxide synthesis and cytokine expression in gastric ulcers

    PubMed Central

    Lee, Jae Min; Lim, Ji-Youn; Kim, Yoonjin; Kim, Ye Ji; Choi, Hyuk Soon; Kim, Eun Sun; Keum, Bora; Seo, Yeon Seok; Jeen, Yoon Tae; Lee, Hong Sik; Um, Soon Ho; Kim, Chang Duck; Ryu, Ho Sang; Sul, Donggeun; Hong, Junghwa; Chun, Hoon Jai

    2016-01-01

    The present study investigated benexate hydrochloride betadex (BHB)-mediated ulcer healing, and changes to microcirculation modulated through nitric oxide synthase (NOS) and anti-inflammatory activity. A rat model of gastric mucosal injury was established through injection of a 60% acetic acid solution into the stomach. Following ulcer induction, the rats were administered BHB orally for 5 days at doses of 0, 100, 300 or 1,000 mg/kg. The highest dose of BHB was also administered with or without L-NG-nitroarginine methyl ester (L-NAME). The area of gastric ulcers was determined by planimetry, and expression of cyclooxygenases (COX), cytokines and NOS in stomach tissues were measured using western blotting. Compared with the control group, gastric ulcer size was significantly decreased in the 1,000 mg/kg BHB-treated group (P<0.05). Administration of BHB led to a significant increase in endothelial (e)NOS expression (P<0.05). Although acetic acid co-treatment with L-NAME induced more severe mucosal damage, BHB decreased COX expression and tumor necrosis factor-α levels when administered with the nitric oxide inhibitor, L-NAME (P<0.05). BHB exhibited protective effects in a rat model of gastric ulcers, which were associated with a decrease in pro-inflammatory cytokine levels and the activation of eNOS. PMID:27446246

  15. Cyclic peptide *CRRETAWAC* attenuates fibronectin-induced cytokine secretion of human airway smooth muscle cells by inhibiting FAK and p38 MAPK.

    PubMed

    Chu, Mengdi; Ji, Jiani; Cao, Wenhao; Zhang, Huojun; Meng, Dan; Xie, Bangruan; Xu, Shuyun

    2017-10-01

    α5β1 integrin is highly expressed in airway smooth muscle cells and mediate the adhesion of airway smooth muscle cells to fibronectin to regulate airway remodelling in asthma. This study aimed to investigate the effects of synthetic cyclic peptide *CRRETAWAC* on fibronectin-induced cytokine secretion of airway smooth muscle cells and the underlying mechanism. Human airway smooth muscle cells were isolated and treated with fibronectin, IL-13, *CRRETAWAC* peptide, α5β1 integrin-blocking antibody, FAK inhibitor or p38 MAPK inhibitor. The transcription and secretion of eotaxin-1 and RANTES were detected by real-time PCR and ELISA, respectively. The phosphorylation of FAK and MAPKs including p38, ERK1/2 and JNK1/2 was detected by Western blot analysis. The transcription and secretion of eotaxin-1 and RANTES increased in airway smooth muscle cells cultured in fibronectin-coated plates. However, α5β1 integrin-blocking antibody, *CRRETAWAC* peptide, FAK inhibitor or p38 MAPK inhibitor significantly reduced mRNA levels and the secretion of eotaxin-1 and RANTES in airway smooth muscle cells cultured in fibronectin-coated plates. In addition, the phosphorylation of FAK and p38 MAPK was significantly increased in airway smooth muscle cells cultured in fibronectin-coated plates compared to the cells cultured in uncoated plates and was significantly reduced in airway smooth muscle cells treated with *CRRETAWAC* peptide. Fibronectin induces cytokine synthesis and secretion of airway smooth muscle cells. Peptide *CRRETAWAC* antagonizes fibronectin-induced cytokine synthesis and secretion of airway smooth muscle cells via the inhibition of FAK and p38 MAPK, and is a potential agent for the therapy of asthma. © 2017 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

  16. 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.

  17. 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.

  18. BH(4) (tetrahydrobiopterin)-dependent activation, but not the expression, of inducible NOS (nitric oxide synthase)-2 in proinflammatory cytokine-stimulated, cultured normal human astrocytes is mediated by MEK-ERK kinases.

    PubMed

    Chiarini, Anna; Dal Pra, Ilaria; Gottardo, Rossella; Bortolotti, Federica; Whitfield, James F; Armato, Ubaldo

    2005-03-01

    Nitric oxide (NO) from astrocytes is one of the signalers used by the brain's extensive glial-neuronal-vascular network, but its excessive production by pro-inflammatory cytokine-stimulated glial cells can be cytodestructive. Here, we show how three pro-inflammatory cytokines (IL-1beta, TNF-alpha, and IFN-gamma) together stimulated the activation, but not the prior expression, of NOS-2 protein via a mechanism involving MEK-ERKs protein kinases in astrocytes from adult human cerebral temporal cortex. The cytokines triggered a transient burst of p38 MAPK activity and the production of NOS-2 mRNA which were followed by bursts of MEK-ERK activities, synthesis of the NOS-2 co-factor tetrahydrobiopterin (BH(4)), a build-up of NOS-2 protein and from it active NOS-2 enzyme. Selectively inhibiting MEK1/MEK2, but not the earlier burst of p38 MAPK activity, with a brief exposure to U0126 between 24 and 24.5 h after adding the cytokine triad affected neither NOS-2 expression nor NOS-2 protein accumulation but stopped BH(4) synthesis and the assembly of the NOS-2 protein into active NOS-2 enzyme. The complete blockage of active NOS-2 production by the brief exposure to U0126 was bypassed by simply adding BH(4) to the culture medium. Therefore, this cytokine triad triggered two completely separable, tandem operating mechanisms in normal human astrocytes, the first being NOS-2 gene expression and accumulation of NOS-2 protein and the second being the synthesis of the BH(4) factor needed to dimerize the NOS-2 protein into active, NO-making NOS-2 enzyme.

  19. Constitutive suppressor of cytokine signaling 3 expression confers a growth advantage to a human melanoma cell line.

    PubMed

    Komyod, Waraporn; Böhm, Markus; Metze, Dieter; Heinrich, Peter C; Behrmann, Iris

    2007-03-01

    The growth of melanocytes and many early stage melanoma cells can be inhibited by cytokines, whereas late stage melanoma cells have often been reported to be "multi-cytokine-resistant." Here, we analyzed the melanoma cell line 1286, resistant towards the growth-inhibitory effects of interleukin 6 (IL-6), and oncostatin M (OSM), to better understand the mechanisms underlying cytokine resistance. Although the relevant receptors gp130 and OSMR are expressed at the cell surface of these cells, cytokine stimulation hardly led to the activation of Janus kinase 1 and signal transducer and activator of transcription (STAT)3 and STAT1. We found a high-level constitutive expression of suppressors of cytokine signaling 3 (SOCS3) that did not further increase after cytokine treatment. Importantly, upon suppression of SOCS3 by short interfering RNA, cells became susceptible towards OSM and IL-6: they showed an enhanced STAT3 phosphorylation and a dramatically increased STAT1 phosphorylation. Moreover, suppression of SOCS3 rendered 1286 cells sensitive to the antiproliferative action of IL-6 and OSM, but not of IFN-alpha. Interestingly, SOCS3-short interfering RNA treatment also increased the growth-inhibitory effect in cytokine-sensitive WM239 cells expressing SOCS3 in an inducible way. Thus, SOCS3 expression confers a growth advantage to these cell lines. Constitutive SOCS3 mRNA expression, although at lower levels than in 1286 cells, was found in nine additional human melanoma cell lines and in normal human melanocytes, although at the protein level, SOCS3 expression was marginal at best. However, in situ analysis of human melanoma specimens revealed SOCS3 immunoreactivity in 3 out of 10 samples, suggesting that in vivo SOCS3 may possibly play a role in IL-6 resistance in at least a fraction of tumors.

  20. Augmented gp130-mediated cytokine signalling accompanies human gastric cancer progression.

    PubMed

    Jackson, C B; Judd, L M; Menheniott, T R; Kronborg, I; Dow, C; Yeomans, N D; Boussioutas, A; Robb, L; Giraud, A S

    2007-10-01

    H. pylori infection accounts for most cases of gastric cancer, but the initiating events remain unclear. The principal H. pylori pathogenicity-associated CagA protein disrupts intracellular SHP-2 signalling pathways including those used by the IL-6 family cytokines, IL-6 and IL-11. Imbalanced IL-6 family cytokine signalling in the gp130(757FF) mouse model of gastric cancer arising from hyperactivation of oncogenic STAT3 after altered SHP-2 : ERK1/2 signalling produces dysplastic antral tumours preceded by gastritis and metaplasia. In a cohort of patient gastric biopsies with known H. pylori and CagA status, we investigated whether (i) STAT3 and ERK1/2 activation is altered in H. pylori-dependent gastritis; (ii) these profiles are more pronounced in CagA+ H. pylori infection; and (iii) the expression of pro-inflammatory cytokines that activate STAT3 and ERK 1/2 pathways is associated with progression to gastric cancer. IL-6, IL-11, and activated STAT3 and ERK1/2 were quantified in antral biopsies from gastritic stomach, metaplastic tissue, and resected gastric cancer tissues. We observed significantly increased STAT3 and ERK1/2 activation (p = 0.001) in H. pylori-dependent gastritis, which was further enhanced in the presence of CagA+ H. pylori strains. Of known gastric ligands that drive STAT3 activation, IL-6 expression was increased after H. pylori infection and both IL-6 and IL-11 were strongly up-regulated in the gastric cancer biopsies. This suggests a mechanism by which IL-11 drives STAT3 activation and proliferation during gastric cancer progression. We addressed this using an in vitro approach, demonstrating that recombinant human IL-11 activates STAT3 and concomitantly increases proliferation of MKN28 gastric epithelial cells. In summary, we show increased STAT3 and ERK1/2 activation in H. pylori-dependent gastritis that is likely driven in an IL-6-dependent fashion. IL-11 expression is associated with adenocarcinoma development, but not gastritic lesions

  1. 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

  2. Endotoxin or cytokines attenuate ozone-induced DNA synthesis in rat nasal transitional epithelium

    SciTech Connect

    Hotchkiss, J.A.; Harkema, J.R. )

    1992-06-01

    Pretreatment of rats with endotoxin (E), a potent inducer of tumor necrosis factor alpha (TNF), and interleukin 1 beta (IL 1), or a combination of TNF and IL1, has been shown to increase levels of lung antioxidant enzymes and protect against pulmonary toxicity associated with hyperoxia. Inhalation of ozone (O3) induces cell injury, followed by increased DNA synthesis, cell proliferation, and secretory cell metaplasia in rat nasal transitional epithelium (NTE). This study was designed to test the effects of E, TNF, and IL1 pretreatment on acute O3-induced NTE cell injury as measured by changes in NTE cell DNA synthesis. Rats were exposed to either 0.8 ppm O3 or air for 6 hr in whole-body inhalation chambers. Immediately before exposure, rats in each group were injected intraperitoneally (ip) with either saline alone or saline containing E, TNF, IL1, or both TNF and IL1. Eighteen hours postexposure, rats were injected ip with bromodeoxyuridine to label cells undergoing DNA synthesis and were euthanized 2 hr later. NTE was processed for light microscopy and immunochemically stained to identify cells that had incorporated BrdU into nuclear DNA. The number of BrdU-labeled NTE nuclei per millimeter of basal lamina was quantitated. There were no significant differences in the number of BrdU-labeled NTE nuclei in air-exposed rats that were injected with E, TNF, IL1, or TNF/IL1 compared with those in saline-injected, air-exposed controls. Rats that were injected with saline and exposed to O3 had approximately 10 times the number of BrdU-labeled NTE nuclei than saline-injected, air-exposed control rats. O3 exposure also induced a significant increase in labeled nuclei in rats that were pretreated with TNF alone. In contrast, pretreatment with E, IL1, or TNF/IL1 attenuated the O3-induced increase in NTE DNA synthesis.

  3. Bile acids and cytokines inhibit the human cholesterol 7 alpha-hydroxylase gene via the JNK/c-jun pathway in human liver cells.

    PubMed

    Li, Tiangang; Jahan, Asmeen; Chiang, John Y L

    2006-06-01

    Cholesterol 7 alpha-hydroxylase (CYP7A1) of the bile acid biosynthesis pathway is suppressed by bile acids and inflammatory cytokines. Bile acids are known to induce inflammatory cytokines to activate the mitogen-activated protein kinase/c-Jun N-terminal kinase (JNK) signaling pathway that inhibits CYP7A1 gene transcription. c-Jun has been postulated to mediate bile acid inhibition of CYP7A1. However, the c-Jun target involved in the regulation of CYP7A1 is unknown. Human primary hepatocytes and HepG2 cells were used as models to study chenodeoxycholic acid (CDCA) and interleukin-1 beta (IL-1 beta) regulation of human CYP7A1 gene expression via real-time polymerase chain reaction, reporter assays, co-immunoprecipitation and chromatin immunocipitation (ChIP) assays. IL-1 beta and CDCA reduced CYP7A1 but induced c-Jun messenger RNA expression in human primary hepatocytes. IL-1beta inhibited human CYP7A1 reporter activity via the HNF4 alpha binding site. A JNK-specific inhibitor blocked the inhibitory effect of IL-1 beta on HNF4 alpha expression and CYP7A1 reporter activity. c-Jun inhibited HNF4 alpha and PPARgamma coactivator-1 alpha (PGC-1 alpha) coactivation of CYP7A1 reporter activity, whereas a dominant negative c-Jun did not. Co-immunoprecipitation and ChIP assays revealed that IL-1 beta and CDCA reduced HNF4 alpha bound to the CYP7A1 chromatin, and that c-Jun interacted with HNF4 alpha and blocked HNF4 alpha recruitment of PGC-1 alpha to the CYP7A1 chromatin. In conclusion, IL-1 beta and CDCA inhibit HNF4 alpha but induce c-Jun, which in turn blocks HNF 4 alpha recruitment of PGC-1 alpha to the CYP7A1 chromatin and results in inhibition of CYP7A1 gene transcription. The JNK/c-Jun signaling pathway inhibits bile acid synthesis and protects hepatocytes against the toxic effect of inflammatory agents.

  4. 3D Human Motion Editing and Synthesis: A Survey

    PubMed Central

    Wang, Xin; Chen, Qiudi; Wang, Wanliang

    2014-01-01

    The ways to compute the kinematics and dynamic quantities of human bodies in motion have been studied in many biomedical papers. This paper presents a comprehensive survey of 3D human motion editing and synthesis techniques. Firstly, four types of methods for 3D human motion synthesis are introduced and compared. Secondly, motion capture data representation, motion editing, and motion synthesis are reviewed successively. Finally, future research directions are suggested. PMID:25045395

  5. Selective effects of tea extract and its phenolic compounds on human peripheral blood mononuclear cell cytokine secretions.

    PubMed

    Neyestani, Tirang R; Gharavi, A'azam; Kalayi, Ali

    2009-01-01

    The effects of black tea extract (BTE) and some of its pure phenolics on human peripheral blood mononuclear cell (PBMC) cytokine secretion were examined in vitro. The main steps of the study included chromatographic analysis of BTE and commercial green tea extract, Polyphenon 60, determination of the total antioxidant capacity of both the extracts and their phenolics, and finally evaluation of their effects on PBMCs. Four major peaks in the chromatogram of BTE belonged to caffeine, gallic acid, epigallocatechin and epigallocatechin gallate, and the latter showed the highest antioxidant capacity. While pure phenolics at the concentration of 20 mM did not significantly affect PBMC cytokine secretion, BTE and Polyphenon 60 suppressed interleukin-1 beta, tumor necrosis factor-alpha and interleukin-6. Interestingly, the secretion of interferon-gamma after 24 h was slightly, but significantly, boosted by the extracts. BTE has selective pro-inflammatory cytokine-suppressing effects on human PBMCs.

  6. Cytokine modulation (IL-6, IL-8, IL-10) by human breast milk lipids on intestinal epithelial cells (Caco-2).

    PubMed

    Barrera, Girolamo J; Sánchez, Gabriela

    2016-01-01

    Human breast milk is the best form of nourishment for infants during the first year of life. It is composed by a complex mixture of carbohydrates, proteins and fats. Breast milk provides nutrients and bioactive factors that themselves modulate maturation and development of the gastrointestinal tract. Many studies have shown that it provides protection against gastrointestinal tract inflammation. In this sense, this study aimed to evaluate the effect of human breast milk lipids on epithelial intestinal cells (Caco-2) cytokine regulation and the fatty acid transporter protein (FATP) involved in this process. Caco-2 cells were cultivated and stimulated with different concentration of human milk lipids from healthy human mothers (18-30-year-olds) or single commercial lipids for 48 h. We measured the concentrations and mRNA levels of IL-6, IL-8 and IL-10 cytokines by immunoassay (ELISA) and quantitative-PCR (qRT-PCR) technique, respectively. We observed a two to three times decrease in pro-inflammatory cytokine levels (p < 0.01) as well as an increase in anti-inflammatory IL-10 levels in cells stimulated with increasing concentrations of breast milk lipids. These results suggest that human breast milk lipids could have an important role on the cytokine modulation in the newborn bowel.

  7. Robotics-based Synthesis of Human Motion

    PubMed Central

    Khatib, O.; Demircan, E.; De Sapio, V.; Sentis, L.; Besier, T.; Delp, S.

    2009-01-01

    The synthesis of human motion is a complex procedure that involves accurate reconstruction of movement sequences, modeling of musculoskeletal kinematics, dynamics and actuation, and characterization of reliable performance criteria. Many of these processes have much in common with the problems found in robotics research. Task-based methods used in robotics may be leveraged to provide novel musculoskeletal modeling methods and physiologically accurate performance predictions. In this paper, we present (i) a new method for the real-time reconstruction of human motion trajectories using direct marker tracking, (ii) a task-driven muscular effort minimization criterion and (iii) new human performance metrics for dynamic characterization of athletic skills. Dynamic motion reconstruction is achieved through the control of a simulated human model to follow the captured marker trajectories in real-time. The operational space control and real-time simulation provide human dynamics at any configuration of the performance. A new criteria of muscular effort minimization has been introduced to analyze human static postures. Extensive motion capture experiments were conducted to validate the new minimization criterion. Finally, new human performance metrics were introduced to study in details an athletic skill. These metrics include the effort expenditure and the feasible set of operational space accelerations during the performance of the skill. The dynamic characterization takes into account skeletal kinematics as well as muscle routing kinematics and force generating capacities. The developments draw upon an advanced musculoskeletal modeling platform and a task-oriented framework for the effective integration of biomechanics and robotics methods. PMID:19665552

  8. Robotics-based synthesis of human motion.

    PubMed

    Khatib, O; Demircan, E; De Sapio, V; Sentis, L; Besier, T; Delp, S

    2009-01-01

    The synthesis of human motion is a complex procedure that involves accurate reconstruction of movement sequences, modeling of musculoskeletal kinematics, dynamics and actuation, and characterization of reliable performance criteria. Many of these processes have much in common with the problems found in robotics research. Task-based methods used in robotics may be leveraged to provide novel musculoskeletal modeling methods and physiologically accurate performance predictions. In this paper, we present (i) a new method for the real-time reconstruction of human motion trajectories using direct marker tracking, (ii) a task-driven muscular effort minimization criterion and (iii) new human performance metrics for dynamic characterization of athletic skills. Dynamic motion reconstruction is achieved through the control of a simulated human model to follow the captured marker trajectories in real-time. The operational space control and real-time simulation provide human dynamics at any configuration of the performance. A new criteria of muscular effort minimization has been introduced to analyze human static postures. Extensive motion capture experiments were conducted to validate the new minimization criterion. Finally, new human performance metrics were introduced to study in details an athletic skill. These metrics include the effort expenditure and the feasible set of operational space accelerations during the performance of the skill. The dynamic characterization takes into account skeletal kinematics as well as muscle routing kinematics and force generating capacities. The developments draw upon an advanced musculoskeletal modeling platform and a task-oriented framework for the effective integration of biomechanics and robotics methods.

  9. Human sebum extract induces barrier disruption and cytokine expression in murine epidermis.

    PubMed

    Guo, Jiun-Wen; Lin, Tzu-Kai; Wu, Chin-Han; Wei, Kai-Che; Lan, Cheng-Che E; Peng, Amy Chia-Ying; Tsai, Jui-Chen; Sheu, Hamm-Ming

    2015-04-01

    Previous studies have shown that human sebum may play a role in barrier function but with much debate. To elucidate the effects of human sebum on skin barrier function. We used hairless mouse skin to study the functional and morphological alternation of epidermis after the application of human sebum. The results showed a significant increase in transepidermal water loss and erythema value, and a decrease in skin hydration, accompanied by epidermal hyperplasia with parakeratosis following sebum application. Nile red staining together with electron microscopic examination confirmed the underlying mechanisms for sebum-induced barrier disruption are related directly to the interaction of sebum with the intracellular lipid lamellae of the SC, thereby leading to the increase in the fluidity of SC intracellular lipids as demonstrated by ATR-FTIR measurement. An inflammatory reaction characterized by an enhanced cytokine cascade, including up-regulation of TNF-α, IL-1α and IL-6, was also observed. On the other hand, there were insignificant expression of thymic stromal lymphopoietin and unchanged serum levels of IgE, suggesting non-immunogenic stimulation by sebum treatment. It may be concluded that inflammation induced by excess amount of sebum is more likely an irritant contact dermatitis rather than an allergic one. Moreover, these findings implicated possible relationships between sebum, irritant contact dermatitis, and seborrheic dermatitis. Copyright © 2015 Japanese Society for Investigative Dermatology. Published by Elsevier Ireland Ltd. All rights reserved.

  10. Neisseria gonorrhoeae survives within and modulates apoptosis and inflammatory cytokine production of human macrophages.

    PubMed

    Château, Alice; Seifert, H Steven

    2016-04-01

    The human-adapted organism Neisseria gonorrhoeae is the causative agent of gonorrhoea, a sexually transmitted infection. It readily colonizes the genital, rectal and nasalpharyngeal mucosa during infection. While it is well established that N. gonorrhoeae recruits and modulates the functions of polymorphonuclear leukocytes during infection, how N. gonorrhoeae interacts with macrophages present in infected tissue is not fully defined. We studied the interactions of N. gonorrhoeae with two human monocytic cell lines, THP-1 and U937, and primary monocytes, all differentiated into macrophages. Most engulfed bacteria were killed in the phagolysosome, but a subset of bacteria was able to survive and replicate inside the macrophages suggesting that those cells may be an unexplored cellular reservoir for N. gonorrhoeae during infection. N. gonorrhoeae was able to modulate macrophage apoptosis: N. gonorrhoeae induced apoptosis in THP-1 cells whereas it inhibited induced apoptosis in U937 cells and primary human macrophages. Furthermore, N. gonorrhoeae induced expression of inflammatory cytokines in macrophages, suggesting a role for macrophages in recruiting polymorphonuclear leukocytes to the site of infection. These results indicate macrophages may serve as a significant replicative niche for N. gonorrhoeae and play an important role in gonorrheal pathogenesis.

  11. 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.

  12. Neisseria gonorrhoeae survives within and modulates apoptosis and inflammatory cytokine production of human macrophages

    PubMed Central

    Château, Alice; Seifert, H. Steven

    2017-01-01

    The human-adapted organism Neisseria gonorrhoeae is the causative agent of gonorrhea a sexually transmitted infection. It readily colonizes the genital, rectal, and nasalpharyngal mucosa during infection. While it is well-established that N. gonorrhoeae recruits and modulates the functions of polymorphonuclear leukocytes (PMNs) during infection, how N. gonorrhoeae interacts with macrophages present in infected tissue is not fully defined. We studied the interactions of N. gonorrhoeae with two human monocytic cell lines, THP-1 and U937, and primary monocytes, all differentiated into macrophages. Most engulfed bacteria were killed in the phagolysosome, but a subset of bacteria were able to survive and replicate inside the macrophages suggesting that those cells may be an unexplored cellular reservoir for N. gonorrhoeae during infection. N. gonorrhoeae was able to modulate macrophage apoptosis, N. gonorrhoeae induced apoptosis in THP-1 cells whereas it inhibited induced apoptosis in U937 cells and primary human macrophages. Furthermore, N. gonorrhoeae induced expression of inflammatory cytokines in macrophages, suggesting a role for macrophages in recruiting PMNs to the site of infection. These results indicate macrophages may serve as a significant replicative niche for N. gonorrhoeae and play an important role in gonorrheal pathogenesis. PMID:26426083

  13. A new method to measure air-borne pyrogens based on human whole blood cytokine response.

    PubMed

    Kindinger, Ilona; Daneshian, Mardas; Baur, Hans; Gabrio, Thomas; Hofmann, Andreas; Fennrich, Stefan; von Aulock, Sonja; Hartung, Thomas

    2005-03-01

    Air-borne microorganisms, as well as their fragments and components, are increasingly recognized to be associated with pulmonary diseases, e.g. organic dust toxic syndrome, humidifier lung, building-related illness, "Monday sickness." We have previously described and validated a new method for the detection of pyrogenic (fever-inducing) microbial contaminations in injectable drugs, based on the inflammatory reaction of human blood to pyrogens. We have now adapted this test to evaluate the total inflammatory capacity of air samples. Air was drawn onto PTFE membrane filters, which were incubated with human whole blood from healthy volunteers inside the collection device. Cytokine release was measured by ELISA. The test detects endotoxins and non-endotoxins, such as fungal spores, Gram-positive bacteria and their lipoteichoic acid moiety and pyrogenic dust particles with high sensitivity, thus reflecting the total inflammatory capacity of a sample. When air from different surroundings such as working environments and animal housing was assayed, the method yielded reproducible data which correlated with other parameters of microbial burden tested. We further developed a standard material for quantification and showed that this assay can be performed with cryopreserved as well as fresh blood. The method offers a test to measure the integral inflammatory capacity of air-borne microbial contaminations relevant to humans. It could thus be employed to assess air quality in different living and work environments.

  14. Cytokine profiles, signalling pathways and effects of fluticasone propionate in respiratory syncytial virus-infected human foetal lung fibroblasts.

    PubMed

    Seki, Erina; Yoshizumi, Masakazu; Tanaka, Ryota; Ryo, Akihide; Ishioka, Taisei; Tsukagoshi, Hiroyuki; Kozawa, Kunihisa; Okayama, Yoshimichi; Okabe-Kado, Junko; Goya, Tomoyuki; Kimura, Hirokazu

    2013-04-01

    To examine cytokine production in response to RSV infection, we assessed the levels of 29 cytokines released from RSV-infected human foetal lung fibroblasts. We also examined the relationships between the effects of fluticasone propionate and various signalling pathways in the cells. Twenty-four hours after infection (1MOI), RSV-infected cells released cytokines, for example proinflammatory cytokines (IL-1β, IL-6 and TNF-α), anti-inflammatory (IL-1ra), Th1 (IFN-γ, IFN-λ1a, IL-2 and IL-12), Th2 (IL-4, IL-5, IL-10 and IL-13), granulopoiesis-inducing (G-CSF and GM-CSF), eosinophil recruitment-inducing (eotaxin and RANTES) and neutrophil recruitment-inducing cytokines (IL-8, IP-10, MCP-1 and MIP-1α). Aberrant release of most was significantly suppressed by fluticasone propionate. Twelve hours after RSV infection, increased phosphorylation of Akt, p38 MAPK, ERK1/2 and IκB-α was noted. Fluticasone propionate suppressed the phosphorylation of Akt, p38 MAPK, and ERK1/2, but not IκB-α, in virus-infected cells. TLR-4 expression was unchanged in control and RSV-infected cells, and TLR-3 and RIG-I expression was not detected. The results indicate that RSV infection induces aberrant production and release of certain cytokines through these signalling pathways in human lung fibroblasts. Overproduction and imbalance of these cytokines may be associated with the pathophysiology of RSV-induced excessive and allergic inflammation. © 2013 International Federation for Cell Biology.

  15. Biological and Technical Variables Affecting Immunoassay Recovery of Cytokines from Human Serum and Simulated Vaginal Fluid: A Multicenter Study

    PubMed Central

    2008-01-01

    The increase of proinflammatory cytokines in vaginal secretions may serve as a surrogate marker of unwanted inflammatory reaction to microbicide products topically applied for the prevention of sexually transmitted diseases, including HIV-1. Interleukin (IL)-1β and IL-6 have been proposed as indicators of inflammation and increased risk of HIV-1 transmission; however, the lack of information regarding detection platforms optimal for vaginal fluids and interlaboratory variation limit their use for microbicide evaluation and other clinical applications. This study examines fluid matrix variants relevant to vaginal sampling techniques and proposes a model for interlaboratory comparisons across current cytokine detection technologies. IL-1β and IL-6 standards were measured by 12 laboratories in four countries, using 14 immunoassays and four detection platforms based on absorbance, chemiluminescence, electrochemiluminescence, and fluorescence. International reference preparations of cytokines with defined biological activity were spiked into (1) a defined medium simulating the composition of human vaginal fluid at pH 4.5 and 7.2, (2) physiologic salt solutions (phosphate-buffered saline and saline) commonly used for vaginal lavage sampling in clinical studies of cytokines, and (3) human blood serum. Assays were assessed for reproducibility, linearity, accuracy, and significantly detectable fold difference in cytokine level. Factors with significant impact on cytokine recovery were determined by Kruskal−Wallis analysis of variance with Dunn’s multiple comparison test and multiple regression models. All assays showed acceptable intra-assay reproducibility; however, most were associated with significant interlaboratory variation. The smallest reliably detectable cytokine differences (P < 0.05) derived from pooled interlaboratory data varied from 1.5- to 26-fold depending on assay, cytokine, and matrix type. IL-6 but not IL-1β determinations were lower in both saline

  16. Pro-inflammatory cytokine TNF-α is a key inhibitory factor for lactose synthesis pathway in lactating mammary epithelial cells.

    PubMed

    Kobayashi, Ken; Kuki, Chinatsu; Oyama, Shoko; Kumura, Haruto

    2016-01-15

    Lactose is a milk-specific carbohydrate synthesized by mammary epithelial cells (MECs) in mammary glands during lactation. Lactose synthesis is downregulated under conditions causing inflammation such as mastitis, in which MECs are exposed to high concentrations of inflammatory cytokines. In this study, we investigated whether inflammatory cytokines (TNF-α, IL-1β, and IL-6) directly influence the lactose synthesis pathway by using two types of murine MEC culture models: the monolayer culture of MECs to induce lactogenesis; and the three-dimensional culture of MECs surrounded by Matrigel to induce reconstitution of the alveolar structure in vitro. TNF-α caused severe down-regulation of lactose synthesis-related genes concurrently with the degradation of glucose transporter 1 (GLUT1) from the basolateral membranes in MECs. IL-1β also caused degradation of GLUT1 along with a decrease in the expression level of β-1,4-galactosylransferase 3. IL-6 caused both up-regulation and down-regulation of the expression levels of lactose synthesis-related genes in MECs. These results indicate that TNF-α, IL-1β, and IL-6 have different effects on the lactose synthesis pathway in MECs. Furthermore, TNF-α triggered activation of NFκB and inactivation of STAT5, suggesting that NFκB and STAT5 signaling pathways are involved in the multiple adverse effects of TNF-α on the lactose synthesis pathway.

  17. Berberine regulates proliferation, collagen synthesis and cytokine secretion of cardiac fibroblasts via AMPK-mTOR-p70S6K signaling pathway.

    PubMed

    Ai, Fen; Chen, Manhua; Yu, Bo; Yang, Yang; Xu, Guizhong; Gui, Feng; Liu, Zhenxing; Bai, Xiangyan; Chen, Zhen

    2015-01-01

    The traditional Chinese medicinal berberine has long been used to treat cardiovascular diseases; however, the mechanism underlying its effects remains unclear. Here, this study would to investigate the effects of berberine on proliferation, collagen synthesis and cytokine secretion of cardiac fibroblasts. We assessed proliferation, collagen synthesis and cytokine secretion in cardiac fibroblasts subjected to angiotensin II (Ang II) subsequent to the consumption of berberine or a control treatment. And then we detected the role of AMPK/mTOR signaling pathway in berberine treatment of cardiac fibroblasts. In the present study, the cellular behaviors of cardiac fibroblasts induced by Ang II were significantly activated including proliferation, transformation into myofibroblasts and collagen synthesis. Additionally, the ability of cytokine secretion was enhanced obviously. It was demonstrated that treatment of cardiac fibroblasts with berberine resulted in deceased proliferation, and attenuated fibroblast α-smooth muscle actin expression and collagen synthesis. And the protein secretion of TGFβ1 was inhibited; however, the protein secretion of IL-10 was increased in cardiac fibroblasts with berberine treatment. Mechanistically, the phosphorylation level of AMPK was increased; and the phosphorylation levels of mTOR and p70S6K were decreased in berberine treatment group. These results illustrated that the protective effects of berberine on cellular behaviors of cardiac fibroblasts were at least in part due to activate AMPK signaling pathway and downregulate mTOR/p70S6K signaling pathway. Berberine might become a new strategy for treating cardiac fibrosis in the future.

  18. Expression of adhesion molecules and chemotactic cytokines in cultured human mesothelial cells.

    PubMed

    Jonjić, N; Peri, G; Bernasconi, S; Sciacca, F L; Colotta, F; Pelicci, P; Lanfrancone, L; Mantovani, A

    1992-10-01

    The mesothelium is a flat epithelial lining of serous cavities that could gate the traffic of molecules and cells between the circulation and these body compartments. The present study was designed to elucidate the capacity of mesothelial cells to express adhesion molecules and chemoattractant cytokines, two fundamental mechanisms of regulation of leukocyte recruitment. Cultured human mesothelial cells express appreciable levels of intercellular adhesion molecule 1 (ICAM-1) and vascular cell adhesion molecule 1 (VCAM-1), and these were increased by in vitro exposure to tumor necrosis factor (TNF), interferon gamma (IFN-gamma), or TNF and IFN-gamma. Interleukin 1 (IL-1) was a less consistent stimulus for adhesion molecule expression in vitro. Unlike endothelial cells, used as a reference cell population, resting or stimulated mesothelial cells did not express E-selectin and ICAM-2, as assessed by flow cytometry. Analysis of VCAM-1 mRNA by reverse transcriptase and polymerase chain reaction using appropriate primers revealed that mesothelial cells expressed both the seven- and the six-Ig domain transcripts, with predominance of the longer species. Monocytes bound appreciably to "resting" and, to a greater extent, to stimulated mesothelial cells. Monocytes exposed to IFN-gamma and lipopolysaccharide, used as prototypic activation signals, showed increased capacity to bind mesothelial cells. Anti-CD18 monoclonal antibody significantly inhibited binding of monocytes to mesothelial cells, and this blocking effect was amplified by anti-very late antigen 4. Mesothelial cells were able to express the chemotactic cytokines IL-8 and monocyte chemotactic protein 1 at the mRNA and protein levels. These results indicate that mesothelial cells can express a set of adhesion molecules (ICAM-1 and VCAM-1) overlapping with, but distinct from, that expressed in vascular endothelium (ICAM-1, ICAM-2, VCAM-1, E-selectin), and that these are functionally relevant for interacting with

  19. TREK-1 Regulates Cytokine Secretion from Cultured Human Alveolar Epithelial Cells Independently of Cytoskeletal Rearrangements

    PubMed Central

    Schwingshackl, Andreas; Roan, Esra; Teng, Bin; Waters, Christopher M.

    2015-01-01

    Background TREK-1 deficient alveolar epithelial cells (AECs) secrete less IL-6, more MCP-1, and contain less F-actin. Whether these alterations in cytokine secretion and F-actin content are related remains unknown. We now hypothesized that cytokine secretion from TREK-1-deficient AECs was regulated by cytoskeletal rearrangements. Methods We determined F-actin and α-tubulin contents of control, TREK-1-deficient and TREK-1-overexpressing human A549 cells by confocal microscopy and western blotting, and measured IL-6 and MCP-1 levels using real-time PCR and ELISA. Results Cytochalasin D decreased the F-actin content of control cells. Jasplakinolide increased the F-actin content of TREK-1 deficient cells, similar to the effect of TREK-1 overexpression in control cells. Treatment of control and TREK-1 deficient cells with TNF-α, a strong stimulus for IL-6 and MCP-1 secretion, had no effect on F-actin structures. The combination of TNF-α+cytochalasin D or TNF-α+jasplakinolide had no additional effect on the F-actin content or architecture when compared to cytochalasin D or jasplakinolide alone. Although TREK-1 deficient AECs contained less F-actin at baseline, quantified biochemically, they contained more α-tubulin. Exposure to nocodazole disrupted α-tubulin filaments in control and TREK-1 deficient cells, but left the overall amount of α-tubulin unchanged. Although TNF-α had no effect on the F-actin or α-tubulin contents, it increased IL-6 and MCP-1 production and secretion from control and TREK-1 deficient cells. IL-6 and MCP-1 secretions from control and TREK-1 deficient cells after TNF-α+jasplakinolide or TNF-α+nocodazole treatment was similar to the effect of TNF-α alone. Interestingly, cytochalasin D decreased TNF-α-induced IL-6 but not MCP-1 secretion from control but not TREK-1 deficient cells. Conclusion Although cytochalasin D, jasplakinolide and nocodazole altered the F-actin and α-tubulin structures of control and TREK-1 deficient AEC, the

  20. Cytokine and Lipid Mediator Regulation of Group 2 Innate Lymphoid Cells (ILC2s) in Human Allergic Airway Disease.

    PubMed

    Cavagnero, Kellen; Doherty, Taylor A

    2017-08-01

    The recent discovery of group 2 innate lymphoid cells (ILC2s) has caused a paradigm shift in the understanding of allergic airway disease pathogenesis. Prior to the discovery of ILC2s, Th2 cells were largely thought to be the primary source of type 2 cytokines; however, activated ILC2s have since been shown to contribute significantly, and in some cases, dominantly to type 2 cytokine production. Since the discovery of ILC2s in 2010, many mediators have been shown to regulate their effector functions. Initial studies identified the epithelial derived cytokines IL-25, IL-33, and TSLP as activators of ILC2s, and recent studies have identified many additional cytokine and lipid mediators that are involved in ILC2 regulation. ILC2s and their mediators represent novel therapeutic targets for allergic airway diseases and intensive investigation is underway to better understand ILC2 biology and upstream and downstream pathways that lead to ILC2-driven airway pathology. In this review, we will focus on the cytokine and lipid mediators that regulate ILC2s in human allergic airway disease, as well as highlight newly discovered mediators of mouse ILC2s that may eventually translate to humans.

  1. Effects of submicrometer particle compositions on cytokine production and lipid peroxidation of human bronchial epithelial cells.

    PubMed Central

    Huang, Song-Lih; Hsu, Miao-Kan; Chan, Chang-Chuan

    2003-01-01

    To identify the size and components related to toxicity of ambient particles, we used a trichotomous impactor to collect 17 sets of particles in three size ranges--submicrometer (diameters < 1 microm; PM1.0, fine (diameters between 1 and 2.5 microm; PM1.0-2.5, and coarse (diameters between 2.5 and 10 microm; PM2.5-10--at stations monitoring background, urban, traffic, and industrial air in Taiwan. Elemental contents, carbon contents, soluble ions, and endotoxin content of particles were determined by X-ray fluorescence spectrometry, thermal analysis, ion chromatography, and the Limulus amebocyte lysate assay, respectively. Human bronchial epithelial BEAS-2B cells were exposed to particle extracts at 100 micro g/mL for 8 hr, and interleukin-8 (IL-8) concentrations in the medium and lipid peroxidation products were measured. Particle-induced tumor necrosis factor-alpha (TNF-alpha) production by mouse macrophage RAW 264.7 cells was also measured. PM1.0 stimulation resulted in significantly higher IL-8 production and lipid peroxidation than PM2.5-10, whereas the responses elicited by PM1.0-2.5 were not significantly higher than blank filters. Untreated and polymyxin B-pretreated PM1.0 also stimulated more TNF-alpha production by RAW 264.7 cells than PM2.5-10 and PM1.0-2.5. Cytokine production was significantly associated with metal contents of PM1.0: IL-8 correlated with Cr and Mn, and TNF-alpha correlated with Fe and Cr. Lipid peroxidation in BEAS-2B cells correlated with elemental and organic carbon contents. Our study found that size and composition of ambient particles were both important factors in inducing cytokine production and lipid peroxidation. PMID:12676602

  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. Viability, Apoptosis, Proliferation, Activation, and Cytokine Secretion of Human Keratoconus Keratocytes after Cross-Linking

    PubMed Central

    Stachon, Tanja; Wang, Jiong; Seitz, Berthold; Szentmáry, Nóra

    2015-01-01

    Purpose. The purpose of this study was to determine the impact of cross-linking (CXL) on viability, apoptosis, proliferation, activation, and cytokine secretion of human keratoconus (KC) keratocytes, in vitro. Methods. Primary KC keratocytes were cultured in DMEM/Ham's F12 medium supplemented with 10% FCS and underwent UVA illumination (370 nm, 2 J/cm2) during exposure to 0.1% riboflavin and 20% Dextran in PBS. Twenty-four hours after CXL, viability was assessed using Alamar blue assay; apoptosis using APO-DIRECT Kit; proliferation using ELISA-BrdU kit; and CD34 and alpha-smooth muscle actin (α-SMA) expression using flow cytometry. Five and 24 hours after CXL, FGFb, HGF, TGFβ1, VEGF, KGF, IL-1β, IL-6, and IL-8 secretion was measured using enzyme-linked-immunoabsorbent assay (ELISA). Results. Following CXL, cell viability and proliferation decreased (P < 0.05; P = 0.009), the percentage of apoptotic keratocytes increased (P < 0.05) significantly, and CD34 and α-SMA expression remained unchanged (P > 0.06). Five hours after CXL, FGFb secretion increased significantly (P = 0.037); however no other cytokine secretion differed significantly from controls after 5 or 24 hours (P > 0.12). Conclusions. Cross-linking decreases viability, triggers apoptosis, and inhibits proliferation, without an impact on multipotent hematopoietic stem cell transformation and myofibroblastic transformation of KC keratocytes. CXL triggers FGFb secretion of KC keratocytes transiently (5 hours), normalizing after 24 hours. PMID:25699261

  4. Effects of proinflammatory cytokines on the claudin-19 rich tight junctions of human retinal pigment epithelium.

    PubMed

    Peng, Shaomin; Gan, Geliang; Rao, Veena S; Adelman, Ron A; Rizzolo, Lawrence J

    2012-07-27

    Chronic, subclinical inflammation contributes to the pathogenesis of several ocular diseases, including age-related macular degeneration. Proinflammatory cytokines affect tight junctions in epithelia that lack claudin-19, but in the retinal pigment epithelium claudin-19 predominates. We examined the effects of cytokines on the tight junctions of human fetal RPE (hfRPE). hfRPE was incubated with interleukin 1-beta (IL-1β), interferon-gamma (IFNγ), or tumor necrosis factor-alpha (TNFα), alone or in combination. Permeability and selectivity of the tight junctions were assessed using nonionic tracers and electrophysiology. Claudins, occludin, and ZO-1 were examined using PCR, immunoblotting, and confocal immunofluorescence microscopy. Only TNFα consistently reduced transepithelial electrical resistance (TER) >80%. A serum-free medium revealed two effects of TNFα: (1) decreased TER was observed only when TNFα was added to the apical side of the monolayer, and (2) expression of TNFα receptors and inhibitors of apoptosis were induced from either side of the monolayer. In untreated cultures, tight junctions were slightly cation selective, and this was affected minimally by TNFα. The results were unexplained by effects on claudin-2, claudin-3, claudin-19, occludin, and ZO-1, but changes in the morphology of the junctions and actin cytoskeleton may have a role. Claudin-19-rich tight junctions have low permeability for ionic and nonionic solutes, and are slightly cation-selective. Claudin-19 is not a direct target of TNFα. TNFα may protect RPE from apoptosis, but makes the monolayer leaky when it is presented to the apical side of the monolayer. Unlike other epithelia, IFNγ failed to augment the effect of TNFα on tight junctions.

  5. Effects of Proinflammatory Cytokines on the Claudin-19 Rich Tight Junctions of Human Retinal Pigment Epithelium

    PubMed Central

    Peng, Shaomin; Gan, Geliang; Rao, Veena S.; Adelman, Ron A.; Rizzolo, Lawrence J.

    2012-01-01

    Purpose. Chronic, subclinical inflammation contributes to the pathogenesis of several ocular diseases, including age-related macular degeneration. Proinflammatory cytokines affect tight junctions in epithelia that lack claudin-19, but in the retinal pigment epithelium claudin-19 predominates. We examined the effects of cytokines on the tight junctions of human fetal RPE (hfRPE). Methods. hfRPE was incubated with interleukin 1-beta (IL-1β), interferon-gamma (IFNγ), or tumor necrosis factor-alpha (TNFα), alone or in combination. Permeability and selectivity of the tight junctions were assessed using nonionic tracers and electrophysiology. Claudins, occludin, and ZO-1 were examined using PCR, immunoblotting, and confocal immunofluorescence microscopy. Results. Only TNFα consistently reduced transepithelial electrical resistance (TER) >80%. A serum-free medium revealed two effects of TNFα: (1) decreased TER was observed only when TNFα was added to the apical side of the monolayer, and (2) expression of TNFα receptors and inhibitors of apoptosis were induced from either side of the monolayer. In untreated cultures, tight junctions were slightly cation selective, and this was affected minimally by TNFα. The results were unexplained by effects on claudin-2, claudin-3, claudin-19, occludin, and ZO-1, but changes in the morphology of the junctions and actin cytoskeleton may have a role. Conclusions. Claudin-19–rich tight junctions have low permeability for ionic and nonionic solutes, and are slightly cation-selective. Claudin-19 is not a direct target of TNFα. TNFα may protect RPE from apoptosis, but makes the monolayer leaky when it is presented to the apical side of the monolayer. Unlike other epithelia, IFNγ failed to augment the effect of TNFα on tight junctions. PMID:22761260

  6. Airborne allergens induce protease activated receptor-2-mediated production of inflammatory cytokines in human gingival epithelium.

    PubMed

    Son, Ga-Yeon; Son, Aran; Yang, Yu-Mi; Park, Wonse; Chang, Inik; Lee, Jae-Ho; Shin, Dong Min

    2016-01-01

    In reaching the airways inhaled allergens pass through and contact with the oral mucosa. Although they are often responsible for initiating asthmatic attacks, it is unknown whether airborne allergens can also trigger chronic inflammation of gingival epithelial cells leading to chronic periodontitis. In this study, we investigated the inflammatory responses of human gingival epithelial cells (HGECs) to airborne allergens, particularly German cockroach extract (GCE) with a focus on calcium signaling. HGECs isolated from healthy donors were stimulated with GCE. Intracellular Ca(2+) concentration ([Ca(2+)]i) was measured with Fura-2-acetoxymethyl ester (Fura-2/AM) staining. Expression of inflammatory cytokines interleukin (IL)-8, IL-1β, IL-6, and NOD-like receptor family, pyridine domain-containing (NLRP) 3 was analyzed using reverse transcription-polymerase chain reaction (RT-PCR). GCE promoted increase in the [Ca(2+)]i in a dose-dependent manner. Depletion of endoplasmic reticulum (ER) Ca(2+) by the ER Ca(2+) ATPase inhibitor thapsigargin (Tg) but not the depletion of extracellular Ca(2+) abolished the GCE-induced increase in [Ca(2+)]i. Treatment of phospholipase C (PLC) inhibitor (U73122) or 1,4,5-trisinositolphosphate (IP3) receptor inhibitor (2-APB) also prevented GCE-induced increase in [Ca(2+)]i. Protease activated receptor (PAR)-2 activation mainly mediated the GCE-induced increase in [Ca(2+)]i and enhanced the expression of IL-8, NLRP3, IL-1β, and IL-6 in HGECs. GCE activates PAR-2, which can induce PLC/IP3-dependent Ca(2+) signaling pathway, ultimately triggering inflammation via the production of pro-inflammatory cytokines such as IL-1β, IL-6, IL-8, and NLRP 3 in HGECs. Copyright © 2015 Elsevier Ltd. All rights reserved.

  7. Manumycin A downregulates release of proinflammatory cytokines from TNF alpha stimulated human monocytes.

    PubMed

    Cecrdlova, Eva; Petrickova, Katerina; Kolesar, Libor; Petricek, Miroslav; Sekerkova, Alena; Svachova, Veronika; Striz, Ilja

    2016-01-01

    Macrolide antibiotics such as azithromycin or clarithromycin are known to have potent anti-inflammatory and immunomodulatory effects but these properties cannot be widely used due to a risk of bacterial resistance. We studied another polyketide antibiotic, structurally related manumycin A known as a streptomycete derived farnesyltransferase inhibitor with limited antibacterial effects, with respect to its potential regulation of mRNA expression of several genes associated with proinflammatory responses. Downregulation of mRNA for IL-6, TLR-8, IL-1 beta and IL-10 was found in THP-1 cells after 4h stimulation with TNF alpha in the presence of manumycin A and downregulated TLR-8 and EGR-1 genes were observed after 8h. Among the genes upregulated in response to manumycin were HMOX-1, TNFRSF10A, IL-1R1, TICAM2, NLRP12 after 4h and only IL-1R1 after 8h. Furthermore, manumycin A was found to inhibit IL-1beta, IL-6, and IL-8 production in TNF alpha stimulated THP-1 cells and peripheral blood monocytes in a dose dependent manner (0.25-1 μM of manumycin A) without affecting cell viability. Cell viability of blood monocytes decreased by about 30% at manumycin A doses of 2-5 μM. Manumycin A also inhibited IL-18 release from THP-1 cells, while in cultures of blood monocytes, this cytokine was not detectable. That manumycin A mediated downregulation of proinflammatory genes in human monocytes confirmed by a measurement of cytokine levels in culture supernatants, together with a very limited effect on cell viability, might suggest potential anti-inflammatory properties of this polyketide antibiotic.

  8. ADHESION AND POLLUTION PARTICLE-INDUCED OXIDANT GENERATION IS NEITHER NECESSARY NOR SUFFICIENT FOR CYTOKINE INDUCTION IN HUMAN ALVEOLAR MACROPHAGES

    EPA Science Inventory

    Adhesion of human monocytes (MOs) results in the rapid transcriptional activation of cytokine genes that are dependent on nuclear factor (NF)-kappaB. Several pathways leading to activation of NF-kappaB have been described, including those involving reactive oxygen intermediates (...

  9. ADHESION AND POLLUTION PARTICLE-INDUCED OXIDANT GENERATION IS NEITHER NECESSARY NOR SUFFICIENT FOR CYTOKINE INDUCTION IN HUMAN ALVEOLAR MACROPHAGES

    EPA Science Inventory

    Adhesion of human monocytes (MOs) results in the rapid transcriptional activation of cytokine genes that are dependent on nuclear factor (NF)-kappaB. Several pathways leading to activation of NF-kappaB have been described, including those involving reactive oxygen intermediates (...

  10. Human Cytomegalovirus Encoded Homologs of Cytokines, Chemokines and their Receptors: Roles in Immunomodulation

    PubMed Central

    McSharry, Brian P.; Avdic, Selmir; Slobedman, Barry

    2012-01-01

    Human cytomegalovirus (HCMV), the largest human herpesvirus, infects a majority of the world’s population. Like all herpesviruses, following primary productive infection, HCMV establishes a life-long latent infection, from which it can reactivate years later to produce new, infectious virus. Despite the presence of a massive and sustained anti-HCMV immune response, productively infected individuals can shed virus for extended periods of time, and once latent infection is established, it is never cleared from the host. It has been proposed that HCMV must therefore encode functions which help to evade immune mediated clearance during productive virus replication and latency. Molecular mimicry is a strategy used by many viruses to subvert and regulate anti-viral immunity and HCMV has hijacked/developed a range of functions that imitate host encoded immunomodulatory proteins. This review will focus on the HCMV encoded homologs of cellular cytokines/chemokines and their receptors, with an emphasis on how these virus encoded homologs may facilitate viral evasion of immune clearance. PMID:23202490

  11. Cellular Mechanics of Primary Human Cervical Fibroblasts: Influence of Progesterone and a Pro-inflammatory Cytokine.

    PubMed

    Shukla, Vasudha; Barnhouse, Victoria; Ackerman, William E; Summerfield, Taryn L; Powell, Heather M; Leight, Jennifer L; Kniss, Douglas A; Ghadiali, Samir N

    2017-09-22

    The leading cause of neonatal mortality, pre-term birth, is often caused by pre-mature ripening/opening of the uterine cervix. Although cervical fibroblasts play an important role in modulating the cervix's extracellular matrix (ECM) and mechanical properties, it is not known how hormones, i.e., progesterone, and pro-inflammatory insults alter fibroblast mechanics, fibroblast-ECM interactions and the resulting changes in tissue mechanics. Here we investigate how progesterone and a pro-inflammatory cytokine, IL-1β, alter the biomechanical properties of human cervical fibroblasts and the fibroblast-ECM interactions that govern tissue-scale mechanics. Primary human fibroblasts were isolated from non-pregnant cervix and treated with estrogen/progesterone, IL-1β or both. The resulting changes in ECM gene expression, matrix remodeling, traction force generation, cell-ECM adhesion and tissue contractility were monitored. Results indicate that IL-1β induces a significant reduction in traction force and ECM adhesion independent of pre-treatment with progesterone. These cell level effects altered tissue-scale mechanics where IL-1β inhibited the contraction of a collagen gel over 6 days. Interestingly, progesterone treatment alone did not modulate traction forces or gel contraction but did result in a dramatic increase in cell-ECM adhesion. Therefore, the protective effect of progesterone may be due to altered adhesion dynamics as opposed to altered ECM remodeling.

  12. 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

  13. Natural innate cytokine response to immunomodulators and adjuvants in human precision-cut lung slices

    SciTech Connect

    Switalla, S.; Lauenstein, L.; Prenzler, F.; Knothe, S.; Foerster, C.; Fieguth, H.-G.; Pfennig, O.; Schaumann, F.; Martin, C.; Guzman, C.A.; Ebensen, T.; Mueller, M.; Hohlfeld, J.M.; Krug, N.; Braun, A.; Sewald, K.

    2010-08-01

    Prediction of lung innate immune responses is critical for developing new drugs. Well-established immune modulators like lipopolysaccharides (LPS) can elicit a wide range of immunological effects. They are involved in acute lung diseases such as infections or chronic airway diseases such as COPD. LPS has a strong adjuvant activity, but its pyrogenicity has precluded therapeutic use. The bacterial lipopeptide MALP-2 and its synthetic derivative BPPcysMPEG are better tolerated. We have compared the effects of LPS and BPPcysMPEG on the innate immune response in human precision-cut lung slices. Cytokine responses were quantified by ELISA, Luminex, and Meso Scale Discovery technology. The initial response to LPS and BPPcysMPEG was marked by coordinated and significant release of the mediators IL-1{beta}, MIP-1{beta}, and IL-10 in viable PCLS. Stimulation of lung tissue with BPPcysMPEG, however, induced a differential response. While LPS upregulated IFN-{gamma}, BPPcysMPEG did not. This traces back to their signaling pathways via TLR4 and TLR2/6. The calculated exposure doses selected for LPS covered ranges occurring in clinical studies with human beings. Correlation of obtained data with data from human BAL fluid after segmental provocation with endotoxin showed highly comparable effects, resulting in a coefficient of correlation > 0.9. Furthermore, we were interested in modulating the response to LPS. Using dexamethasone as an immunosuppressive drug for anti-inflammatory therapy, we found a significant reduction of GM-CSF, IL-1{beta}, and IFN-{gamma}. The PCLS-model offers the unique opportunity to test the efficacy and toxicity of biological agents intended for use by inhalation in a complex setting in humans.

  14. The inner gel component of Aloe vera suppresses bacterial-induced pro-inflammatory cytokines from human immune cells.

    PubMed

    Habeeb, Fatema; Stables, Graham; Bradbury, Fiona; Nong, Stephanie; Cameron, Pamela; Plevin, Robin; Ferro, Valerie A

    2007-08-01

    The present study was carried out to examine the anti-inflammatory activity of the inner leaf gel component of Aloe barbadensis Miller. A simple in vitro assay was designed to determine the effect of the inner gel on bacterial-induced pro-inflammatory cytokine production, namely TNF-alpha and IL-1 beta, from peripheral blood leukocytes stimulated with Shigella flexneri or LPS. This report describes the suppression of both cytokines with a freeze-dried inner gel powder and a commercial health drink from the same source. Comparison was made with a human monocytic cell-line (THP-1 cells) and a similar trend in responses was demonstrated.

  15. Type 1 and type 2 cytokine profiles in children exposed to or infected with vertically transmitted human immunodeficiency virus.

    PubMed Central

    Lee, B N; Lu, J G; Kline, M W; Paul, M; Doyle, M; Kozinetz, C; Shearer, W T; Reuben, J M

    1996-01-01

    In human immunodeficiency virus (HIV)-infected adults, cytokine production profiles switch from predominantly type 1 (interleukin-2 [IL-2] and gamma interferon [IFN-gamma]) to type 2 (IL-4 and IL-10) cytokines with disease progression. To test this hypothesis in vertically HIV-infected children, we measured cytokine transcription and production in rapid progressors (RPs), seroreverters (SRs), and those children exposed to HIV in utero (P0s). Production of type 1 and type 2 cytokines was measured in peripheral blood mononuclear cell cultures of 8 SR, 25 P0, and 11 RP children. Unstimulated cultures, irrespective of infection and stage of disease, produced similar levels of IL-2, IFN-gamma, IL-4, and IL-10. Upon stimulation with phytohemagglutinin (PHA) plus phorbol-12-myristate-13-acetate (PMA), RP children produced less IL-2 (P < 0.01) and IFN-gamma (P < 0.02) than SR children and also expressed significantly less IFN-gamma mRNA (P < 0.01) than SR children. RP children expressed significantly higher levels of IL-4 mRNA than P0 children (P < 0.03). There were no differences in the production of IL-10 by PHA-PMA-stimulated peripheral blood mononuclear cell cultures among the three groups of children. Our data with these pediatric patients suggest that a deficiency in mitogen-stimulated type 1 cytokine production and excess type 2 cytokine (IL-4) transcription correlate with disease progression. Additional studies with larger sample sizes are needed to test further the hypothesis of the type 1-to-type 2 cytokine switch in children infected with HIV. PMID:8877124

  16. Broad-spectrum sunscreens prevent the secretion of proinflammatory cytokines in human keratinocytes exposed to ultraviolet A and phototoxic lomefloxacin.

    PubMed

    Reinhardt, P; Cybulski, M; Miller, S M; Ferrarotto, C; Wilkins, R; Deslauriers, Y

    2006-02-01

    The combination of phototoxic drugs and ultraviolet (UV) radiation can trigger the release of proinflammatory cytokines. The present study measured the ability of sunscreens to prevent cytokine secretion in human keratinocytes following cotreatment of these cells with a known photoreactive drug and UVA. Keratinocytes were treated for 1 h with increasing concentrations of lomefloxacin (LOM) or norfloxacin (NOR), exposed to 15 J/cm2 UVA, and incubated for 24 h. NOR, owing to the absence of a fluorine atom in position 8, was non-phototoxic and used as a negative control. Cell viability and the release of 3 cytokines were assessed, namely interleukin-1alpha (IL-1alpha), interleukin-6 (IL-6), and tumour necrosis factor-alpha (TNF-alpha). The measurement of these cytokines may be a useful tool for detecting photoreactive compounds. To measure their ability to prevent cytokine secretion, various sunscreens were inserted between the UVA source and the cells. Treatment with NOR, NOR plus UVA, or LOM had no effect on the cells. LOM plus UVA, however, had an effect on cell viability and on cytokine secretion. IL-1alpha levels increased with LOM concentration. The release of TNF-alpha and IL-6 followed the same pattern at lower concentrations of LOM but peaked at 15 micromol/L and decreased at higher concentrations. Sunscreens protected the cells from the effects of LOM plus UVA, as cell viability and levels of cytokines remained the same as in the control cells. In conclusion, the application of broad-spectrum sunscreen by individuals exposed to UVA radiation may prevent phototoxic reactions initiated by drugs such as LOM.

  17. Entamoeba lysyl-tRNA synthetase contains a cytokine-like domain with chemokine activity towards human endothelial cells.

    PubMed

    Castro de Moura, Manuel; Miro, Francesc; Han, Jung Min; Kim, Sunghoon; Celada, Antonio; Ribas de Pouplana, Lluís

    2011-11-01

    Immunological pressure encountered by protozoan parasites drives the selection of strategies to modulate or avoid the immune responses of their hosts. Here we show that the parasite Entamoeba histolytica has evolved a chemokine that mimics the sequence, structure, and function of the human cytokine HsEMAPII (Homo sapiens endothelial monocyte activating polypeptide II). This Entamoeba EMAPII-like polypeptide (EELP) is translated as a domain attached to two different aminoacyl-tRNA synthetases (aaRS) that are overexpressed when parasites are exposed to inflammatory signals. EELP is dispensable for the tRNA aminoacylation activity of the enzymes that harbor it, and it is cleaved from them by Entamoeba proteases to generate a standalone cytokine. Isolated EELP acts as a chemoattractant for human cells, but its cell specificity is different from that of HsEMAPII. We show that cell specificity differences between HsEMAPII and EELP can be swapped by site directed mutagenesis of only two residues in the cytokines' signal sequence. Thus, Entamoeba has evolved a functional mimic of an aaRS-associated human cytokine with modified cell specificity.

  18. Effect Of α2-Adrenergic Agonists And Antagonists On Cytokine Release From Human Lung Macrophages Cultured In Vitro

    PubMed Central

    Piazza, O.; Staiano, R.I.; De Robertis, E.; Conti, G.; Di Crescenzo, V.; Loffredo, S.; Marone, G.; Marinosci, G. Zito; Cataldi, M. M.

    2016-01-01

    The most trusted hypothesis to explain how α2-adrenergic agonists may preserve pulmonary functions in critically ill patients is that they directly act on macrophages by interfering with an autocrine/paracrine adrenergic system that controls cytokine release through locally synthetized noradrenaline and α1- and α2-adrenoreceptors. We tested this hypothesis in primary cultures of resident macrophages from human lung (HLMs). HLMs were isolated by centrifugation on percoll gradients from macroscopically healthy human lung tissue obtained from four different patients at the time of lung resection for cancer. HLMs from these patients showed a significant expression of α2A, α2B and α2C adrenoreceptors both at the mRNA and at the protein level. To evaluate whether α2 adrenoreceptors controlled cytokine release from HMLs, we measured IL-6, IL-8 and TNF-α concentrations in the culture medium in basal conditions and after preincubation with several α2-adrenergic agonists or antagonists. Neither the pretreatment with the α2-adrenergic agonists clonidine, medetomidine or dexdemetomidine or with the α2-adrenergic antagonist yohimbine caused significant changes in the response of any of these cytokines to LPS. These results show that, different from what reported in rodents, clonidine and dexdemetomidine do not directly suppress cytokine release from human pulmonary macrophages. This suggests that alternative mechanisms such as effects on immune cells activation or the modulation of autonomic neurotransmission could be responsible for the beneficial effects of these drugs on lung function in critical patients. PMID:27896229

  19. Expression of proinflammatory cytokines and receptors by human fallopian tubes in organ culture following challenge with Neisseria gonorrhoeae.

    PubMed

    Maisey, Kevin; Nardocci, Gino; Imarai, Monica; Cardenas, Hugo; Rios, Miguel; Croxatto, Horacio B; Heckels, John E; Christodoulides, Myron; Velasquez, Luis A

    2003-01-01

    Infection of the Fallopian tubes (FT) by Neisseria gonorrhoeae can lead to acute salpingitis, an inflammatory condition, which is a major cause of infertility. Challenge of explants of human FT with gonococci induced mRNA expression and protein secretion for the proinflammatory cytokines interleukin (IL)-1alpha, IL-1beta, and tumor necrosis factor alpha (TNF-alpha) but not for granulocyte-macrophage colony-stimulating factor. In contrast, FT expression of IL-6 and of the cytokine receptors IL-6R, TNF receptor I (TNF-RI), and TNF-RII was constitutive and was not increased by gonococcal challenge. These studies suggest that several proinflammatory cytokines are likely to contribute to the cell and tissue damage observed in gonococcal salpingitis.

  20. Expression of Proinflammatory Cytokines and Receptors by Human Fallopian Tubes in Organ Culture following Challenge with Neisseria gonorrhoeae

    PubMed Central

    Maisey, Kevin; Nardocci, Gino; Imarai, Monica; Cardenas, Hugo; Rios, Miguel; Croxatto, Horacio B.; Heckels, John E.; Christodoulides, Myron; Velasquez, Luis A.

    2003-01-01

    Infection of the Fallopian tubes (FT) by Neisseria gonorrhoeae can lead to acute salpingitis, an inflammatory condition, which is a major cause of infertility. Challenge of explants of human FT with gonococci induced mRNA expression and protein secretion for the proinflammatory cytokines interleukin (IL)-1α, IL-1β, and tumor necrosis factor alpha (TNF-α) but not for granulocyte-macrophage colony-stimulating factor. In contrast, FT expression of IL-6 and of the cytokine receptors IL-6R, TNF receptor I (TNF-RI), and TNF-RII was constitutive and was not increased by gonococcal challenge. These studies suggest that several proinflammatory cytokines are likely to contribute to the cell and tissue damage observed in gonococcal salpingitis. PMID:12496205

  1. Distinct cytokine pattern in response to different bacterial pathogens in human brain abscess.

    PubMed

    Bajpai, Anamika; Prasad, Kashi Nath; Mishra, Priyanka; Singh, Aloukick Kumar; Gupta, Rakesh Kumar; Ojha, Bal Krishan

    2014-08-15

    Brain abscess is a focal suppurative process. Host inflammatory response in Gram type and specific bacteria has not been studied in brain abscess. A total of 57 brain abscess patients with monomicrobial infections were studied for Th1 (TNF-α, IFN-γ, IL1-β), Th2 (IL-4, IL-10) and Th17 (IL-17, IL-23) cytokine response by reverse-transcriptase PCR and ELISA. Th1 and Th17 cytokines were significantly elevated in Gram positive (Staphylococcus aureus and Streptococcus intermedius) and Th2 cytokine (IL-10) in Gram negative (Bacteroides fragilis and Escherichia coli) infections (p<0.05). Cytokine levels were significantly higher in abscess than blood (p<0.001). Elevated levels of several inflammatory cytokines (TNF-α, IFN-γ, IL1-β, IL-17 and IL-23) were associated with the duration of symptoms; predisposing factors also influenced the levels of several cytokines. The expression of inflammatory cytokines in abscess was influenced by the bacterial pathogen, duration of symptoms and predisposing factors. Local milieu of brain plays significant role in secretion of various cytokines. Copyright © 2014 Elsevier B.V. All rights reserved.

  2. A comparitive assessement of cytokine expression in human-derived cell lines exposed to alpha particles and X-rays.

    PubMed

    Chauhan, Vinita; Howland, Matthew; Wilkins, Ruth

    2012-01-01

    Alpha- (α-) particle radiation exposure has been linked to the development of lung cancer and has been identified as a radiation type likely to be employed in radiological dispersal devices. Currently, there exists a knowledge gap concerning cytokine modulations associated with exposure to α-particles. Bio-plex technology was employed to investigate changes in proinflammatory cytokines in two human-derived cell lines. Cells were irradiated at a dose of 1.5 Gy to either α-particles or X-rays at equivalent dose rates. The two cell lines exhibited a unique pattern of cytokine expression and the response varied with radiation type. Of the 27 cytokines assessed, only vascular endothelin growth factor (VEGF) was observed to be modulated in both cell lines solely after α-particle exposure, and the expression of VEGF was shown to be dose responsive. These results suggest that certain proinflammatory cytokines may be involved in the biological effects related to α- particle exposure and the responses are cell type and radiation type specific.

  3. 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.

  4. Cytokine responses of human intestinal epithelial-like Caco-2 cells to the nonpathogenic bacterium Bacillus subtilis (natto).

    PubMed

    Hosoi, Tomohiro; Hirose, Rieko; Saegusa, Shizue; Ametani, Akio; Kiuchi, Kan; Kaminogawa, Shuichi

    2003-05-15

    Intestinal epithelial cells produce cytokines in response to pathogenic bacteria. However, cellular responses of these cells to nonpathogenic strains, such as Bacillus subtilis, are yet to be determined. In this study, we investigate whether epithelial-like human colon carcinoma Caco-2 cells produce cytokines in response to B. subtilis or B. subtilis (natto). The latter strain is utilized for manufacturing the fermented soy food "natto". Live cells of nonpathogenic B. subtilis JCM 1465(T), B. subtilis (natto) and E. coli JCM 1649(T), as well as pathogenic S. enteritidis JCM 1652 and P. aeruginosa JCM 5516 strains, induced secretion of interleukin-6 (IL-6) and/or IL-8, but not IL-7, IL-15 or tumor necrosis factor alpha (TNF-alpha). Transepithelial electrical resistance (TER) of Caco-2 cell monolayers cultured with E. coli, S. enteritidis or P. aeruginosa decreased more rapidly than that of cells cultured with B. subtilis or B. subtilis (natto). The amounts of cytokine induced by B. subtilis (natto) cells were strain-dependent. Moreover, B. subtilis (natto) cells subjected to hydrochloric acid treatment, but not autoclaving, induced a higher secretion of IL-6 and IL-8 than intact cells. Tyrosine kinase inhibitors, including AG126 and genistein, suppressed cytokine secretion. Our results suggest that the nonpathogenic B. subtilis (natto) bacterium induces cytokine responses in intestinal epithelial cells via activation of an intracellular signaling pathway, such as that of nuclear factor-kappa B (NF-kappaB).

  5. A Comparitive Assessement of Cytokine Expression in Human-Derived Cell Lines Exposed to Alpha Particles and X-Rays

    PubMed Central

    Chauhan, Vinita; Howland, Matthew; Wilkins, Ruth

    2012-01-01

    Alpha- (α-) particle radiation exposure has been linked to the development of lung cancer and has been identified as a radiation type likely to be employed in radiological dispersal devices. Currently, there exists a knowledge gap concerning cytokine modulations associated with exposure to α-particles. Bio-plex technology was employed to investigate changes in proinflammatory cytokines in two human-derived cell lines. Cells were irradiated at a dose of 1.5 Gy to either α-particles or X-rays at equivalent dose rates. The two cell lines exhibited a unique pattern of cytokine expression and the response varied with radiation type. Of the 27 cytokines assessed, only vascular endothelin growth factor (VEGF) was observed to be modulated in both cell lines solely after α-particle exposure, and the expression of VEGF was shown to be dose responsive. These results suggest that certain proinflammatory cytokines may be involved in the biological effects related to α- particle exposure and the responses are cell type and radiation type specific. PMID:22619631

  6. 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.

  7. Cytokine polymorphisms are associated with poor sleep maintenance in adults living with human immunodeficiency virus/acquired immunodeficiency syndrome.

    PubMed

    Lee, Kathryn A; Gay, Caryl; Pullinger, Clive R; Hennessy, Mary Dawn; Zak, Rochelle S; Aouizerat, Bradley E

    2014-03-01

    Cytokine activity and polymorphisms have been associated with sleep outcomes in prior animal and human research. The purpose of this study was to determine whether circulating plasma cytokines and cytokine polymorphisms are associated with the poor sleep maintenance commonly experienced by adults living with human immunodeficiency virus/acquired immunodeficiency syndrome (HIV/AIDS). Cross-sectional descriptive study. HIV clinics and community sites in the San Francisco Bay area. A convenience sample of 289 adults (193 men, 73 women, and 23 transgender) living with HIV/AIDS. None. A wrist actigraph was worn for 72 h to estimate the percentage of wake after sleep onset (WASO%) and total sleep time (TST), plasma cytokines were analyzed, and genotyping was conducted for 15 candidate genes involved in cytokine signaling: interferon-gamma (IFNG), IFNG receptor 1 (IFNGR1), interleukins (IL1B, IL1R2, IL1R2, IL2, IL4, IL6, IL8, IL10, IL13, IL17A), nuclear factor of kappa light polypeptide gene enhancer in B cells (NFKB1 and NFKB2), and tumor necrosis factor-alpha (TNFA). Controlling for demographic variables such as race and sex, and clinical variables such as CD4+ count and medications, higher WASO% was associated with single nucleotide polymorphisms (SNPs) of IL1R2 rs11674595 and TNFA rs1041981 and less WASO% was associated with IL2 rs2069776. IL1R2 rs11674595 and TNFA rs1041981 were also associated with short sleep duration. This study strengthens the evidence for an association between inflammation and sleep maintenance problems. In this chronic illness population, cytokine polymorphisms associated with wake after sleep onset provide direction for intervention research aimed at comparing anti-inflammatory mechanisms with hypnotic agents for improving sleep maintenance and total sleep time.

  8. 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.

  9. 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.

  10. Cancer Cytokines and the Relevance of 3D Cultures for Studying those Implicated in Human Cancers.

    PubMed

    Maddaly, Ravi; Subramaniyan, Aishwarya; Balasubramanian, Harini

    2017-03-06

    Cancers are complex conditions and involving several factors for oncogenesis and progression. Of the various factors influencing the physiology of cancers, cytokines are known to play significant roles as mediators of functions. Intricate cytokine networks have been identified in cancers and interest in cytokines associated with cancers has been gaining ground. Of late, some of these cytokines are even identified as potential targets for cancer therapy apart from a few others such as IL-6 being identified as markers for disease prognosis. Of the major contributors to cancer research, cancer cell lines occupy the top slot as the most widely used material in vitro. In vitro cell cultures have seen significant evolution by the introduction of 3 dimensional (3D) culture systems. 3D cell cultures are now widely accepted as excellent material for cancer research which surpasses the traditional monolayer cultures. Cancer research has benefitted from 3D cell cultures for understanding the various hallmarks of cancers. However, the potential of these culture systems are still unexploited for cancer cytokine research compared to the other aspects of cancers such as gene expression changes, drug-induced toxicity, morphology, angiogenesis and invasion. Considering the importance of cancer cytokines, 3D cell cultures can be better utilized in understanding their roles and functions. Some of the possibilities where 3D cell cultures can contribute to cancer cytokine research arise from the distinct morphology of the tumor spheroids, the extracellular matrix (ECM), and the spontaneous occurrence of nutrient and oxygen gradients. Also, the 3D culture models enable one to co-culture different types of cells as a simulation of in vivo conditions, enhancing their utility to study cancer cytokines. We review here the cancer associated cytokines the contributions of 3D cancer cell cultures for studying cancer cytokines. This article is protected by copyright. All rights reserved.

  11. 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.

  12. Expression of suppressor of cytokine signaling genes in human elderly and Alzheimer's disease brains and human microglia.

    PubMed

    Walker, D G; Whetzel, A M; Lue, L-F

    2015-08-27

    Multiple cellular systems exist to prevent uncontrolled inflammation in brain tissues; the suppressor of cytokine signaling (SOCS) proteins have key roles in these processes. SOCS proteins are involved in restricting cellular signaling pathways by enhancing the degradation of activated receptors and removing the stimuli for continued activation. There are eight separate SOCS genes that code for proteins with similar structures and properties. All SOCS proteins can reduce signaling of activated transcription factors Janus kinase (JAK) and signal transducer and activator of transcription (STAT), but they also regulate many other signaling pathways. SOCS-1 and SOCS-3 have particular roles in regulating inflammatory processes. Chronic inflammation is a key feature of the pathology present in Alzheimer's disease (AD)-affected brains resulting from responses to amyloid plaques or neurofibrillary tangles, the pathological hallmarks of AD. The goal of this study was to examine SOCS gene expression in human non-demented (ND) and AD brains and in human brain-derived microglia to determine if AD-related pathology resulted in a deficit of these critical molecules. We demonstrated that SOCS-1, SOCS-2, SOCS-3 and cytokine-inducible SH2 containing protein (CIS) mRNA expression was increased in amyloid beta peptide (Aβ)- and inflammatory-stimulated microglia, while SOCS-6 mRNA expression was decreased by both types of treatments. Using human brain samples from the temporal cortex from ND and AD cases, SOCS-1 through SOCS-7 and CIS mRNA and SOCS-1 through SOCS-7 protein could be detected constitutively in ND and AD human brain samples. Although, the expression of key SOCS genes did not change to a large extent as a result of AD pathology, there were significantly increased levels of SOCS-2, SOCS-3 and CIS mRNA and increased protein levels of SOCS-4 and SOCS-7 in AD brains. In summary, there was no evidence of a deficit of these key inflammatory regulating proteins in aged or AD

  13. Transcriptional regulation of human inducible nitric oxide synthase (NOS2) gene by cytokines: initial analysis of the human NOS2 promoter.

    PubMed Central

    de Vera, M E; Shapiro, R A; Nussler, A K; Mudgett, J S; Simmons, R L; Morris, S M; Billiar, T R; Geller, D A

    1996-01-01

    The expression of inducible nitric oxide synthase (NOS2) is complex and is regulated in part by gene transcription. In this investigation we studied the regulation of NOS2 in a human liver epithelial cell line (AKN-1) which expresses high levels of NOS2 mRNA and protein in response to tumor necrosis factor alpha, interleukin 1 beta, and interferon gamma (cytokine mix, CM). Nuclear run-on analysis revealed that CM transcriptionally activated the human NOS2 gene. To delineate the cytokine-responsive regions of the human NOS2 promoter, we stimulated AKN-1 cells with CM following transfection of NOS2 luciferase constructs. Analysis of the first 3.8 kb upstream of the NOS2 gene demonstrated basal promoter activity but failed to show any cytokine-inducible activity. However, 3- to 5-fold inductions of luciferase activity were seen in constructs extending up to -5.8 and -7.0 kg, and a 10-fold increase was seen upon transfection of a -16 kb construct. Further analysis of various NOS2 luciferase constructs ligated upstream of the thymidine kinase promoter identified three regions containing cytokine-responsive elements in the human NOS2 gene: -3.8 to -5.8, -5.8 to -7.0, and -7.0 to -16 kb. These results are in marked contrast with the murine macrophage NOS2 promoter in which only 1 kb of the proximal 5' flanking region is necessary to confer inducibility to lipopolysaccharide and interferon gamma. These data demonstrate that the human NOS2 gene is transcriptionally regulated by cytokines and identify multiple cytokine-responsive regions in the 5' flanking region of the human NOS2 gene. Images Fig. 1 Fig. 2 PMID:8577713

  14. Dual role for RhoA in suppression and induction of cytokines in the human neutrophil

    PubMed Central

    Fessler, Michael B.; Arndt, Patrick G.; Just, Ingo; Nick, Jerry A.; Malcolm, Kenneth C.; Scott Worthen, G.

    2007-01-01

    Production of tumor necrosis factor-α (TNFα) by the neutrophil (PMN) is a pivotal event in innate immunity, but the signals regulating TNFα induction in this primary cell are poorly understood. Herein, we use protein transduction to identify novel, opposing anti– and pro–cytokine-inducing roles for RhoA in the resting and lipopolysaccharide (LPS)–stimulated human PMN, respectively. In the resting cell, RhoA suppresses Cdc42 activation, IκBα degradation, nuclear factor-κB (NF-κB) activation, and induction of TNFα and NF-κB–dependent chemokines. Suppression of TNFα induction by RhoA is Rho kinase α (ROCKα) independent, but Cdc42 dependent, because TNFα induction by C3 transferase is attenuated by inhibition of Cdc42, and constitutively active Cdc42 suffices to activate NF-κB and induce TNFα. By contrast, we also place RhoA downstream of p38 mitogen-activated protein kinase and Cdc42 in a novel LPS-activated pathway in which p38, Cdc42, and ROCKα all promote TNFα protein expression. The p65 subunit of NF-κB coprecipitates with RhoA in a manner sensitive to the RhoA activation state. Our findings suggest a new, 2-faced role for RhoA as a checkpoint in innate immunity. PMID:17018860

  15. Subfractions of enamel matrix derivative differentially influence cytokine secretion from human oral fibroblasts

    PubMed Central

    Villa, Oscar; Brookes, Steven J; Thiede, Bernd; Heijl, Lars; Lyngstadaas, Staale P

    2015-01-01

    Enamel matrix derivative is used to promote periodontal regeneration during the corrective phase of the treatment of periodontal defects. Our main goal was to analyze the bioactivity of different molecular weight fractions of enamel matrix derivative. Enamel matrix derivative, a complex mixture of proteins, was separated into 13 fractions using size-exclusion chromatography and characterized by sodium dodecyl sulfate–polyacrylamide gel electrophoresis and liquid chromatography–electrospray ionization–tandem mass spectrometry. Human periodontal ligament fibroblasts were treated with either enamel matrix derivative or the different fractions. Proliferation and cytokine secretion to the cell culture medium were measured and compared to untreated cells. The liquid chromatography–electrospray ionization–tandem mass spectrometry analyses revealed that the most abundant peptides were amelogenin and leucine-rich amelogenin peptide related. The fractions containing proteins above 20 kDa induced an increase in vascular endothelial growth factor and interleukin-6 secretion, whereas lower molecular weight fractions enhanced proliferation and secretion of interleukin-8 and monocyte chemoattractant protein-1 and reduced interleukin-4 release. The various molecular components in the enamel matrix derivative formulation might contribute to reported effects on tissue regeneration through their influence on vascularization, the immune response, and chemotaxis. PMID:26090085

  16. Applications of monoclonal antibodies and recombinant cytokines for the treatment of human colorectal and other carcinomas

    SciTech Connect

    Greiner, J.W.; Smalley, R.V.; Borden, E.C.; Martin, E.W.; Guadagni, F.; Roselli, M.; Schlom, J. )

    1991-01-01

    Monoclonal antibodies (MAbs) which recognize a human tumor antigen, termed tumor-associated glycoprotein-72 (TAG-72), have successfully been used to localize primary as well as metastatic colorectal tumor lesions in patients. The localization of the anti-TAG-72 MAbs has also been exploited intraoperatively using a hand-held gamma probe. That procedure, termed radioimmunoguided surgery (RIGS), has identified occult tumors which were not detected using standard external imaging techniques. In another clinical trial, interferon-gamma (IFN-gamma) was administered intraperitoneally to patients diagnosed with either gastrointestinal or ovarian carcinoma with secondary ascites. Analysis of the tumor cells isolated from the malignant ascites revealed a substantial increase in TAG-72 expression on the surface of tumor cells isolated from seven of eight patients. The results provide evidence that the combination of an anti-carcinoma MAb with the administration of a cytokine, such as IFN-gamma, may be an effective approach for the detection and subsequent treatment, of colorectal carcinoma. 15 references.

  17. Fu-Ling, a Chinese herbal drug, modulates cytokine secretion by human peripheral blood monocytes.

    PubMed

    Yu, S J; Tseng, J

    1996-01-01

    Fu-Ling, the sclederma of Poria cocos (Schw.) Wolf, has long been used as a sedative and diuretic in traditional Chinese herbal medicine. Our study demonstrated that the substances extracted from Fu-Ling by 50% hot ethanol significantly augmented the secretion of interleukins IL-1 beta and IL-6 6 h after in vitro cultivation of human peripheral blood monocytes. The augmented effect was dose dependent. Tumour necrosis factor-alpha (TNF-alpha) secretion was also increased as the cells were treated with 0.4 mg/ml or higher doses of Fu-Ling extract. By contrast, Fu-Ling extract significantly suppressed the secretion of transforming growth factor-beta (TGF-beta) 3 h after the in vitro drug treatment. The suppressive effect was shown at doses as low as 0.2 mg/ml of Fu-Ling extract. Since Fu-Ling extract enhanced the secretion of immune stimulators (IL-1 beta, IL-6 and TNF-alpha) but suppressed the secretion of an immune suppressor (TGF-beta), the substance in 50% hot ethanol extract of Fu-Ling might have potentiated the immune response. Fu-Ling extract was further fractionated by reverse-phase column chromatography and high-performance liquid chromatography (HPLC). The components showing activity in modulating the cytokine secretion were relatively high in hydrophobicity.

  18. Antibody and cytokine levels in humans fed on by the common bedbug, Cimex lectularius L.

    PubMed

    Sheele, J M; Ridge, G E; Coppolino, K; Bonfield, T; Young, A B; Gaines, S L; McCormick, T S

    2017-03-01

    Little is known about cimicosis, the resultant dermal reaction from feeding activity by the common bedbug, Cimex lectularius L. We fed C. lectularius on human study subjects four times over four weeks and measured serum cytokine and antibody levels, and subjects recorded any cimicosis. The average time for subjects to develop cimicosis decreased with each feeding from 8.4, to 2.1, 1.5 and 1.3 days, respectively. There were no significant changes in total IgG, IgG1, IgG2, IgG4 or IgE levels between the first and fourth bedbug feedings, but there was a significant decrease in total IgG3 levels (P<.001). IgG4 was not required for cimicosis. Higher IgG2 and IgG4 levels at study visit 4 were associated with an increased duration of cimicosis (P=.04) and lower pruritis (P=.03), respectively. There were no significant changes in serum TNF-α, IL-1β, IL-4, IL-5, IL-6, IL-10, IFN-γ and IL-17A levels before and one hour after the C. lectularius feeding. Lower post-C. lectularius feeding IL-6 levels were associated with increased pruritis (P=.001) and the time to maximum pruritis (P=.04), respectively. Higher post-C. lectularius feeding IL-5 levels were associated with a longer duration of pruritis (P=.05).

  19. Vitamin D attenuates cytokine-induced remodeling in human fetal airway smooth muscle cells.

    PubMed

    Britt, Rodney D; Faksh, Arij; Vogel, Elizabeth R; Thompson, Michael A; Chu, Vivian; Pandya, Hitesh C; Amrani, Yassine; Martin, Richard J; Pabelick, Christina M; Prakash, Y S

    2015-06-01

    Asthma in the pediatric population remains a significant contributor to morbidity and increasing healthcare costs. Vitamin D3 insufficiency and deficiency have been associated with development of asthma. Recent studies in models of adult airway diseases suggest that the bioactive Vitamin D3 metabolite, calcitriol (1,25-dihydroxyvitamin D3 ; 1,25(OH)2 D3 ), modulates responses to inflammation; however, this concept has not been explored in developing airways in the context of pediatric asthma. We used human fetal airway smooth muscle (ASM) cells as a model of the early postnatal airway to explore how calcitriol modulates remodeling induced by pro-inflammatory cytokines. Cells were pre-treated with calcitriol and then exposed to TNFα or TGFβ for up to 72 h. Matrix metalloproteinase (MMP) activity, production of extracellular matrix (ECM), and cell proliferation were assessed. Calcitriol attenuated TNFα enhancement of MMP-9 expression and activity. Additionally, calcitriol attenuated TNFα and TGFβ-induced collagen III expression and deposition, and separately, inhibited proliferation of fetal ASM cells induced by either inflammatory mediator. Analysis of signaling pathways suggested that calcitriol effects in fetal ASM involve ERK signaling, but not other major inflammatory pathways. Overall, our data demonstrate that calcitriol can blunt multiple effects of TNFα and TGFβ in developing airway, and point to a potentially novel approach to alleviating structural changes in inflammatory airway diseases of childhood.

  20. Granzyme K synergistically potentiates LPS-induced cytokine responses in human monocytes

    PubMed Central

    Wensink, Annette C.; Kemp, Vera; Fermie, Job; García Laorden, M. Isabel; van der Poll, Tom; Hack, C. Erik; Bovenschen, Niels

    2014-01-01

    Granzymes are serine proteases released by cytotoxic lymphocytes to induce apoptosis in virus-infected cells and tumor cells. Evidence is emerging that granzymes also play a role in controlling inflammation. Granzyme serum levels are elevated in patients with autoimmune diseases and infections, including sepsis. However, the function of extracellular granzymes in inflammation largely remains unknown. Here, we show that granzyme K (GrK) binds to Gram-negative bacteria and their cell-wall component lipopolysaccharide (LPS). GrK synergistically enhances LPS-induced cytokine release in vitro from primary human monocytes and in vivo in a mouse model of LPS challenge. Intriguingly, these extracellular effects are independent of GrK catalytic activity. GrK disaggregates LPS from micelles and augments LPS–CD14 complex formation, thereby likely boosting monocyte activation by LPS. We conclude that extracellular GrK is an unexpected direct modulator of LPS–TLR4 signaling during the antimicrobial innate immune response. PMID:24711407

  1. Subfractions of enamel matrix derivative differentially influence cytokine secretion from human oral fibroblasts.

    PubMed

    Villa, Oscar; Brookes, Steven J; Thiede, Bernd; Heijl, Lars; Lyngstadaas, Staale P; Reseland, Janne E

    2015-01-01

    Enamel matrix derivative is used to promote periodontal regeneration during the corrective phase of the treatment of periodontal defects. Our main goal was to analyze the bioactivity of different molecular weight fractions of enamel matrix derivative. Enamel matrix derivative, a complex mixture of proteins, was separated into 13 fractions using size-exclusion chromatography and characterized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and liquid chromatography-electrospray ionization-tandem mass spectrometry. Human periodontal ligament fibroblasts were treated with either enamel matrix derivative or the different fractions. Proliferation and cytokine secretion to the cell culture medium were measured and compared to untreated cells. The liquid chromatography-electrospray ionization-tandem mass spectrometry analyses revealed that the most abundant peptides were amelogenin and leucine-rich amelogenin peptide related. The fractions containing proteins above 20 kDa induced an increase in vascular endothelial growth factor and interleukin-6 secretion, whereas lower molecular weight fractions enhanced proliferation and secretion of interleukin-8 and monocyte chemoattractant protein-1 and reduced interleukin-4 release. The various molecular components in the enamel matrix derivative formulation might contribute to reported effects on tissue regeneration through their influence on vascularization, the immune response, and chemotaxis.

  2. 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.

  3. 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

  4. Substance P Receptor Antagonist Suppresses Inflammatory Cytokine Expression in Human Disc Cells.

    PubMed

    Kepler, Christopher K; Markova, Dessislava Z; Koerner, John D; Mendelis, Joseph; Chen, Chiu-Ming; Vaccaro, Alexander R; Risbud, Makarand V; Albert, Todd J; Anderson, D Greg

    2015-08-15

    Laboratory study. To evaluate whether blockade of the Substance P (SP) NK1R attenuates its proinflammatory effect on human intervertebral disc cells (IVD), and to evaluate the signaling pathways associated with SP. SP and its receptors are expressed in human IVD cells, and cause upregulation of inflammatory mediators; however, the effects of blocking these receptors have not been studied in human IVD cells. Human annulus fibrosus (AF) and nucleus pulposus (NP) cells were expanded in monolayer, and then suspended in alginate beads. The alginate beads were treated with culture medium first containing a high affinity NK1R antagonist (L-760735) at different concentrations, and then with medium containing both NK1R antagonist and SP at 2 concentrations. Ribonucleic acid was isolated and transcribed into cDNA. Quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) was performed to evaluate expression of interleukin (IL)-1β, IL-6, and IL-8. Western blot analysis was performed to examine levels of the phosphorylated p38 mitogen-activated protein kinase (MAPK), extracellular signal regulated kinase 1/2 (ERK1/2) and nuclear factor kappa-light-chain-enhancer of activated B cells (NFκB p65). The cells were pretreated with specific inhibitors of p38 (SB203580), ERK1/2 (PD98059), and p65 (SM7368) and then stimulated with SP. We detected expression of NK1R, neurokinin receptor 2 (NK2R), and neurokinin receptor 3 (NK3R) in AF and NP cells. Treatment of disc cells with the NK1R antagonist was able to suppress expression of IL-1β, IL-6, and IL-8 in a dose-dependent manner. SP stimulation increased phosphorylation of p38-MAPK and ERK1/2, but not of NFκB p65. This indicates that p38-MAPK and ERK1/2 control SP-induced cytokine expression independently from NF-kB p65. Inhibition of p38 and ERK1/2 activation reduced SP-induced IL-6 production in human disc cells. NK1R is responsible for the proinflammatory effect of SP on IVD cells and this effect can be blocked by

  5. Effects of resolvin D1 on cell survival and cytokine expression of human gingival fibroblasts.

    PubMed

    Khaled, Mohamed; Shibani, Nouf-Al; Labban, Nawaf; Batarseh, Ghada; Song, Fengyu; Ruby, John; Windsor, L Jack

    2013-12-01

    Tissue breakdown in periodontitis is initiated by bacteria, such as Porphyromonas gingivalis, and is caused largely by host responses. Resolvins protect the host against acute inflammation by blocking the migration of polymorphonuclear neutrophils to initiate resolution. The effects of resolvins on human gingival fibroblasts (HGFs) are unknown. This study examines the effects of resolvin D1 on HGF survival and cytokine expression when treated with or without P. gingivalis supernatant. Cytotoxicity of resolvin D1 on HGFs with or without a toxic level of P. gingivalis supernatant was measured with lactate dehydrogenase assays. Cytokine arrays were performed on HGF-conditioned media treated with or without resolvin D1 and with or without P. gingivalis supernatant. Resolvin D1 had no cytotoxic effects on HGFs at concentrations between 1 and 1,000 nM (all P > 0.05). Resolvin D1 (1,000 nM) significantly inhibited the toxic effects of 13.5% (v/v) P. gingivalis supernatant on HGFs (P = 0.002). Resolvin D1 significantly reduced the expression of interleukin (IL)-6 (P = 0.010) and monocyte chemoattractant protein (MCP)-1 (P = 0.04) in untreated fibroblasts. P. gingivalis (10%) supernatant significantly increased the expression levels of granulocyte-macrophage colony-stimulating factor (CSF), granulocyte CSF, growth-regulated oncogene (GRO), IL-5, IL-6, IL-7, IL-8, IL-10, MCP-1, MCP-2, MCP-3, and monokine induced by γ-interferon. Resolvin D1 significantly reduced the expression of GRO (P = 0.04), marginally reduced the levels of MCP-1 (P = 0.10), and marginally increased the levels of transforming growth factor (TGF)-β1 (P = 0.07) from HGFs treated with P. gingivalis supernatant. Resolvin D1 altered the cytotoxicity of P. gingivalis supernatant on HGFs. Resolvin D1 significantly reduced GRO, marginally reduced MCP-1, and marginally increased TGF-β1 from P. gingivalis-treated HGFs, which could alter the ability of P. gingivalis to induce inflammation.

  6. An Ethyl Acetate Fraction of Moringa oleifera Lam. Inhibits Human Macrophage Cytokine Production Induced by Cigarette Smoke

    PubMed Central

    Kooltheat, Nateelak; Pankla Sranujit, Rungnapa; Chumark, Pilaipark; Potup, Pachuen; Laytragoon-Lewin, Nongnit; Usuwanthim, Kanchana

    2014-01-01

    Moringa oleifera Lam. (MO) has been reported to harbor anti-oxidation and anti-inflammatory activity and useful in the treatment of inflammatory diseases. However, despite these findings there has been little work done on the effects of MO on immune cellular function. Since macrophages, TNF and related cytokines play an important pathophysiologic role in lung damage induced by cigarette smoke, we examined the effects of MO on cigarette smoke extract (CSE)—induced cytokine production by human macrophages. An ethyl acetate fraction of MO (MOEF) was prepared from fresh leaves extract of Moringa and shown to consist of high levels of phenolic and antioxidant activities. Human monocyte derived macrophages (MDM) pre-treated with varying concentrations of MOEF showed decreased production of TNF, IL-6 and IL-8 in response to both LPS and CSE. The decrease was evident at both cytokine protein and mRNA levels. Furthermore, the extract inhibited the expression of RelA, a gene implicated in the NF-κB p65 signaling in inflammation. The findings highlight the ability of MOEF to inhibit cytokines (IL-8) which promote the infiltration of neutrophils into the lungs and others (TNF, IL-6) which mediate tissue disease and damage. PMID:24553063

  7. Flagella from Five Cronobacter Species Induce Pro-Inflammatory Cytokines in Macrophage Derivatives from Human Monocytes

    PubMed Central

    Cruz-Córdova, Ariadnna; Rocha-Ramírez, Luz M.; Ochoa, Sara A.; Gónzalez-Pedrajo, Bertha; Espinosa, Norma; Eslava, Carlos; Hernández-Chiñas, Ulises; Mendoza-Hernández, Guillermo; Rodríguez-Leviz, Alejandra; Valencia-Mayoral, Pedro; Sadowinski-Pine, Stanislaw; Hernández-Castro, Rigoberto; Estrada-García, Iris; Muñoz-Hernández, Onofre; Rosas, Irma; Xicohtencatl-Cortes, Juan

    2012-01-01

    Cronobacter spp. are opportunistic pathogens linked to lie-threatening infections in neonates and contaminated powdered infant formula that has been epidemiologically associated with these cases. Clinical symptoms of Cronobacter include necrotizing enterocolitis, bacteremia, and meningitis. Flagella from C. sakazakii are involved in biofilm formation and its adhesion to epithelial cells. We investigated the role of flagella from C. sakazakii ST1 and ST4, C. malonaticus, C. muytjensii, C. turicensis and C. dublinensis during the activation of cytokines (IL-8, TNF-α, and IL-10) in macrophage derivatives from human monocytes, which has not been extensively studied. The production and identity of flagella from the five Cronobacter species were visualized and recognized with anti-flagella antibodies by immunogold labeling through transmission electron microscopy. Purified flagella were dissociated into monomers in 12% SDS-PAGE Coomassie blue-stained gels showing a band of ∼28 kDa and, in addition, mass spectrometry revealed the presence of several peptides that correspond to flagellin. Flagella (100 ng) induced the release of IL-8 (3314–6025 pg/ml), TNF-α (39–359 pg/ml), and IL-10 (2–96 pg/ml), in macrophage isolates from human monocytes and similar results were obtained when flagella were dissociated into monomers. Inhibition assays using three dilutions of anti-flagella antibodies (1∶10, 1∶100, and 1∶200) suppressed the secretion of IL-8, TNF-α, and IL-10 between 95–100% using 100 ng of protein. A transfection assay using 293-hTLR5 cells showed IL-8 release of 197 pg/ml and suppression in the secretion of IL-8 when anti-hTLR5-IgA antibodies were used at different concentrations. These observations suggest that flagella and flagellin are involved in an inflammatory response dependent on TLR5 recognition, which could contribute to the pathogenesis of the bacteria. PMID:23284883

  8. Flagella from five Cronobacter species induce pro-inflammatory cytokines in macrophage derivatives from human monocytes.

    PubMed

    Cruz-Córdova, Ariadnna; Rocha-Ramírez, Luz M; Ochoa, Sara A; González-Pedrajo, Bertha; Gónzalez-Pedrajo, Bertha; Espinosa, Norma; Eslava, Carlos; Hernández-Chiñas, Ulises; Mendoza-Hernández, Guillermo; Rodríguez-Leviz, Alejandra; Valencia-Mayoral, Pedro; Sadowinski-Pine, Stanislaw; Hernández-Castro, Rigoberto; Estrada-García, Iris; Muñoz-Hernández, Onofre; Rosas, Irma; Xicohtencatl-Cortes, Juan

    2012-01-01

    Cronobacter spp. are opportunistic pathogens linked to lie-threatening infections in neonates and contaminated powdered infant formula that has been epidemiologically associated with these cases. Clinical symptoms of Cronobacter include necrotizing enterocolitis, bacteremia, and meningitis. Flagella from C. sakazakii are involved in biofilm formation and its adhesion to epithelial cells. We investigated the role of flagella from C. sakazakii ST1 and ST4, C. malonaticus, C. muytjensii, C. turicensis and C. dublinensis during the activation of cytokines (IL-8, TNF-α, and IL-10) in macrophage derivatives from human monocytes, which has not been extensively studied. The production and identity of flagella from the five Cronobacter species were visualized and recognized with anti-flagella antibodies by immunogold labeling through transmission electron microscopy. Purified flagella were dissociated into monomers in 12% SDS-PAGE Coomassie blue-stained gels showing a band of ∼28 kDa and, in addition, mass spectrometry revealed the presence of several peptides that correspond to flagellin. Flagella (100 ng) induced the release of IL-8 (3314-6025 pg/ml), TNF-α (39-359 pg/ml), and IL-10 (2-96 pg/ml), in macrophage isolates from human monocytes and similar results were obtained when flagella were dissociated into monomers. Inhibition assays using three dilutions of anti-flagella antibodies (1∶10, 1∶100, and 1∶200) suppressed the secretion of IL-8, TNF-α, and IL-10 between 95-100% using 100 ng of protein. A transfection assay using 293-hTLR5 cells showed IL-8 release of 197 pg/ml and suppression in the secretion of IL-8 when anti-hTLR5-IgA antibodies were used at different concentrations. These observations suggest that flagella and flagellin are involved in an inflammatory response dependent on TLR5 recognition, which could contribute to the pathogenesis of the bacteria.

  9. Human Invasive Muscular Sarcocystosis Induces Th2 Cytokine Polarization and Biphasic Cytokine Changes, Based on an Investigation among Travelers Returning from Tioman Island, Malaysia.

    PubMed

    Tappe, Dennis; Slesak, Günther; Pérez-Girón, José Vicente; Schäfer, Johannes; Langeheinecke, Andreas; Just-Nübling, Gudrun; Muñoz-Fontela, César; Püllmann, Kerstin

    2015-06-01

    Sarcocystis nesbitti is a parasite responsible for a biphasic eosinophilic febrile myositis syndrome in two recent outbreaks in Malaysia. We demonstrate Th2 cytokine polarization in infected travelers, an overall cytokine production decrease in the early phase of the disease suggestive of initial immunosuppression, and elevated levels of proinflammatory and chemotactic cytokines in the later myositic phase. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  10. Cytokine Polymorphisms are Associated with Poor Sleep Maintenance in Adults Living with Human Immunodeficiency Virus/Acquired Immunodeficiency Syndrome

    PubMed Central

    Lee, Kathryn A.; Gay, Caryl; Pullinger, Clive R.; Hennessy, Mary Dawn; Zak, Rochelle S.; Aouizerat, Bradley E.

    2014-01-01

    Study Objectives: Cytokine activity and polymorphisms have been associated with sleep outcomes in prior animal and human research. The purpose of this study was to determine whether circulating plasma cytokines and cytokine polymorphisms are associated with the poor sleep maintenance commonly experienced by adults living with human immunodeficiency virus/acquired immunodeficiency syndrome (HIV/AIDS). Design: Cross-sectional descriptive study. Setting: HIV clinics and community sites in the San Francisco Bay area. Participants: A convenience sample of 289 adults (193 men, 73 women, and 23 transgender) living with HIV/AIDS. Interventions: None. Measurements and Results: A wrist actigraph was worn for 72 h to estimate the percentage of wake after sleep onset (WASO%) and total sleep time (TST), plasma cytokines were analyzed, and genotyping was conducted for 15 candidate genes involved in cytokine signaling: interferon-gamma (IFNG), IFNG receptor 1 (IFNGR1), interleukins (IL1B, IL1R2, IL1R2, IL2, IL4, IL6, IL8, IL10, IL13, IL17A), nuclear factor of kappa light polypeptide gene enhancer in B cells (NFKB1 and NFKB2), and tumor necrosis factor-alpha (TNFA). Controlling for demographic variables such as race and sex, and clinical variables such as CD4+ count and medications, higher WASO% was associated with single nucleotide polymorphisms (SNPs) of IL1R2 rs11674595 and TNFA rs1041981 and less WASO% was associated with IL2 rs2069776. IL1R2 rs11674595 and TNFA rs1041981 were also associated with short sleep duration. Conclusions: This study strengthens the evidence for an association between inflammation and sleep maintenance problems. In this chronic illness population, cytokine polymorphisms associated with wake after sleep onset provide direction for intervention research aimed at comparing anti-inflammatory mechanisms with hypnotic agents for improving sleep maintenance and total sleep time. Citation: Lee KA; Gay C; Pullinger CR; Hennessy MD; Zak RS; Aouizerat BE

  11. Glucagon-like peptide-1 protects human islets against cytokine-mediated β-cell dysfunction and death: a proteomic study of the pathways involved.

    PubMed

    Rondas, Dieter; Bugliani, Marco; D'Hertog, Wannes; Lage, Kasper; Masini, Mathilde; Waelkens, Etienne; Marchetti, Piero; Mathieu, Chantal; Overbergh, Lut

    2013-09-06

    Glucagon-like peptide-1 (GLP-1) has been shown to protect pancreatic β-cells against cytokine-induced dysfunction and destruction. The mechanisms through which GLP-1 exerts its effects are complex and still poorly understood. The aim of this study was to analyze the protein expression profiles of human islets of Langerhans treated with cytokines (IL-1β and IFN-γ) in the presence or absence of GLP-1 by 2D difference gel electrophoresis and subsequent protein interaction network analysis to understand the molecular pathways involved in GLP-1-mediated β-cell protection. Co-incubation of cytokine-treated human islets with GLP-1 resulted in a marked protection of β-cells against cytokine-induced apoptosis and significantly attenuated cytokine-mediated inhibition of glucose-stimulated insulin secretion. The cytoprotective effects of GLP-1 coincided with substantial alterations in the protein expression profile of cytokine-treated human islets, illustrating a counteracting effect on proteins from different functional classes such as actin cytoskeleton, chaperones, metabolic proteins, and islet regenerating proteins. In summary, GLP-1 alters in an integrated manner protein networks in cytokine-exposed human islets while protecting them against cytokine-mediated cell death and dysfunction. These data illustrate the beneficial effects of GLP-1 on human islets under immune attack, leading to a better understanding of the underlying mechanisms involved, a prerequisite for improving therapies for diabetic patients.

  12. Cytokine receptor expression in human lymphoid tissue: analysis by fluorescence microscopy.

    PubMed

    Zola, H; Ridings, J; Weedon, H; Fusco, M; Byard, R W; Macardle, P J

    1995-08-01

    A highly-sensitive flourescence method, capable of detecting cytokine receptors present at low concentrations (around 100 molecules per cell) by flow cytometry, was adapted for use on tissue sections. This method was used to examine the expression of several cytokine receptors in lymphoid tissues. IL-2 receptors were distributed broadly, with higher concentrations in T cell areas. IL-1 receptor Type 1 was detected in T cell areas and in the follicular mantle, and was strongly expressed on vascular endothelium. IL-6 receptor was found at very low concentration, both within and outside germinal centres. The gp 130 molecule, which is involved in the functional receptor complex for IL-6 and several other cytokines, was present at higher concentrations, particularly in the germinal centre. Analysis of receptor expression in secondary lymphoid tissue provides evidence bearing on the physiological roles of cytokines, as these tissues contain cells at various stages of physiological activation located in well-defined functional zones.

  13. Contribution of human osteoblasts and macrophages to bone matrix degradation and proinflammatory cytokine release after exposure to abrasive endoprosthetic wear particles.

    PubMed

    Jonitz-Heincke, Anika; Lochner, Katrin; Schulze, Christoph; Pohle, Diana; Pustlauk, Wera; Hansmann, Doris; Bader, Rainer

    2016-08-01

    One of the major reasons for failure after total joint arthroplasty is aseptic loosening of the implant. At articulating surfaces, defined as the interface between implant and surrounding bone cement, wear particles can be generated and released into the periprosthetic tissue, resulting in inflammation and osteolysis. The aim of the present study was to evaluate the extent to which osteoblasts and macrophages are responsible for the osteolytic and inflammatory reactions following contact with generated wear particles from Ti‑6Al‑7Nb and Co‑28Cr‑6Mo hip stems. To this end, human osteoblasts and THP‑1 monocytic cells were incubated with the experimentally generated wear particles as well as reference particles (0.01 and 0.1 mg/ml) for 48 h under standard culture conditions. To evaluate the impact of these particles on the two cell types, the release of different bone matrix degrading matrix metalloproteinases (MMPs), tissue inhibitors of MMPs (TIMPs), and relevant cytokines were determined by multiplex enzyme‑linked immunosorbent assays. Following incubation with wear particles, human osteoblasts showed a significant upregulation of MMP1 and MMP8, whereas macrophages reacted with enhanced MMP3, MMP8 and MMP10 production. Moreover, the synthesis of TIMPs 1 and 2 was inhibited. The osteoblasts and macrophages also responded with modified expression of the inflammatory mediators interleukin (IL)‑6, IL‑8, monocyte chemoattractant protein‑1 and vascular endothelial growth factor. These results demonstrate that the release of wear particles affects the release of proinflammatory cytokines and has a negative impact on bone matrix formation during the first 48 h of particle exposure. Human osteoblasts are directly involved in the proinflammatory cascade of bone matrix degradation. The simultaneous activation and recruitment of monocytes/macrophages boosted osteolytic processes in the periprosthetic tissue. By the downregulation of TIMP production and the

  14. HIV-1 gp120 envelope glycoprotein determinants for cytokine burst in human monocytes

    PubMed Central

    Coutu, Mathieu; Prévost, Jérémie; Brassard, Nathalie; Peres, Adam; Stegen, Camille; Madrenas, Joaquín; Kaufmann, Daniel E.; Finzi, Andrés

    2017-01-01

    The first step of HIV infection involves the interaction of the gp120 envelope glycoprotein to its receptor CD4, mainly expressed on CD4+ T cells. Besides its role on HIV-1 entry, the gp120 has been shown to be involved in the production of IL-1, IL-6, CCL20 and other innate response cytokines by bystander, uninfected CD4+ T cells and monocytes. However, the gp120 determinants involved in these functions are not completely understood. Whether signalling leading to cytokine production is due to CD4 or other receptors is still unclear. Enhanced chemokine receptor binding and subsequent clustering receptors may lead to cytokine production. By using a comprehensive panel of gp120 mutants, here we show that CD4 binding is mandatory for cytokine outburst in monocytes. Our data suggest that targeting monocytes in HIV-infected patients might decrease systemic inflammation and the potential tissue injury associated with the production of inflammatory cytokines. Understanding how gp120 mediates a cytokine burst in monocytes might help develop new approaches to improve the chronic inflammation that persists in these patients despite effective suppression of viremia by antiretroviral therapy. PMID:28346521

  15. Protective effect of ixerisoside A against UVB-induced pro-inflammatory cytokine production in human keratinocytes.

    PubMed

    Kim, Sung-Bae; Kim, Ji-Eun; Kang, Ok-Hwa; Mun, Su-Hyun; Seo, Yun-Soo; Kang, Da-Hye; Yang, Da-Wun; Ryu, Shi-Yong; Lee, Young-Mi; Kwon, Dong-Yeul

    2015-05-01

    Human skin is the first line of defense for the protection of the internal organs of the body from different stimuli. Ultraviolet B (UVB), one of the harmful radiations for skin, is widely known to induce abnormally increased cytokine release from keratinocytes leading to inflammatory skin disorders. IL-6 and IL-8 induce an acute-phase response and stimulate leukocyte infiltration in the skin. Previous studies have shown that chronic exposure to UVB radiation increases cyclooxygenase-2 (COX‑2) expression through various cell signaling pathways, resulting in skin cancer. Recent studies have shown that the activation of extracellular signal-regulated kinase (ERK), c-Jun NH2-terminal kinase (JNK), and p38 MAPK is strongly correlated with acute inflammation and development of skin cancer caused by an increased expression of COX-2. Ixerisoside A (IXA) is an active constituent of Ixeris dentata of the Compositae (Asteraceae) family. The effect of IXA on skin inflammation has yet to be elucidated. To determine the anti-inflammatory effects of IXA, we examined its effect on UVB-induced pro-inflammatory cytokine production in human keratinocytes (HaCaT cells) by observing these cells in the presence or absence of IXA. In this study, pro-inflammatory cytokine production was determined by enzyme-linked immunosorbent assay (ELISA), reverse transcription-polymerase chain reaction (rt-pcr), and western blot analysis to evaluate the activation of mitogen-activated protein kinases (MAPKs). IXA inhibited UVB-induced production of the pro-inflammatory cytokines IL-6 and IL-8 in a dose-dependent manner. Moreover, IXA inhibited the expression of COX-2, ERK, JNK, and p38 MAPKs, indicating that the secretion of the pro-inflammatory cytokines IL-6 and IL-8, and COX-2 expression was inhibited by blocking MAPK phosphorylation. These results indicated that IXA potentially protects against UVB-induced skin inflammation.

  16. Suppression of Cytokine Release by Fluticasone Furoate vs. Mometasone Furoate in Human Nasal Tissue Ex-Vivo

    PubMed Central

    Zhang, Nan; Van Crombruggen, Koen; Holtappels, Gabriele; Lan, Feng; Katotomichelakis, Michail; Zhang, Luo; Högger, Petra; Bachert, Claus

    2014-01-01

    Background Topical glucocorticosteroids are the first line therapy for airway inflammation. Modern compounds with higher efficacy have been developed, but head-to-head comparison studies are sparse. Objective To compare the activity of two intranasal glucocorticoids, fluticasone furoate (FF) and mometasone furoate (MF) with respect to the inhibition of T helper (Th)1, Th2 and Th17 cytokine release in airway mucosa. Methods We used an ex-vivo human nasal mucosal tissue model and employed pre- and post- Staphylococcus aureus enterotoxin B (SEB)-challenge incubations with various time intervals and drug concentrations to mimic typical clinical situations of preventive or therapeutic use. Results At a fixed concentration of 10−10 M, FF had significantly higher suppressive effects on interferon (IFN)-γ, interleukin (IL)-2 and IL-17 release, but not IL-5 or tumor necrosis factor (TNF)-α, vs. MF. While the maximal suppressive activity was maintained when FF was added before or after tissue stimulation, the cytokine suppression capacity of MF appeared to be compromised when SEB-induced cell activation preceded the addition of the drug. In a pre-challenge incubation setting with removal of excess drug concentrations, MF approached inhibition of IL-5 and TNF-α after 6 and 24 hours while FF maximally blocked the release of these cytokines right after pre-incubation. Furthermore, FF suppressed a wider range of T helper cytokines compared to MF. Conclusion The study demonstrates the potential of our human mucosal model and shows marked differences in the ability to suppress the release of various cytokines in pre- and post-challenge settings between FF and MF mimicking typical clinical situations of preventive or therapeutic use. PMID:24710117

  17. Pro-inflammatory Macrophages Sustain Pyruvate Oxidation through Pyruvate Dehydrogenase for the Synthesis of Itaconate and to Enable Cytokine Expression.

    PubMed

    Meiser, Johannes; Krämer, Lisa; Sapcariu, Sean C; Battello, Nadia; Ghelfi, Jenny; D'Herouel, Aymeric Fouquier; Skupin, Alexander; Hiller, Karsten

    2016-02-19

    Upon stimulation with Th1 cytokines or bacterial lipopolysaccharides, resting macrophages shift their phenotype toward a pro-inflammatory state as part of the innate immune response. LPS-activated macrophages undergo profound metabolic changes to adapt to these new physiological requirements. One key step to mediate this metabolic adaptation is the stabilization of HIF1α, which leads to increased glycolysis and lactate release, as well as decreased oxygen consumption. HIF1 abundance can result in the induction of the gene encoding pyruvate dehydrogenase kinase 1 (PDK1), which inhibits pyruvate dehydrogenase (PDH) via phosphorylation. Therefore, it has been speculated that pyruvate oxidation through PDH is decreased in pro-inflammatory macrophages. However, to answer this open question, an in-depth analysis of this metabolic branching point was so far lacking. In this work, we applied stable isotope-assisted metabolomics techniques and demonstrate that pyruvate oxidation is maintained in mature pro-inflammatory macrophages. Glucose-derived pyruvate is oxidized via PDH to generate citrate in the mitochondria. Citrate is used for the synthesis of the antimicrobial metabolite itaconate and for lipogenesis. An increased demand for these metabolites decreases citrate oxidation through the tricarboxylic acid cycle, whereas increased glutamine uptake serves to replenish the TCA cycle. Furthermore, we found that the PDH flux is maintained by unchanged PDK1 abundance, despite the presence of HIF1. By pharmacological intervention, we demonstrate that the PDH flux is an important node for M(LPS) macrophage activation. Therefore, PDH represents a metabolic intervention point that might become a research target for translational medicine to treat chronic inflammatory diseases.

  18. Pro-inflammatory Macrophages Sustain Pyruvate Oxidation through Pyruvate Dehydrogenase for the Synthesis of Itaconate and to Enable Cytokine Expression*

    PubMed Central

    Meiser, Johannes; Krämer, Lisa; Sapcariu, Sean C.; Battello, Nadia; Ghelfi, Jenny; D'Herouel, Aymeric Fouquier; Skupin, Alexander; Hiller, Karsten

    2016-01-01

    Upon stimulation with Th1 cytokines or bacterial lipopolysaccharides, resting macrophages shift their phenotype toward a pro-inflammatory state as part of the innate immune response. LPS-activated macrophages undergo profound metabolic changes to adapt to these new physiological requirements. One key step to mediate this metabolic adaptation is the stabilization of HIF1α, which leads to increased glycolysis and lactate release, as well as decreased oxygen consumption. HIF1 abundance can result in the induction of the gene encoding pyruvate dehydrogenase kinase 1 (PDK1), which inhibits pyruvate dehydrogenase (PDH) via phosphorylation. Therefore, it has been speculated that pyruvate oxidation through PDH is decreased in pro-inflammatory macrophages. However, to answer this open question, an in-depth analysis of this metabolic branching point was so far lacking. In this work, we applied stable isotope-assisted metabolomics techniques and demonstrate that pyruvate oxidation is maintained in mature pro-inflammatory macrophages. Glucose-derived pyruvate is oxidized via PDH to generate citrate in the mitochondria. Citrate is used for the synthesis of the antimicrobial metabolite itaconate and for lipogenesis. An increased demand for these metabolites decreases citrate oxidation through the tricarboxylic acid cycle, whereas increased glutamine uptake serves to replenish the TCA cycle. Furthermore, we found that the PDH flux is maintained by unchanged PDK1 abundance, despite the presence of HIF1. By pharmacological intervention, we demonstrate that the PDH flux is an important node for M(LPS) macrophage activation. Therefore, PDH represents a metabolic intervention point that might become a research target for translational medicine to treat chronic inflammatory diseases. PMID:26679997

  19. Titanium, chromium and cobalt ions modulate the release of bone-associated cytokines by human monocytes/macrophages in vitro.

    PubMed

    Wang, J Y; Wicklund, B H; Gustilo, R B; Tsukayama, D T

    1996-12-01

    Osteolysis has become a major cause of aseptic loosening in total joint arthroplasty (TJA). Titanium, cobalt and chromium are commonly used in orthopaedic implants (e.g. joint prostheses). The release of bone-associated cytokines has been associated with the development of osteolysis in patients with prostheses. We evaluated the effects of these metals on the release of bone-associated cytokines (IL-1 beta, IL-6, TNF-alpha and TGF-beta 1) by human blood monocytes/macrophages and monocyte-like U937 cells upon lipopolysaccharide (LPS) stimulation, the cell proliferation, and their cytotoxic effects on these cells in vitro. We found that the release of IL-1 beta was enhanced by titanium, chromium and cobalt, the release of TNF-alpha was enhanced by titanium and chromium, and the release of IL-6 was enhanced by titanium. All three metal ions inhibited the release of TGF-beta 1. We also found that titanium and chromium, but not cobalt, enhanced blood monocyte/macrophage proliferation in response to LPS while only titanium enhanced U937 cell proliferation in response to LPS. The metals in concentrations ranging from 0.01 to 100 ngml-1 did not stimulate the cells to secrete detectable cytokines in the absence of LPS. Furthermore, a 4-h pre-exposure of blood monocytes/macrophages or U937 cells to the metals did not alter cytokine release when the metals were removed from the media prior to the addition of LPS. Similarly, a 4-h pre-exposure of blood monocytes/macrophages or U937 cells to LPS did not alter cytokine release when LPS was removed from the media prior to the addition of the metals. The metals did not reduce cell viability and induce cell injury after 72h incubation with the cells. The data suggest that the three metals at clinically relevant concentrations modulated cytokine expression, whereas they did not induce any cytotoxic effects. A metal-induced enhancement of bone-resorbing cytokine release with a concomitant inhibition of bone-forming cytokine release

  20. Anti-Fas/APO-1 antibody-mediated apoptosis of cultured human glioma cells. Induction and modulation of sensitivity by cytokines.

    PubMed Central

    Weller, M; Frei, K; Groscurth, P; Krammer, P H; Yonekawa, Y; Fontana, A

    1994-01-01

    Fas/APO-1 is a transmembrane protein of the nerve growth factor/TNF alpha receptor family which signals apoptotic cell death in susceptible target cells. We have investigated the susceptibility of seven human malignant glioma cell lines to Fas/APO-1-dependent apoptosis. Sensitivity to Fas/APO-1 antibody-mediated cell killing correlated with cell surface expression of Fas/APO-1. Expression of Fas/APO-1 as well as Fas/APO-1-dependent cytotoxicity were augmented by preexposure of human malignant glioma cells to IFN gamma and TNF alpha. Further, pretreatment with TGF beta 2, IL1 and IL8 enhanced Fas/APO-1 antibody-induced glioma cell apoptosis whereas other cytokines including TNF beta, IL6, macrophage colony-stimulating factor, IL10 and IL13 had no such effect. None of the human malignant glioma cell lines was susceptible to TNF alpha-induced cytotoxicity. Fas/APO-1 antibody-sensitive glioma cell lines (n = 5), but not Fas/APO-1 antibody-resistant glioma cell lines (n = 2), became sensitive to TNF alpha when co-treated with inhibitors of RNA and protein synthesis. Resistance of human glioma cells to Fas/APO-1 antibody-mediated apoptosis was mainly related to low level expression of Fas/APO-1 and appeared not to be linked to overexpression of the anti-apoptotic protooncogene, bcl-2. Given the resistance of human malignant glioma to surgery, irradiation, chemotherapy and immunotherapy, we propose that Fas/APO-1 may be a promising target for a novel locoregionary approach to human malignant glioma. This strategy gains support from the demonstration of Fas/APO-1 expression in ex vivo human malignant glioma specimens and from the absence of Fas/APO-1 in normal human brain parenchyma. Images PMID:7521890

  1. Proteolytic enzymes and amylase induce cytokine production in human peripheral blood mononuclear cells in vitro.

    PubMed

    Desser, L; Rehberger, A; Paukovits, W

    1994-01-01

    In vitro treatment of human peripheral blood mononuclear cells (PBMNC) with proteolytic enzymes (bromelain, papain) and amylase leads to the production of large amounts of tumor necrosis factor-alpha (TNF-alpha), interleukin-1-beta (IL-1 beta), and interleukin-6 (IL-6) in a time and dose dependent manner. Increased TNF-alpha and IL-6 production was already found after 4-6 hours of incubation, and plateau levels were reached after 12-16 hours. Plateau levels up to 1500 pg TNF-alpha/ml/10(6) PBMNC, 13000 pg IL-1 beta/ml/10(6) PBMNC, and 23000 pg IL-6/ml/10(6) PBMNC were observed. Control cultures contained below 35 pg/ml/10(6) PBMNC of TNF-alpha, IL-1 beta or IL-6. In contrast to TNF-alpha which was undetectable after more than 24 hours, peak levels of IL-1 beta and IL-6 were still present at 24 hours. After incubation of the enzyme solution for some hours at 56 degrees C the cytokine inducing capacity disappeared. Neutralization experiments with inactivating antibodies, radioimmunoassay, and western blotting after electrophoretic separation showed that the TNF-like activity found in the lytic assay was due to TNF-alpha. Interferon-alpha (IFN-alpha) and Interferon-gamma (IFN-gamma), which had no effect alone, synergistically increased TNF-alpha production when applied together with the enzymes. A commercial mixture of these enzymes (Wobenzym), which was also investigated, showed a similar concentration and time dependence, as well as synergism with the interferons. A synergistic effect on TNF-alpha production was also found with the enzymes and phorbol ester (PMA).

  2. Diminished Production of Monocyte Proinflammatory Cytokines during Human Immunodeficiency Virus Viremia Is Mediated by Type I Interferons▿ †

    PubMed Central

    Tilton, John C.; Johnson, Alison J.; Luskin, Marlise R.; Manion, Maura M.; Yang, Jun; Adelsberger, Joseph W.; Lempicki, Richard A.; Hallahan, Claire W.; McLaughlin, Mary; Mican, JoAnn M.; Metcalf, Julia A.; Iyasere, Christiana; Connors, Mark

    2006-01-01

    The effect of human immunodeficiency virus (HIV) infection and high-level HIV replication on the function of monocytes was investigated. HIV-positive patients had elevated levels of spontaneous production of some or all of the monocyte proinflammatory cytokines measured (interleukin-1β [IL-1β], IL-6, and tumor necrosis factor alpha [TNF-α]) compared to uninfected controls. In patients on therapy with high frequencies of monocytes producing proinflammatory cytokines, this frequency was diminished in the context of viremia during an interruption of therapy. Diminished production of proinflammatory cytokines during viremia was restored by culture with autologous CD4+ T cells or monocytes from an on-therapy time point or lipopolysaccharide (LPS). Microarray analysis demonstrated that diminished monocyte production of proinflammatory cytokines was correlated with elevated type I interferon-stimulated gene transcripts. The addition of exogenous alpha 2A interferon diminished the spontaneous production of IL-1β, IL-6, and TNF-α but did not affect responses to LPS, recapitulating the changes observed for HIV-viremic patients. These results suggest that monocyte function is diminished during high-level HIV viremia and that this effect is mediated by chronic stimulation by type I interferons. This effect on monocytes during viremia may play a role in diminished innate or adaptive immune system functions in HIV-infected patients. In addition, the restoration of these functions may also play a role in some immune reconstitution syndromes observed during initiation of therapy. PMID:17005663

  3. Analysis of complex biomarkers for human immune-mediated disorders based on cytokine responsiveness of peripheral blood cells123

    PubMed Central

    Davis, John M.; Knutson, Keith L.; Strausbauch, Michael A.; Crowson, Cynthia S.; Therneau, Terry M.; Wettstein, Peter J.; Matteson, Eric L.; Gabriel, Sherine E.

    2010-01-01

    The advent of improved biomarkers promises to enhance the clinical care for patients with rheumatoid arthritis (RA) and other immune-mediated disorders. We have developed an innovative approach to broadly assess the cytokine responsiveness of human PBMC using a multi-stimulant panel and multiplexed immunoassays. The objective of this study was to demonstrate this concept by determining whether cytokine profiles could discriminate RA patients according to disease stage (early vs. late) or severity. A 10-cytokine profile, consisting of IL-12, CCL4, TNFα, IL-4, and IL-10 release in response to stimulation with anti-CD3/anti-CD28, CXCL8 and IL-6 in response to CMV/EBV lysate, and IL-17A, GM-CSF, and CCL2 in response to HSP60, easily discriminated the early RA group from controls. These data were used to create an immune response score, which performed well in distinguishing the early RA patients from controls and also correlated with several markers of disease severity among the patients with late RA. In contrast, the same 10-cytokine profile assessed in serum was far less effective in discriminating the groups. Thus, our approach lays the foundation for the development of immunologic ‘signatures’ that could be useful in predicting disease course and monitoring the outcomes of therapy among patients with immune-mediated diseases. PMID:20495063

  4. Lung Infection by Human Bocavirus Induces the Release of Profibrotic Mediator Cytokines In Vivo and In Vitro

    PubMed Central

    Karagiannidis, Christian; Bayh, Inga; Brockmann, Michael; Pieper, Monika; Windisch, Wolfram; Schildgen, Oliver; Schildgen, Verena

    2016-01-01

    Human Bocavirus subtype 1 (HBoV1) is associated with respiratory diseases and may contribute to chronic lung diseases by persisting in the infected host. Here the question was addressed if HBoV infections could contribute to fibrogenesis processes as suggested by previously published clinical observations. Cytokine profiles induced by HBoV infection in CuFi-8 air-liquid interphase cell cultures and in bronchoalveolar lavage fluid (BALF) of 20 HBoV-positive and 12 HBoV-negative patients were analysed by semi-quantitative Western spot blot analyses. Although lots of cytokines were regulated independently of HBoV status, several cytokines associated with lung fibrosis and tumour development, e.g., EGF, VEGF, TARC (CCL17), TNF-α, TNF-β, TIMP-1, were clearly upregulated in the HBoV-positive cohort. These findings suggest that the development of lung fibrosis might be triggered by HBoV induced cytokine expression. PMID:26807786

  5. Low frequency pulsed electromagnetic field affects proliferation, tissue-specific gene expression, and cytokines release of human tendon cells.

    PubMed

    de Girolamo, L; Stanco, D; Galliera, E; Viganò, M; Colombini, A; Setti, S; Vianello, E; Corsi Romanelli, M M; Sansone, V

    2013-07-01

    Low frequency pulsed electromagnetic field (PEMF) has proven to be effective in the modulation of bone and cartilage tissue functional responsiveness, but its effect on tendon tissue and tendon cells (TCs) is still underinvestigated. PEMF treatment (1.5 mT, 75 Hz) was assessed on primary TCs, harvested from semitendinosus and gracilis tendons of eight patients, under different experimental conditions (4, 8, 12 h). Quantitative PCR analyses were conducted to identify the possible effect of PEMF on tendon-specific gene transcription (scleraxis, SCX and type I collagen, COL1A1); the release of pro- and anti-inflammatory cytokines and of vascular endothelial growth factor (VEGF) was also assessed. Our findings show that PEMF exposure is not cytotoxic and is able to stimulate TCs' proliferation. The increase of SCX and COL1A1 in PEMF-treated cells was positively correlated to the treatment length. The release of anti-inflammatory cytokines in TCs treated with PEMF for 8 and 12 h was significantly higher in comparison with untreated cells, while the production of pro-inflammatory cytokines was not affected. A dramatically higher increase of VEGF-A mRNA transcription and of its related protein was observed after PEMF exposure. Our data demonstrated that PEMF positively influence, in a dose-dependent manner, the proliferation, tendon-specific marker expression, and release of anti-inflammatory cytokines and angiogenic factor in a healthy human TCs culture model.

  6. Genetic toxicity of cytokines.

    PubMed

    Lazutka, J R

    1996-12-12

    Review of the literature shows that such cytokines as human interferons alpha and gamma, tumor necrosis factor alpha, epidermal growth factor and interleukin-2 may exhibit genotoxic properties in human peripheral blood lymphocyte cultures. For all above cytokines, except interleukin-2, parabolic-like relationship between the dose and the frequency of sister chromatid exchanges was found. Although the mechanisms of these genotoxic actions remain largely unknown, generation of free radicals or interaction with enzymes such as DNA topoisomerase II may be suspected. Human interferon alpha also may be considered as an antimutagenic compound in human cells. Human tumor necrosis factor alpha has been reported to enhance cytotoxicity and DNA fragmentation produced by DNA topoisomerase II-targeted anticancer drugs. At the same time, it has some radio- and chemoprotective properties in vitro and in vivo. Despite these facts, the question about genotoxicity of cytokines is not answered. Some problems must be resolved before receiving the final answer. First, much more cytokines must be tested for their genotoxic activity. Second, appropriate test-systems must be designed. Third, genotoxicity studies of cytokines must account for cytokine interaction in the cytokine network as well as for such cytokine-induced effects as cytotoxicity and apoptosis. Fourth, in each case, it is necessary to have experimental evidence that observed genotoxic effects were caused by cytokine under investigation and not by the other factors.

  7. Immunostimulatory Activity of the Cytokine-Based Biologic, IRX-2, on Human Papillomavirus-Exposed Langerhans Cells

    PubMed Central

    Da Silva, Diane M.; Woodham, Andrew W.; Naylor, Paul H.; Egan, James E.; Berinstein, Neil L.

    2016-01-01

    Langerhans cells (LCs) are the antigen-presenting cells of the epithelial layer and are responsible for initiating immune responses against skin and mucosa-invading viruses. Human papillomavirus (HPV)-mediated suppression of LC function is a crucial mechanism of HPV immune evasion, which can lead to persistent infection and development of several human cancers, including cervical, anal, and head and neck cancers. The cell-derived cytokine-based biologic, IRX-2, consists of multiple well-defined cytokines and is broadly active on various immune cell subsets. In this study, we investigated primary human LC activation after exposure to HPV16, followed by treatment with IRX-2 in vitro, and evaluated their subsequent ability to induce HPV16-specific T cells. In contrast to its activity on dendritic cells, HPV16 alone is not sufficient to induce phenotypic and functional activation of LCs. However, IRX-2 induces a significant upregulation of antigen presentation and costimulatory molecules, T helper 1 (Th1)-associated cytokine release, and chemokine-directed migration of LCs pre-exposed to HPV16. Furthermore, LCs treated with IRX-2 after HPV16 exposure induced CD8+ T-cell responses against specific HLA-A*0201-binding HPV16 T-cell epitopes. The present study suggests that IRX-2 is an attractive immunomodulator for assisting the immune response in eradication of HPV-infected cells, thereby potentially preventing HPV-induced cancers. PMID:26653678

  8. Combination of lipid metabolism alterations and their sensitivity to inflammatory cytokines in human lipin-1-deficient myoblasts.

    PubMed

    Michot, Caroline; Mamoune, Asmaa; Vamecq, Joseph; Viou, Mai Thao; Hsieh, Lu-Sheng; Testet, Eric; Lainé, Jeanne; Hubert, Laurence; Dessein, Anne-Frédérique; Fontaine, Monique; Ottolenghi, Chris; Fouillen, Laetitia; Nadra, Karim; Blanc, Etienne; Bastin, Jean; Candon, Sophie; Pende, Mario; Munnich, Arnold; Smahi, Asma; Djouadi, Fatima; Carman, George M; Romero, Norma; de Keyzer, Yves; de Lonlay, Pascale

    2013-12-01

    Lipin-1 deficiency is associated with massive rhabdomyolysis episodes in humans, precipitated by febrile illnesses. Despite well-known roles of lipin-1 in lipid biosynthesis and transcriptional regulation, the pathogenic mechanisms leading to rhabdomyolysis remain unknown. Here we show that primary myoblasts from lipin-1-deficient patients exhibit a dramatic decrease in LPIN1 expression and phosphatidic acid phosphatase 1 activity, and a significant accumulation of lipid droplets (LD). The expression levels of LPIN1-target genes [peroxisome proliferator-activated receptors delta and alpha (PPARδ, PPARα), peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α), acyl-coenzyme A dehydrogenase, very long (ACADVL), carnitine palmitoyltransferase IB and 2 (CPT1B and CPT2)] were not affected while lipin-2 protein level, a closely related member of the family, was increased. Microarray analysis of patients' myotubes identified 19 down-regulated and 51 up-regulated genes, indicating pleiotropic effects of lipin-1 deficiency. Special attention was paid to the up-regulated ACACB (acetyl-CoA carboxylase beta), a key enzyme in the fatty acid synthesis/oxidation balance. We demonstrated that overexpression of ACACB was associated with free fatty acid accumulation in patients' myoblasts whereas malonyl-carnitine (as a measure of malonyl-CoA) and CPT1 activity were in the normal range in basal conditions accordingly to the normal daily activity reported by the patients. Remarkably ACACB invalidation in patients' myoblasts decreased LD number and size while LPIN1 invalidation in controls induced LD accumulation. Further, pro-inflammatory treatments tumor necrosis factor alpha+Interleukin-1beta(TNF1α+IL-1ß) designed to mimic febrile illness, resulted in increased malonyl-carnitine levels, reduced CPT1 activity and enhanced LD accumulation, a phenomenon reversed by dexamethasone and TNFα or IL-1ß inhibitors. Our data suggest that the pathogenic mechanism

  9. Combination of lipid metabolism alterations and their sensitivity to inflammatory cytokines in human lipin-1-deficient myoblasts

    PubMed Central

    Michot, Caroline; Mamoune, Asmaa; Vamecq, Joseph; Viou, Mai Thao; Hsieh, Lu-Sheng; Testet, Eric; Lainé, Jeanne; Hubert, Laurence; Dessein, Anne-Frédérique; Fontaine, Monique; Ottolenghi, Chris; Fouillen, Laetitia; Nadra, Karim; Blanc, Etienne; Bastin, Jean; Candon, Sophie; Pende, Mario; Munnich, Arnold; Smahi, Asma; Djouadi, Fatima; Carman, George M.; Romero, Norma; de Keyzer, Yves; de Lonlay, Pascale

    2014-01-01

    Lipin-1 deficiency is associated with massive rhabdomyolysis episodes in humans, precipitated by febrile illnesses. Despite well-known roles of lipin-1 in lipid biosynthesis and transcriptional regulation, the pathogenic mechanisms leading to rhabdomyolysis remain unknown. Here we show that primary myoblasts from lipin-1-deficient patients exhibit a dramatic decrease in LPIN1 expression and phosphatidic acid phosphatase 1 activity, and a significant accumulation of lipid droplets (LD). The expression levels of LPIN1-target genes [peroxisome proliferator-activated receptors delta and alpha (PPARδ, PPARα), peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α), acyl-coenzyme A dehydrogenase, very long (ACADVL), carnitine palmitoyltransferase IB and 2 (CPT1B and CPT2)] were not affected while lipin-2 protein level, a closely related member of the family, was increased. Microarray analysis of patients’ myotubes identified 19 down-regulated and 51 up-regulated genes, indicating pleiotropic effects of lipin-1 deficiency. Special attention was paid to the up-regulated ACACB (acetyl-CoA carboxylase beta), a key enzyme in the fatty acid synthesis/oxidation balance. We demonstrated that overexpression of ACACB was associated with free fatty acid accumulation in patients’ myoblasts whereas malonyl-carnitine (as a measure of malonyl-CoA) and CPT1 activity were in the normal range in basal conditions accordingly to the normal daily activity reported by the patients. Remarkably ACACB invalidation in patients’ myoblasts decreased LD number and size while LPIN1 invalidation in controls induced LD accumulation. Further, pro-inflammatory treatments tumor necrosis factor alpha + Interleukin-1beta(TNF1α + IL-1β) designed to mimic febrile illness, resulted in increased malonyl-carnitine levels, reduced CPT1 activity and enhanced LD accumulation, a phenomenon reversed by dexamethasone and TNFα or IL-1β inhibitors. Our data suggest that the pathogenic

  10. Cytokines and feeding.

    PubMed

    Plata-Salamán, C R

    2001-12-01

    Various categories of cytokines participate in the control of feeding, including interleukin-1 and -6 and other activators of gp 130, leptin (ob protein), interleukin-8 and other chemokines, tumor necrosis factor-alpha, and interferon-alpha. These feeding-inhibitory cytokines may play a role in the regulation of food intake during physiological (eg a role proposed for leptin) and pathophysiological (eg proinflammatory cytokines) conditions. Data show that various cytokines participate in acute and chronic disease-associated anorexia such as during infection, inflammation or malignancy. Food intake suppression (reported as anorexia) is also a common central manifestation observed during cytokine immunotherapy in humans. The concept of local production of various cytokines within specific brain regions in response to peripheral challenges and pathophysiological processes has broad implications for the interpretation of brain cytokines as mediators or participants in CNS modulation of feeding and anorexia.

  11. Early cytokine signatures of ischemia/reperfusion injury in human orthotopic liver transplantation

    PubMed Central

    Sosa, Rebecca A.; Zarrinpar, Ali; Lassman, Charles R.; Naini, Bita V.; Datta, Nakul; Rao, Ping; Harre, Nicholas; Zheng, Ying; Hoffmann, Alexander; Busuttil, Ronald W.; Gjertson, David W.; Zhai, Yuan; Kupiec-Weglinski, Jerzy W.; Reed, Elaine F.

    2016-01-01

    BACKGROUND. Orthotopic liver transplant (OLT) is the primary therapy for end-stage liver disease and acute liver failure. However, ischemia/reperfusion injury (IRI) can severely compromise allograft survival. To understand the evolution of immune responses underlying OLT-IRI, we evaluated longitudinal cytokine expression profiles from adult OLT recipients before transplant through 1 month after transplant. METHODS. We measured the expression of 38 cytokines, chemokines, and growth factors in preoperative and postoperative recipient circulating systemic blood (before transplant and 1 day, 1 week, and 1 month after transplant) and intraoperative portal blood (before and after reperfusion) of 53 OLT patients and analyzed this expression in relation to biopsy-proven IRI (n = 26 IRI+; 27 IRI–), clinical liver function tests early (days 1–7) after transplant, and expression of genes encoding cytokine receptors in biopsies of donor allograft taken before and after reperfusion. RESULTS. Bilirubin and arginine transaminase levels early after transplant correlated with IRI. Fourteen cytokines were significantly increased in the systemic and/or portal blood of IRI+ recipients that shifted from innate to adaptive-immune responses over time. Additionally, expression of cognate receptors for 10 of these cytokines was detected in donor organ biopsies by RNAseq. CONCLUSION. These results provide a mechanistic roadmap of the early immunological events both before and after IRI and suggest several candidates for patient stratification, monitoring, and treatment. FUNDING. Ruth L. Kirschstein National Research Service Award T32CA009120, Keck Foundation award 986722, and a Quantitative & Computational Biosciences Collaboratory Postdoctoral Fellowship. PMID:27942590

  12. Dissimilar cytokine patterns in different human liver and colon cancer cell lines.

    PubMed

    Guerriero, Eliana; Capone, Francesca; Rusolo, Fabiola; Colonna, Giovanni; Castello, Giuseppe; Costantini, Susan

    2013-11-01

    An accurate and simultaneous estimate of cellular levels of a large cytokine number is very useful to obtain information about an organ dysfunction leading to cancer because through the understanding of the evolution of cytokine patterns we can recognize and predict the disease progression. Cancer cell lines are commonly used to study the cancer microenvironment, to analyze their chemosensitivity and carcinogenesis as well as to test in vitro the effect of molecules, such as drugs or anti-oxidants, on the inflammation status and its progression. We noted that various cell lines commonly used as a model for studies on liver and colon cancer possess different patterns of cytokines. This aspect may generate data not comparable in laboratories using different cell lines; thus, to investigate the origin of these abnormalities we compared the cell lines HepG2 and Huh7, and HT-29 and HCT-116, for liver and colon cancer, respectively. In this context we have evaluated and compared the levels of cytokines, chemokines and growth factors in the supernatants of these cellular lines. Our aim was to identify what cytokines were significantly different correlating similarities and differences to the specific inflammation status of each cellular model of cancer. Copyright © 2013 Elsevier Ltd. All rights reserved.

  13. Activated platelets signal chemokine synthesis by human monocytes.

    PubMed Central

    Weyrich, A S; Elstad, M R; McEver, R P; McIntyre, T M; Moore, K L; Morrissey, J H; Prescott, S M; Zimmerman, G A

    1996-01-01

    Human blood monocytes adhere rapidly and for prolonged periods to activated platelets that display P-selectin, an adhesion protein that recognizes a specific ligand on leukocytes, P-selectin glycoprotein-1. We previously demonstrated that P-selectin regulates expression and secretion of cytokines by stimulated monocytes when it is presented in a purified, immobilized form or by transfected cells. Here we show that thrombin-activated platelets induce the expression and secretion of monocyte chemotactic protein-1 and IL-8 by monocytes. Enhanced monokine synthesis requires engagement of P-selectin glycoprotein-1 on the leukocyte by P-selectin on the platelet. Secretion of the chemokines is not, however, directly signaled by P-selectin; instead, tethering of the monocytes by P-selectin is required for their activation by RANTES (regulated upon activation normal T cell expressed presumed secreted), a platelet chemokine not previously known to induce immediate-early gene products in monocytes. Adhesion of monocytes to activated platelets results in nuclear translocation of p65 (RelA), a component of the NF-kappaB family of transcription factors that binds kappaB sequences in the regulatory regions of monocyte chemotactic protein-1, IL-8, and other immediate-early genes. However, expression of tissue factor, a coagulation protein that also has a kappaB sequence in the 5' regulatory region of its gene, is not induced in monocytes adherent to activated platelets. Thus, contact of monocytes with activated platelets differentially affects the expression of monocyte products. These experiments suggest that activated platelets regulate chemokine secretion by monocytes in inflammatory lesions in vivo and provide a model for the study of gene regulation in cell-cell interactions. PMID:8617886

  14. PCR phenotyping of cytokines, growth factors and their receptors and bone matrix proteins in human osteoblast-like cell lines.

    PubMed

    Bilbe, G; Roberts, E; Birch, M; Evans, D B

    1996-11-01

    The expression of a total of 58 cytokines, growth factors, and their corresponding receptors and bone matrix proteins was assessed using reverse transcription-linked polymerase chain reaction (RT-PCR) analysis to determine the similarity in the expression profile between clonal osteosarcoma-derived human osteoblast-like cell lines and primary human osteoblast-like cell cultures derived from human trabecular bone explants. The spectrum of cytokines, growth factors, and bone-related proteins expressed by three human osteosarcoma-derived cell lines, TE-85, MG-63, SaOS-2, and primary human osteoblast-like cells was found to be highly comparable and for the first time the expression of EGF, ECGF, FGF beta, oncostatin M, TNF beta, and SCF by human osteoblast-like cells was detected. Also the expression of several receptor types including IL-4R, IL-7R, IFN alpha/beta R, and SCFR was detected that has not been previously described for human osteoblast-like cells. For the factors examined, no qualitative variations in the expression profile were observed in the six primary human osteoblast-like cell cultures used in this study. Of the 58 factors examined, only 13 showed some degree of nonuniformity of expression between all of the three cell lines and primary cell cultures. These differences were seen especially in the expression of cytokine receptor mRNA and to a lesser extent with some cytokines. Differences in receptor expression would suggest that the possible spectrum of response to exogenously added factors, or even autocrine/ paracrine networks would be determined by the repertoire of receptors expressed by each cell type. Whether the differences are related to the status of cell maturation within the osteoblast development lineage or to their abberant regulation of expression cannot be concluded at this stage. However, this PCR-phenotyping approach rapidly provides a resource of information, which can be subsequently used for further in depth studies to facilitate

  15. CD163 and CD206 expression does not correlate with tolerance and cytokine production in LPS-tolerant human monocytes.

    PubMed

    Alves-Januzzi, Amanda Barba; Brunialti, Milena Karina Colo; Salomao, Reinaldo

    2017-05-01

    Lipopolysaccharide (LPS)-tolerant monocytes produce small amounts of inflammatory cytokines, which is one of the characteristics of the alternative activated macrophages (AAM). These cells exhibited an increased expression of CD206 and CD163. Given the functional similarities of AAMs with the modulation of monocytes' functions observed during sepsis and LPS-tolerance, we evaluated whether the inhibition of inflammatory cytokine production by LPS-tolerant monocytes is associated with the phenotype of cells expressing CD206 and CD163. We investigated whether tolerant human monocytes would modulate their expression of CD206 and CD163, markers of alternative activation, and whether the level of their expression would be related to cytokines detection. Tolerance to LPS was induced in peripheral blood mononuclear cell by pre-incubating the cells with increasing concentrations of LPS. The expression of CD206 and CD163 and intracellular TNF-α and IL-6 was determined 24 h after LPS challenge by flow cytometry. No differences in CD163 expression were observed between tolerant and non-tolerant cells, while the expression of CD206, which was decreased following LPS stimulation in non-tolerized cells, was further reduced in tolerant cells. Decreased production of inflammatory cytokines was observed in the tolerized cells, regardless of the expression of CD163 and CD206, with the exception of IL-6 in CD206+ monocytes, which was similarly expressed in both tolerized and non-tolerized cells. The effect of LPS in the expression of CD163 and CD206 on monocytes is not reverted in LPS tolerant cells, and the inhibition of inflammatory cytokines in tolerant cells is not related with modulation of these receptors. © 2016 International Clinical Cytometry Society. © 2016 International Clinical Cytometry Society.

  16. Surface layer proteins from Clostridium difficile induce inflammatory and regulatory cytokines in human monocytes and dendritic cells.

    PubMed

    Ausiello, Clara Maria; Cerquetti, Marina; Fedele, Giorgio; Spensieri, Fabiana; Palazzo, Raffaella; Nasso, Maria; Frezza, Simona; Mastrantonio, Paola

    2006-09-01

    Clostridium difficile, an etiological agent of most cases of antibiotic-associated diarrhea, exerts its pathological action mainly by the activity of toxin A and toxin B. Less known is the role that S-layer proteins (SLPs), predominant surface components of the bacterium, may play in pathogenesis. Here, we evaluate the ability of SLPs to modulate the function of human monocytes and dendritic cells (DC) and to induce inflammatory and regulatory cytokines, influencing the natural and adaptive immune response. To this aim, SLPs were extracted from the clinical isolate C253 and characterized for their effects on immune cells. SLPs induced the release of elevated amounts of interleukin (IL)-1beta and IL-6 pro-inflammatory cytokines by resting monocytes, induced maturation of human monocyte-derived DC (MDDC), and enhanced proliferation of allogeneic T cells. C253-SLP-treated MDDC also secreted large amounts of IL-10 and IL-12p70 and induced a mixed Th1/Th2 orientation of immune response in naïve CD4 T cells. In conclusion, C. difficile SLPs may contribute to the pathogenicity of the bacterium by perturbing the fine balance of inflammatory and regulatory cytokines. These data are of interest also in the light of the possible use of SLPs in a multicomponent vaccine against C. difficile infections for high-risk patients.

  17. 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.

  18. Role of cytosolic phospholipase A2 in cytokine-stimulated prostaglandin release by human gallbladder cells.

    PubMed

    Grossmann, E M; Longo, W E; Mazuski, J E; Panesar, N; Kaminski, D L

    2000-01-01

    Eicosanoids are involved in gallbladder inflammation, epithelial water transport, and mucous secretion. Phospholipase Asubscript2 enzymes liberate arachidonic acid from membrane phospholipids for the synthesis of eicosanoids. The purpose of this study was to determine the effect of selective cytoplasmic and secretory phospholipase A2 inhibitors on basal and stimulated arachidonic acid and prostaglandin E2 release in gallbladder cells. Western immunoblotting was employed to evaluate both cytosolic and secretory phospholipase A2 enzymes in human gallbladder cells. Cells were incubated for 22 hours with (3)H-labeled arachidonic acid. Arachidonic acid and prostaglandin E2 release was then measured in the supernate after 2 hours of exposure to human interleukin-1beta, alone or after pretreatment for 1 hour with the inhibitors. Unstimulated gallbladder cells express both 85 kDa cytosolic and 14 kDa secretory phospholipase A2++. The 85 kDa phospholipase A2 was induced by interleukin-1beta, whereas there was no apparent change in secretory phospholipase A2 enzyme concentrations. Both the secretory phospholipase A2 inhibitor p-bromophenylacyl bromide and the cytosolic phospholipase A2 inhibitor arachidonyl trifluoromethyl ketone decreased basal and interleukin-1beta-stimulated arachidonic acid release. In contrast, only inhibition of cytosolic phospholipase A2 led to a decrease in interleukin-1beta-stimulated prostaglandin E2 release. Basal and interleukin-1beta-stimulated arachidonic acid release appears to be the result of the activity of both cytosolic and secretory phospholipase A2. Interleukin-1beta-stimulated prostaglandin E2 release appears to be dependent on the activity of cytosolic phospholipase A2.

  19. Cytokine Effects on Gap Junction Communication and Connexin Expression in Human Bladder Smooth Muscle Cells and Suburothelial Myofibroblasts

    PubMed Central

    Heinrich, Marco; Oberbach, Andreas; Schlichting, Nadine; Stolzenburg, Jens-Uwe; Neuhaus, Jochen

    2011-01-01

    Background The last decade identified cytokines as one group of major local cell signaling molecules related to bladder dysfunction like interstitial cystitis (IC) and overactive bladder syndrome (OAB). Gap junctional intercellular communication (GJIC) is essential for the coordination of normal bladder function and has been found to be altered in bladder dysfunction. Connexin (Cx) 43 and Cx45 are the most important gap junction proteins in bladder smooth muscle cells (hBSMC) and suburothelial myofibroblasts (hsMF). Modulation of connexin expression by cytokines has been demonstrated in various tissues. Therefore, we investigate the effect of interleukin (IL) 4, IL6, IL10, tumor necrosis factor-alpha (TNFα) and transforming growth factor-beta1 (TGFβ1) on GJIC, and Cx43 and Cx45 expression in cultured human bladder smooth muscle cells (hBSMC) and human suburothelial myofibroblasts (hsMF). Methodology/Principal Findings HBSMC and hsMF cultures were set up from bladder tissue of patients undergoing cystectomy. In cytokine stimulated cultured hBSMC and hsMF GJIC was analyzed via Fluorescence Recovery after Photo-bleaching (FRAP). Cx43 and Cx45 expression was assessed by quantitative PCR and confocal immunofluorescence. Membrane protein fraction of Cx43 and Cx45 was quantified by Dot Blot. Upregulation of cell-cell-communication was found after IL6 stimulation in both cell types. In hBSMC IL4 and TGFβ1 decreased both, GJIC and Cx43 protein expression, while TNFα did not alter communication in FRAP-experiments but increased Cx43 expression. GJ plaques size correlated with coupling efficacy measured, while Cx45 expression did not correlate with modulation of GJIC. Conclusions/Significance Our finding of specific cytokine effects on GJIC support the notion that cytokines play a pivotal role for pathophysiology of OAB and IC. Interestingly, the effects were independent from the classical definition of pro- and antiinflammatory cytokines. We conclude, that connexin

  20. Regulation of muscle protein synthesis in humans.

    PubMed

    Phillips, Bethan E; Hill, Derek S; Atherton, Philip J

    2012-01-01

    Investigations into the regulation of muscle protein synthesis (MPS) are a cornerstone of understanding the control of muscle mass. Rates of MPS are finely tuned according to levels of activity, nutrient availability and health status. For instance, rates of MPS are positively regulated by exercise and nutrition, and negatively regulated by inactivity (e.g. disuse), ageing (i.e. sarcopenia) and in muscle-wasting related diseases (e.g. cancer). Skeletal muscles display a high degree of intrinsic regulation. Increases in MPS after exercise occur independently of the systemic milieu for example growth hormone/testosterone concentrations. In the absence of exercise, increases in MPS after feeding are of finite duration despite enduring precursor availability; that is muscles can sense they are 'full'. Intriguingly, exercise delays this 'muscle-full' response to allow for building and repair. In contrast, muscle-wasting conditions exhibit a premature 'muscle-full' response to nutrition and exercise (i.e. anabolic resistance), which may cause atrophy. Observations of 'dissociations' between MPS and anabolic signalling pathways have cast doubt on how much we understand of the molecular regulation of human MPS. Anabolic and anticatabolic interventions in health and disease should be aimed at manipulating the 'muscle-full' set point to maximize muscle maintenance/hypertrophy.

  1. Extrapituitary growth hormone synthesis in humans.

    PubMed

    Pérez-Ibave, Diana Cristina; Rodríguez-Sánchez, Iram Pablo; Garza-Rodríguez, María de Lourdes; Barrera-Saldaña, Hugo Alberto

    2014-01-01

    The gene for pituitary growth hormone (GH-N) in man belongs to a multigene locus located at chromosome 17q24.2, which also harbors four additional genes: one for a placental variant of GH-N (named GH-V) and three of chorionic somatommamotropin (CSH) type. Their tandem arrangement from 5' to 3' is: GH-N, CSH-L, CSH-1, GH-V and CSH-2. GH-N is mainly expressed in the pituitary from birth throughout life, while the remaining genes are expressed in the placenta of pregnant women. Pituitary somatotrophs secrete GH into the bloodstream to act at receptor sites in most tissues. GH participates in the regulation of several complex physiological processes, including growth and metabolism. Recently, the presence of GH has been described in several extrapituitary sites, such as neural, ocular, reproductive, immune, cardiovascular, muscular, dermal and skeletal tissues. It has been proposed that GH has an autocrine action in these tissues. While the body of evidence for its presence is constantly growing, research of its possible function and implications lag behind. In this review we highlight the evidence of extrapituitary synthesis of GH in humans.

  2. Human TNF cytokine neutralization with a vNAR from Heterodontus francisci shark: a potential therapeutic use.

    PubMed

    Camacho-Villegas, Tanya; Mata-Gonzalez, Teresa; Paniagua-Solis, Jorge; Sanchez, Edna; Licea, Alexei

    2013-01-01

    The therapeutic use of single domain antibodies (sdAbs) is a promising new approach because these small antibodies maintain antigen recognition and neutralization capacity, have thermal and chemical stability and have good solubility. In this study, using phage display technology, we isolated a variable domain of a IgNAR (vNAR) from a Heterodontus francisci shark immunized against the recombinant human cytokine TNFα (rhTNFα). One clone T43, which expresses the vNAR protein in the periplasmic space, was isolated from the fourth round of panning. T43 had the capacity to recognize rhTNF and neutralize it in vitro, indicating that T43 has potential as a therapeutic that can be used for diseases in which this pro-inflammatory cytokine needs to be controlled.

  3. Effects of Japanese mistletoe lectin on cytokine gene expression in human colonic carcinoma cells and in the mouse intestine.

    PubMed

    Monira, Pervin; Koyama, Yu; Fukutomi, Ryuuta; Yasui, Kensuke; Isemura, Mamoru; Yokogoshi, Hidehiko

    2009-10-01

    Mistletoe lectins have various biological activities including anti-cancer and immunomodulatory effects. We previously isolated a lectin (ML-J) from Japanese mistletoe. In the present study, we examined the effects of ML-J on cytokine gene expression in human colon adenocarcinoma Caco-2 cells and in the mouse intestine. The results of reverse transcription-polymerase chain reaction and quantitative real-time polymerase chain reaction indicated that ML-J caused an upregulation of the gene expression of the proinflammatory cytokines interleukin (IL)-8, tumor necrosis factor-alpha (TNF-alpha) and IL-6 in Caco-2 cells and TNF-alpha and IL-6 in the duodenum. This study provides the first example to show that a perorally administered plant lectin affects gene expression in the duodenum.

  4. Sulfur mustard primes human neutrophils for increased degranulation and stimulates cytokine release via TRPM2/p38 MAPK signaling

    SciTech Connect

    Ham, Hwa-Yong; Hong, Chang-Won; Lee, Si-Nae; Kwon, Min-Soo; Kim, Yeon-Ja; Song, Dong-Keun

    2012-01-01

    Sulfur mustard (2,2′-bis-chloroethyl-sulfide; SM) has been a military threat since the World War I. The emerging threat of bioterrorism makes SM a major threat not only to military but also to civilian world. SM injury elicits an inflammatory response characterized by infiltration of neutrophils. Although SM was reported to prime neutrophils, the mechanism has not been identified yet. In the present study, we investigated the mechanism of SM-induced priming in human neutrophils. SM increased [Ca{sup 2+}]{sub i} in human neutrophils in a concentration-dependent fashion. Transient receptor potential melastatin (TRPM) 2 inhibitors (clotrimazole, econazole and flufenamic acid) and silencing of TRPM2 by shRNA attenuated SM-induced [Ca{sup 2+}]{sub i} increase. SM primed degranulation of azurophil and specific granules in response to activation by fMLP as previously reported. SB203580, an inhibitor of p38 MAPK, inhibited SM-induced priming. Neither PD98057, an ERK inhibitor, nor SP600215, a JNK inhibitor, inhibited SM-induced priming. In addition, SM enhanced phosphorylation of NF-kB p65 and release of TNF-α, interleukin (IL)-6 and IL-8. SB203580 inhibited SM-induced NF-kB phosphorylation and cytokine release. These results suggest the involvement of TRPM2/p38 MAPK pathway in SM-induced priming and cytokines release in neutrophils. -- Highlights: ► SM increased [Ca{sup 2+}]{sub i} in human neutrophils through TPRM2-mediated calcium influx. ► SM primed degranulation of azurophil and specific granules. ► SM enhanced p38 MAPK and NF-κB p65 phosphorylation in human neutrophils. ► SM enhanced release of TNF-α, interleukin (IL)-6 and IL-8 from human neutrophils. ► SB203580 inhibited SM-induced priming, NF-κB p65 phosphorylation and cytokine release.

  5. Regulation of cytokine production in human gingival fibroblasts following treatment with nicotine and lipopolysaccharide.

    PubMed

    Wendell, K J; Stein, S H

    2001-08-01

    Patients who smoke are at increased risk for chronic periodontitis (CP). Most studies suggest that the microbial flora in these patients is similar to that found in non-smoking CP patients. Thus, the increased risk for development of CP is not dependent on an altered microbial profile, but rather to some change in the host response to these periopathogens. There is evidence that human gingival fibroblasts (HGF) derived from diseased sites produce greater amounts of interleukin (IL)-6 and IL-8 in vitro than cells derived from healthy sites. This suggests that HGF subpopulations may be selected based upon the inflammatory milieu in which they reside. The hypothesis to be tested was that the combination of nicotine and lipopolysaccharide (LPS) could regulate HGF inflammatory mediator production. HGF cell cultures were established from explants derived from 10 patients with CP. HGF cell cultures were stimulated with 1 mM, 1 microM, or 1 nM nicotine +/- Escherichia coli or Porphyromonas gingivalis LPS. At 12, 24, or 48-hour time points, the cells were counted and the supernatant was collected for subsequent IL-6 and IL-8 determination in an enzyme-linked immunosorbent assay. At the 24-hour time point, 1 nM nicotine stimulated IL-6 production compared to control (P=0.02). E. coli LPS alone caused a 3- to 4-fold increase in IL-6 and IL-8 production, whereas P gingivalis LPS did not augment IL-6 or IL-8. A synergistic effect upregulating IL-6 was observed with combined treatment of 1 mM nicotine and E. coli LPS or P gingivalis LPS at the 24-hour time point (P<0.0005 and P=0.002, respectively). Similar effects were seen when IL-8 production was evaluated following HGF stimulation with high doses of nicotine and E. coli LPS or P gingivalis LPS. These results demonstrate that nicotine by itself can stimulate HGF IL-6 and IL-8 production. Moreover, the combination of high doses of nicotine and either E. coli or P gingivalis LPS can synergistically upregulate cytokine

  6. Impact of elvitegravir on human adipocytes: Alterations in differentiation, gene expression and release of adipokines and cytokines.

    PubMed

    Moure, Ricardo; Domingo, Pere; Gallego-Escuredo, José M; Villarroya, Joan; Gutierrez, Maria Del Mar; Mateo, Maria G; Domingo, Joan C; Giralt, Marta; Villarroya, Francesc

    2016-08-01

    Elvitegravir is a recently developed integrase inhibitor used for antiretroviral treatment of HIV infection. Secondary effects, including disturbances in lipid metabolism and, ultimately, in adipose tissue distribution and function, are common concerns associated with antiretroviral treatments. Here, we provide the first study of the effects of elvitegravir (in comparison with efavirenz, a non-nucleoside analog inhibitor of reverse transcriptase; and raltegravir, another integrase inhibitor) on human adipocyte differentiation, gene expression and secretion of adipokines and cytokines. Elvitegravir impaired adipogenesis and adipocyte metabolism in human SGBS adipocytes in a concentration-dependent manner (delaying acquisition of adipocyte morphology and reducing the expression of adipogenesis marker genes such as PPARγ, glucose transporter GLUT4, lipoprotein lipase, and the adipokines adiponectin and leptin). Compared with efavirenz, the effects of elvitegravir were similar but tended to occur at higher concentrations than those elicited by efavirenz, or were somewhat less intense than those caused by efavirenz at similar concentration. Elvitegravir tended to cause a more moderate induction of pro-inflammatory cytokines than efavirenz. Efavirenz induced a marked concentration-dependent increase in interleukin-8 expression and release whereas elvitregravir had little effect. Raltegravir had totally neutral actions of adipogenesis, adipocyte metabolism-related gene expression and release of adipokines and cytokines. In conclusion, elvitegravir alters adipocyte differentiation and function and promotes induction of pro-inflammatory cytokines similarly to efavirenz, but several effects were less intense. Further assessment of lipid metabolism and adipose tissue function in patients administered elvitegravir-based regimes is advisable considering that totally neutral effects of elvitegravir on lipid homeostasis cannot be anticipated from the current study in vitro.

  7. Pimecrolimus inhibits up-regulation of OX40 and synthesis of inflammatory cytokines upon secondary T cell activation by allogeneic dendritic cells

    PubMed Central

    KALTHOFF, F S; CHUNG, J; STUETZ, A

    2002-01-01

    Pimecrolimus is a new non-steroidal inhibitor of T cell and mast cell activation. In the present study, we compared the potency of pimecrolimus and cyclosporin A (CyA) to inhibit cytokine synthesis of alloantigen-primed T cells and the expression of CD134 (OX40), an inducible co-receptor molecule thought to be critical for the survival and expansion of inflammation-mediating T cells. To mimic the physiological situation of recurrent antigenic stimulation, we have used dendritic cells (DC) as stimulators of purified CD4+ T cells in the primary and secondary allogeneic mixed lymphocyte culture (allo-MLC). Pimecrolimus inhibited surface expression of OX40 and prevented the up-regulation of CD25 and CD54 with a 10-fold higher potency compared to CyA. Similarly, 50% inhibition of allo-DC-mediated T cell proliferation by pimecrolimus was obtained at 0·55 nm, compared to about 12 nm for CyA. Furthermore, pimecrolimus blocked the increase of OX40 on primed T cells restimulated on day 10 in secondary allo-MLC. Allo-DC-primed T cells showed a restricted cytokine profile characterized by the production of TNF-α, IFN-γ and IL-2 but low to undetectable levels of IL-4 and IL-10. The synthesis of TNF-α and IFN-γ and the up-regulation of OX40 on T cells after secondary allogeneic stimulation were almost entirely blocked by 10 nm pimecrolimus. Taken together, pimecrolimus inhibits T cell proliferation and Th1 cytokine synthesis and also prevents the up-regulation of the OX40 co-receptor on primed T cells indicating its potential in the therapy of chronic inflammation and autoimmunity. PMID:12296857

  8. Experimental colitis in rats induces de novo synthesis of cytokines at distant intestinal sites: role of capsaicin-sensitive primary afferent fibers.

    PubMed

    Mourad, Fadi H; Hamdi, Tamim; Barada, Kassem A; Saadé, Nayef E

    2016-06-01

    Increased levels of pro- and anti-inflammatory cytokines were observed in various segments of histologically-intact small intestine in animal models of acute and chronic colitis. Whether these cytokines are produced locally or spread from the inflamed colon is not known. In addition, the role of gut innervation in this upregulation is not fully understood. To examine whether cytokines are produced de novo in the small intestine in two rat models of colitis; and to investigate the role of capsaicin-sensitive primary afferents in the synthesis of these inflammatory cytokines. Colitis was induced by rectal instillation of iodoacetamide (IA) or trinitrobenzene sulphonic acid (TNBS) in adult Sprague-Dawley rats. Using reverse transcriptase (RT) and real-time PCR, TNF-α, and IL-10 mRNA expression was measured in mucosal scrapings of the duodenum, jejunum, ileum and colon at different time intervals after induction of colitis. Capsaicin-sensitive primary afferents (CSPA) were ablated using subcutaneous injections of capsaicin at time 0, 8 and 32 h, and the experiment was repeated at specific time intervals to detect any effect on cytokines expression. TNF-α mRNA expression increased by 3-40 times in the different intestinal segments (p<0.05 to p<0.001), 48h after IA-induced colitis. CSPA ablation completely inhibited this upregulation in the small intestine, but not in the colon. Similar results were obtained in TNBS-induced colitis at 24 h. Intestinal IL-10 mRNA expression significantly decreased at 12 h and then increased by 6-43 times (p<0.05 to p<0.001) 48h after IA administration. This increase was abolished in rats subjected to CSPA ablation except in the colon, where IL-10 further increased by twice (p<0.05). In the TNBS group, there was 4-12- and 4-7-fold increases in small intestinal IL-10 mRNA expression at 1 and 21 days after colitis induction, respectively (both p<0.01). This increase was not observed in rats pretreated with capsaicin. Capsaicin-treated and

  9. A New Application for Albumin Dialysis in Extracorporeal Organ Support: Characterization of a Putative Interaction Between Human Albumin and Proinflammatory Cytokines IL-6 and TNFα.

    PubMed

    Pfensig, Claudia; Dominik, Adrian; Borufka, Luise; Hinz, Michael; Stange, Jan; Eggert, Martin

    2016-04-01

    Albumin dialysis in extracorporeal organ support is often performed in the treatment of liver failure as it facilitates the removal of toxic components from the blood. Here, we describe a possible effect of albumin dialysis on proinflammatory cytokine levels in vitro. Initially, albumin samples were incubated with different amounts of cytokines and analyzed by enzyme-linked immunosorbent assay (ELISA). Analysis of interleukin 6 (IL-6) and tumor necrosis factor alpha (TNFα) levels indicated that increased concentrations of albumin reduce the measureable amount of the respective cytokines. This led to the hypothesis that the used proinflammatory cytokines may interact with albumin. Size exclusion chromatography of albumin spiked with cytokines was carried out using high-performance liquid chromatography analysis. The corresponding fractions were evaluated by immunoblotting. We detected albumin and cytokines in the same fractions indicating an interaction of the small-sized cytokines IL-6 and TNFα with the larger-sized albumin. Finally, a two-compartment albumin dialysis in vitro model was used to analyze the effect of albumin on proinflammatory cytokines in the recirculation circuit during 6-h treatment. These in vitro albumin dialysis experiments indicated a significant decrease of IL-6, but not of TNFα, when albumin was added to the dialysate solution. Taken together, we were able to show a putative in vitro interaction of human albumin with the proinflammatory cytokine IL-6, but with less evidence for TNFα, and demonstrated an additional application for albumin dialysis in liver support therapy where IL-6 removal might be indicated.

  10. Pro-inflammatory cytokine dysregulation is associated with novel avian influenza A (H7N9) virus in primary human macrophages.

    PubMed

    Zhao, Chihao; Qi, Xian; Ding, Meng; Sun, Xinlei; Zhou, Zhen; Zhang, Shuo; Zen, Ke; Li, Xihan

    2016-02-01

    Since March 2013, more than 500 laboratory-confirmed human H7N9 influenza A virus infection cases have been recorded, with a case fatality rate of more than 30%. Clinical research has shown that cytokine and chemokine dysregulation contributes to the pathogenicity of the H7N9 virus. Here, we investigated cytokine profiles in primary human macrophages infected with the novel H7N9 virus, using cytokine antibody arrays. The levels of several pro-inflammatory cytokines, particularly TNF-α, were increased in H7N9-infected macrophages. Induction of the transcriptional and translational levels of the pro-inflammatory cytokines by H7N9 virus seemed to be intermediate between those induced by highly pathogenic avian H5N1 and pandemic human H1N1 viruses, which were detected by ELISA and real-time quantitative PCR, respectively. Additionally, compared with H5N1, the upregulation of pro-inflammatory cytokines caused by H7N9 infection occurred rapidly but mildly. Our results identified the overall profiles of cytokine and chemokine induction by the H7N9 influenza virus in an in vitro cell-culture model, and could provide potential therapeutic targets for the control of severe human H7N9 disease.

  11. Immunostimulatory DNA inhibits IL-4-dependent IgE synthesis by human B cells.

    PubMed

    Horner, A A; Widhopf, G F; Burger, J A; Takabayashi, K; Cinman, N; Ronaghy, A; Spiegelberg, H L; Raz, E

    2001-09-01

    Immunostimulatory sequence oligodeoxynucleotide (ISS-ODN) is a potent antiallergic immunomodulating agent in mice. However, few studies have addressed its antiallergic potential in human subjects. We sought to determine whether a phosphoro-thioate ISS-ODN could inhibit IL-4-dependent IgE synthesis by human B cells. Initially, nonatopic- and atopic-donor PBMCs were incubated with ISS-ODN or mutated oligodeoxynucleotide, and cytokine production and B-cell expression of IFN-gamma receptor and IL-4 receptor were measured by using ELISA and flow cytometry, respectively. In subsequent studies atopic-donor PBMCs were incubated with IL-4 alone or with ISS-ODN or mutated oligodeoxynucleotide. After 14 days, IgE production and IgM, IgG, and IgA production were determined by using ELISA. In select IgE studies cytokines were neutralized with mAbs. ISS-ODN induced IL-12, IFN-alpha, IFN-gamma, IL-10, and IL-6 production from both nonatopic- and atopic-donor PBMCs. ISS-ODN also increased IFN-gamma receptor and inhibited IL-4 receptor expression on B cells from both donor populations. Furthermore, ISS-ODN inhibited IL-4-dependent IgE production by atopic-donor PBMCs. Neutralization of IL-12, IFN-alpha, IFN-gamma, and IL-10, but not IL-6, attenuated the inhibitory activity of ISS-ODN on IgE production. In contrast to its inhibition of IgE synthesis, ISS-ODN stimulated the production of IgM, IgG, and IgA. These in vitro studies demonstrate that phos-phorothioate ISS-ODN elicits an innate immune response by PBMCs, which inhibits IL-4-dependent IgE synthesis. In addition, these results provide further support for consideration of ISS-ODN therapy for the treatment of allergic disease in clinical practice.

  12. Pulmonary infection of mice with human metapneumovirus induces local cytotoxic T-cell and immunoregulatory cytokine responses similar to those seen with human respiratory syncytial virus.

    PubMed

    Herd, Karen A; Nelson, Michelle; Mahalingam, Suresh; Tindle, Robert W

    2010-05-01

    Human metapneumovirus (hMPV) is a major cause of upper and lower respiratory-tract infection in infants, the elderly and immunocompromised individuals. Virus-directed cellular immunity elicited by hMPV infection is poorly understood, in contrast to the phylogenetically and clinically related pathogen human respiratory syncytial virus (hRSV). In a murine model of acute lower respiratory-tract infection with hMPV, we demonstrate the accumulation of gamma interferon (IFN-gamma)-producing CD8+ T cells in the airways and lungs at day 7 post-infection (p.i.), associated with cytotoxic T lymphocytes (CTLs) directed to an epitope of the M2-1 protein. This CTL immunity was accompanied by increased pulmonary expression of Th1 cytokines IFN-gamma and interleukin (IL)-12 and antiviral cytokines (IFN-beta), as well as chemokines Mip-1alpha, Mip-1beta, Mig, IP-10 and CX3CL1. There was also a moderate increase in Th2-type cytokines IL-4 and IL-10 compared with uninfected mice. At 21 days p.i., a strong CTL response could be recalled from the spleen. A similar pattern of CTL induction to the homologous M2-1 CTL epitope of hRSV, and of cytokine/chemokine induction, was observed following infection with hRSV, highlighting similarities in the cellular immune response to the two related pathogens.

  13. Transfusion of cryopreserved human red blood cells into healthy humans is associated with rapid extravascular hemolysis without a proinflammatory cytokine response.

    PubMed

    Hult, Andreas; Malm, Christer; Oldenborg, Per-Arne

    2013-01-01

    Transfusion of stored red blood cells (RBCs) can be associated with adverse side effects. Recent studies in mice transfused with stored RBCs showed that a strong proinflammatory cytokine storm was induced due to extravascular hemolysis already at 2 hours after transfusion. Therefore, we here investigated if transfusion of 2 units of cryopreserved autologous RBCs induced a proinflammatory response in healthy human volunteers. Two units of autologous RBCs, cryopreserved for 16 weeks, were transfused into 10 healthy human volunteers. Serum and blood samples taken at 2 hours before and at 2 and 48 hours after transfusion were analyzed for signs of extravascular hemolysis and the presence of proinflammatory cytokines. At 2 hours after transfusion, transferin-bound serum iron, as well as transferin saturation and total bilirubin, were already significantly increased. These measures all returned back toward that in pretransfusion samples at 48 hours after transfusion. No increases in the production of the proinflammatory cytokines interleukin (IL)-1β, IL-6, IL-8, monocyte chemotactic protein-1, macrophage inflammatory protein-1β, or tumor necrosis factor-α were detected at any time point after transfusion. Although a significant level of extravascular hemolysis already occurred at 2 hours after transfusion of cryopreserved RBCs, there were no signs of proinflammatory cytokine production up to 48 hours after transfusion. © 2012 American Association of Blood Banks.

  14. Involvement of reactive oxygen species in brominated diphenyl ether-47-induced inflammatory cytokine release from human extravillous trophoblasts in vitro

    SciTech Connect

    Park, Hae-Ryung Kamau, Patricia W.; Loch-Caruso, Rita

    2014-01-15

    Polybrominated diphenyl ethers (PBDEs) are widely used flame retardant compounds. Brominated diphenyl ether (BDE)-47 is one of the most prevalent PBDE congeners found in human breast milk, serum and placenta. Despite the presence of PBDEs in human placenta, effects of PBDEs on placental cell function are poorly understood. The present study investigated BDE-47-induced reactive oxygen species (ROS) formation and its role in BDE-47-stimulated proinflammatory cytokine release in a first trimester human extravillous trophoblast cell line, HTR-8/SVneo. Exposure of HTR-8/SVneo cells for 4 h to 20 μM BDE-47 increased ROS generation 1.7 fold as measured by the dichlorofluorescein (DCF) assay. Likewise, superoxide anion production increased approximately 5 fold at 10 and 15 μM and 9 fold at 20 μM BDE-47 with a 1-h exposure, as measured by cytochrome c reduction. BDE-47 (10, 15 and 20 μM) decreased the mitochondrial membrane potential by 47–64.5% at 4, 8 and 24 h as assessed with the fluorescent probe Rh123. Treatment with 15 and 20 μM BDE-47 stimulated cellular release and mRNA expression of IL-6 and IL-8 after 12 and 24-h exposures: the greatest increases were a 35-fold increased mRNA expression at 12 h and a 12-fold increased protein concentration at 24 h for IL-6. Antioxidant treatments (deferoxamine mesylate, (±)α-tocopherol, or tempol) suppressed BDE-47-stimulated IL-6 release by 54.1%, 56.3% and 37.7%, respectively, implicating a role for ROS in the regulation of inflammatory pathways in HTR-8/SVneo cells. Solvent (DMSO) controls exhibited statistically significantly decreased responses compared with non-treated controls for IL-6 release and IL-8 mRNA expression, but these responses were not consistent across experiments and times. Nonetheless, it is possible that DMSO (used to dissolve BDE-47) may have attenuated the stimulatory actions of BDE-47 on cytokine responses. Because abnormal activation of proinflammatory responses can disrupt trophoblast functions

  15. Enhanced production of the chemotactic cytokines interleukin-8 and monocyte chemoattractant protein-1 in human abdominal aortic aneurysms.

    PubMed Central

    Koch, A. E.; Kunkel, S. L.; Pearce, W. H.; Shah, M. R.; Parikh, D.; Evanoff, H. L.; Haines, G. K.; Burdick, M. D.; Strieter, R. M.

    1993-01-01

    Inflammatory leukocytes play a central role in the pathogenesis of human atherosclerotic disease, from early atherogenesis to the late stages of atherosclerosis, such as aneurysm formation. We have shown previously that human abdominal aortic aneurysms are characterized by the presence of numerous chronic inflammatory cells throughout the vessel wall (Am J Pathol 1990, 137: 1199-1213). The signals that attract lymphocytes and monocytes into the aortic wall in aneurysmal disease remain to be precisely defined. We have studied the production of the chemotactic cytokines interleukin-8 (IL-8) and monocyte chemoattractant protein-1 (MCP-1) by aortic tissues obtained from 47 subjects. We compared the antigenic production of these cytokines by explants of: 1) human abdominal aneurysmal tissue, 2) occlusive (atherosclerotic) aortas, and 3) normal aortas. IL-8, which is chemotactic for neutrophils, lymphocytes, and endothelial cells was liberated in greater quantities by abdominal aortic aneurysms than by occlusive or normal aortas. Using immunohistochemistry, macrophages, and to a lesser degree endothelial cells, were found to be positive for the expression of antigenic IL-8. Similarly, MCP-1, a potent chemotactic cytokine for monocytes/macrophages, was released by explants from abdominal aortic aneurysms in greater quantities than by explants from occlusive or normal aortas. Using immunohistochemistry, the predominant MCP-1 antigen-positive cells were macrophages and to a lesser extent smooth muscle cells. Our results indicate that human abdominal aortic aneurysms produce IL-8 and MCP-1, both of which may serve to recruit additional inflammatory cells into the abdominal aortic wall, hence perpetuating the inflammatory reaction that may result in the pathology of vessel wall destruction and aortic aneurysm formation. Images Figure 2 Figure 3 Figure 4 Figure 5 PMID:8494046

  16. A Human In Vitro Whole Blood Assay to Predict the Systemic Cytokine Response to Therapeutic Oligonucleotides Including siRNA

    PubMed Central

    Schwickart, Anna; Putschli, Bastian; Renn, Marcel; Höller, Tobias; Barchet, Winfried

    2013-01-01

    Therapeutic oligonucleotides including siRNA and immunostimulatory ligands of Toll-like receptors (TLR) or RIG-I like helicases (RLH) are a promising novel class of drugs. They are in clinical development for a broad spectrum of applications, e.g. as adjuvants in vaccines and for the immunotherapy of cancer. Species-specific immune activation leading to cytokine release is characteristic for therapeutic oligonucleotides either as an unwanted side effect or intended pharmacology. Reliable in vitro tests designed for therapeutic oligonucleotides are therefore urgently needed in order to predict clinical efficacy and to prevent unexpected harmful effects in clinical development. To serve this purpose, we here established a human whole blood assay (WBA) that is fast and easy to perform. Its response to synthetic TLR ligands (R848: TLR7/8, LPS: TLR4) was on a comparable threshold to the more time consuming peripheral blood mononuclear cell (PBMC) based assay. By contrast, the type I IFN profile provoked by intravenous CpG-DNA (TLR9 ligand) in humans in vivo was more precisely replicated in the WBA than in stimulated PBMC. Since Heparin and EDTA, but not Hirudin, displaced oligonucleotides from their delivery agent, only Hirudin qualified as the anticoagulant to be used in the WBA. The Hirudin WBA exhibited a similar capacity as the PBMC assay to distinguish between TLR7-activating and modified non-stimulatory siRNA sequences. RNA-based immunoactivating TLR7/8- and RIG-I-ligands induced substantial amounts of IFN-α in the Hirudin-WBA dependent on delivery agent used. In conclusion, we present a human Hirudin WBA to determine therapeutic oligonucleotide-induced cytokine release during preclinical development that can readily be performed and offers a close reflection of human cytokine response in vivo. PMID:23940691

  17. Unique proliferation response in odontoblastic cells derived from human skeletal muscle stem cells by cytokine-induced matrix metalloproteinase-3

    SciTech Connect

    Ozeki, Nobuaki; Hase, Naoko; Kawai, Rie; Yamaguchi, Hideyuki; Hiyama, Taiki; Kondo, Ayami; Nakata, Kazuhiko; Mogi, Makio

    2015-02-01

    A pro-inflammatory cytokine mixture (CM: interleukin (IL)-1β, tumor necrosis factor-α and interferon-γ) and IL-1β-induced matrix metalloproteinase (MMP)-3 activity have been shown to increase the proliferation of rat dental pulp cells and murine stem cell-derived odontoblast-like cells. This suggests that MMP-3 may regulate wound healing and regeneration in the odontoblast-rich dental pulp. Here, we determined whether these results can be extrapolated to human dental pulp by investigating the effects of CM-induced MMP-3 up-regulation on the proliferation and apoptosis of purified odontoblast-like cells derived from human skeletal muscle stem cells. We used siRNA to specifically reduce MMP-3 expression. We found that CM treatment increased MMP-3 mRNA and protein levels as well as MMP-3 activity. Cell proliferation was also markedly increased, with no changes in apoptosis, upon treatment with CM and following the application of exogenous MMP-3. Endogenous tissue inhibitors of metalloproteinases were constitutively expressed during all experiments and unaffected by MMP-3. Although treatment with MMP-3 siRNA suppressed cell proliferation, it also unexpectedly increased apoptosis. This siRNA-mediated increase in apoptosis could be reversed by exogenous MMP-3. These results demonstrate that cytokine-induced MMP-3 activity regulates cell proliferation and suppresses apoptosis in human odontoblast-like cells. - Highlights: • Pro-inflammatory cytokines induce MMP-3 activity in human odontoblast-like cells. • Increased MMP-3 activity can promote cell proliferation in odontoblasts. • Specific loss of MMP-3 increases apoptosis in odontoblasts. • MMP-3 has potential as a promising new target for pupal repair and regeneration.

  18. Regulation of IL-13 synthesis in human lymphocytes: implications for asthma therapy

    PubMed Central

    Pahl, Andreas; Zhang, Meixia; Kuss, Hildegard; Szelenyi, Istvan; Brune, Kay

    2002-01-01

    IL-13 is an important mediator in inflammatory diseases such as asthma. IL-13 is mainly produced by T cells. However, signalling pathways leading to induction of this cytokine are not well-characterized. We analysed the regulation of IL-13 in human peripheral blood mononuclear cells and CD4+ T cells.Cyclosporine (CsA) and FK-506 inhibited IL-13 synthesis, when cells were stimulated by TPA/ionomycin. However, stimulation by α-CD3/α-CD28 led to an enhanced IL-13 synthesis.NF-κB inhibitor N-tosyl-L-lysine chloromethylketone (TLCK) inhibited IL-13 synthesis more effectively after TPA/ionomycin stimulation. After α-CD3/α-CD28 stimulation, only 300 μM TLCK inhibited IL-13 synthesis. Dexamethasone inhibited IL-13 equally effective after α-CD3/α-CD28 and TPA/ionomycin stimulation.p38 MAPK inhibitor SB203580 inhibited IL-13 synthesis only partially. MEK inhibitor U0126 inhibited TPA/ionomycin induced IL-13 synthesis very effectively, whereas α-CD3/α-CD28 stimulated IL-13 induction was resistant to this drug.These results were confirmed in purified CD4+ T cells. In difference to PBMCs α-CD3/α-CD28 stimulated IL-13 synthesis was effectively inhibited by CsA, FK-506 and U0126.Therefore U0126 was tested in an animal model of allergic asthma. We could demonstrate for the first time that inhibition of the MEK – ERK cascade is a therapeutic option for asthma. Intraperitoneal administration of 10 mg kg−1 U0126 reduced lung eosinophilia in ovalbumin-challenged Brown Norway rats by 44%.These results demonstrate that different signalling pathways are involved in regulating IL-13 synthesis in primary human T cells. Characterizing highly potent inhibitors of IL-13 synthesis can be exploited to identify new drugs to treat immunological diseases such as asthma. PMID:11959794

  19. Human periodontal ligament cells facilitate leukocyte recruitment and are influenced in their immunomodulatory function by Th17 cytokine release.

    PubMed

    Konermann, A; Beyer, M; Deschner, J; Allam, J P; Novak, N; Winter, J; Jepsen, S; Jäger, A

    2012-01-01

    The objective of this in vitro study was to examine the immunomodulatory impact of human periodontal ligament (PDL) cells on the nature and magnitude of the leukocyte infiltrate in periodontal inflammation, particularly with regard to Th17 cells. PDL cells were challenged with pro-inflammatory cytokines (IL-1ß, IL-17A, and IFN-γ) and analyzed for the expression of cytokines involved in periodontal immunoinflammatory processes (IL-6, MIP-3 alpha, IL-23A, TGFß1, IDO, and CD274). In order to further investigate a direct involvement of PDL cells in leukocyte function, co-culture experiments were conducted. The expression of the immunomodulatory cytokines studied was significantly increased under pro-inflammatory conditions in PDL cells. Although PDL cells did not stimulate leukocyte proliferation or Th17 differentiation, these cells induced the recruitment of leukocytes. The results of our study suggest that PDL cells might be involved in chronic inflammatory mechanisms in periodontal tissues and thus in the transition to an adaptive immune response in periodontitis.

  20. The influence of recombinant human erythropoietin on apoptosis and cytokine production of CD4+ lymphocytes from hemodialyzed patients.

    PubMed

    Lisowska, Katarzyna A; Dębska-Ślizień, Alicja; Jasiulewicz, Aleksandra; Daca, Agnieszka; Bryl, Ewa; Witkowski, Jacek M

    2013-04-01

    Recombinant human erythropoietin (rhEPO) treatment of hemodialyzed (HD) patients normalizes the altered phenotype of CD4(+) lymphocytes and restores the balance of Th1/Th2 cytokines. We decided to test how the presence of rhEPO in cell culture modulates cytokine production of CD4(+) lymphocytes in HD patients with stable hemoglobin level and expression of activation antigens of stimulated CD4(+) lymphocytes similar to those observed in healthy individuals. We also tested whether the presence of rhEPO in cell culture protects stimulated CD4(+) lymphocytes of HD patients from apoptosis. Peripheral blood mononuclear cells (PBMC) of HD patients were stimulated with an immobilized anti-CD3 antibody with or without addition of rhEPO. The percentage of apoptotic CD4(+) lymphocytes and the level of Th1/Th2 cytokines in culture supernatants were measured with flow cytometry. HD patients showed a decrease in the percentage of apoptotic CD4(+) cells after stimulation with the anti-CD3 antibody combined with rhEPO. The level of IFN-γ and IL-10 was increased while the level of TNF-α was decreased in the presence of rhEPO in cell culture from HD patients. These results confirm the role of rhEPO signaling in T lymphocytes of HD patients.

  1. Propionibacterium acnes and lipopolysaccharide induce the expression of antimicrobial peptides and proinflammatory cytokines/chemokines in human sebocytes.

    PubMed

    Nagy, István; Pivarcsi, Andor; Kis, Kornélia; Koreck, Andrea; Bodai, László; McDowell, Andrew; Seltmann, Holger; Patrick, Sheila; Zouboulis, Christos C; Kemény, Lajos

    2006-07-01

    Acne is a common skin disorder of the pilosebaceous unit. In addition to genetic, hormonal and environmental factors, abnormal colonization by Propionibacterium acnes has been implicated in the occurrence of acne via the induction of inflammatory mediators. To gain more insight into the role that sebocytes play in the innate immune response of the skin, particularly in acne, we compared the antimicrobial peptide and proinflammatory cytokine/chemokine expression at mRNA and protein levels, as well as the viability and differentiation of SZ95 sebocytes in response to co-culture with representative isolates of P. acnes type IA and type IB as well as Escherichia coli-derived lipopolysaccharide (LPS). We found that, in vitro, P. acnes type IA and IB isolates and LPS induced human beta-defensin-2 and proinflammatory cytokine/chemokine expression, and influenced sebocyte viability and differentiation. Our results provide evidence that sebocytes are capable of producing proinflammatory cytokines/chemokines and antimicrobial peptides, which may have a role in acne pathogenesis. Furthermore, since P. acnes types IA and IB differentially affect both the differentiation and viability of sebocytes, our data demonstrate that different strains of P. acnes vary in their capacity to stimulate an inflammatory response within the pilosebaceous follicle.

  2. Antioxidants inhibit SAA formation and pro-inflammatory cytokine release in a human cell model of alkaptonuria.

    PubMed

    Spreafico, Adriano; Millucci, Lia; Ghezzi, Lorenzo; Geminiani, Michela; Braconi, Daniela; Amato, Loredana; Chellini, Federico; Frediani, Bruno; Moretti, Elena; Collodel, Giulia; Bernardini, Giulia; Santucci, Annalisa

    2013-09-01

    Alkaptonuria (AKU) is an ultra-rare autosomal recessive disease that currently lacks an appropriate therapy. Recently we provided experimental evidence that AKU is a secondary serum amyloid A (SAA)-based amyloidosis. The aim of the present work was to evaluate the use of antioxidants to inhibit SAA amyloid and pro-inflammatory cytokine release in AKU. We adopted a human chondrocytic cell AKU model to evaluate the anti-amyloid capacity of a set of antioxidants that had previously been shown to counteract ochronosis in a serum AKU model. Amyloid presence was evaluated by Congo red staining. Homogentisic acid-induced SAA production and pro-inflammatory cytokine release (overexpressed in AKU patients) were evaluated by ELISA and multiplex systems, respectively. Lipid peroxidation was evaluated by means of a fluorescence-based assay. Our AKU model allowed us to prove the efficacy of ascorbic acid combined with N-acetylcysteine, taurine, phytic acid and lipoic acid in significantly inhibiting SAA production, pro-inflammatory cytokine release and membrane lipid peroxidation. All the tested antioxidant compounds were able to reduce the production of amyloid and may be the basis for establishing new therapies for AKU amyloidosis.

  3. UVB-induced inflammatory cytokine release, DNA damage and apoptosis of human oral compared with skin tissue equivalents.

    PubMed

    Breger, Joyce; Baeva, Larissa; Agrawal, Anant; Shindell, Eli; Godar, Dianne E

    2013-01-01

    People can get oral cancers from UV (290-400 nm) exposures. Besides high outdoor UV exposures, high indoor UV exposures to oral tissues can occur when consumers use UV-emitting tanning devices to either tan or whiten their teeth. We compared the carcinogenic risks of skin to oral tissue cells after UVB (290-320 nm) exposures using commercially available 3D-engineered models for human skin (EpiDerm™), gingival (EpiGing™) and oral (EpiOral™) tissues. To compare the relative carcinogenic risks, we investigated the release of cytokines, initial DNA damage in the form of cyclobutane pyrimidine dimers (CPDs), repair of CPDs and apoptotic cell numbers. We measured cytokine release using cytometric beads with flow cytometry and previously developed a fluorescent immunohistochemical assay to quantify simultaneously CPD repair rates and apoptotic cell numbers. We found that interleukin-8 (IL-8) release and the initial CPDs are significantly higher, whereas the CPD repair rates and apoptotic cell numbers are significantly lower for oral compared with skin tissue cells. Thus, the increased release of the inflammatory cytokine IL-8 along with inefficient CPD repair and decreased death rates for oral compared with skin tissue cells suggests that mutations are accumulating in the surviving population of oral cells increasing people's risks for getting oral cancers.

  4. Obesity and inflammation: reduced cytokine expression due to resveratrol in a human in vitro model of inflamed adipose tissue

    PubMed Central

    Zagotta, Ivana; Dimova, Elitsa Y.; Debatin, Klaus-Michael; Wabitsch, Martin; Kietzmann, Thomas; Fischer-Posovszky, Pamela

    2015-01-01

    Obesity is associated with an inflammatory status and linked with a number of pathophysiological complications among them cardiovascular disease, type 2 diabetes mellitus, or the metabolic syndrome. Resveratrol was proposed to improve obesity-related inflammatory problems, but the effect of resveratrol on cytokine expression in obesity is not completely understood. In this study, we used an in vitro model of human adipose tissue inflammation to examine the effects of resveratrol on the production of the inflammatory cytokines interleukin 6 (IL-6), IL-8, and monocyte chemoattractant protein 1 (MCP-1). We found that resveratrol reduced IL-6, IL-8, and MCP-1 levels in a concentration-dependent manner in adipocytes under inflammatory conditions. Further experiments showed that the action of resveratrol was mainly due to its NFκB inhibitory potential. Thus, our data support the concept that resveratrol can alleviate obesity-induced up-regulation of inflammatory cytokines providing a new insight toward novel treatment options in obesity. PMID:25926797

  5. 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.

  6. A cytokine-independent approach to identify antigen-specific human germinal center Tfh cells and rare antigen-specific CD4+ T cells in blood

    PubMed Central

    Dan, Jennifer M.; Arlehamn, Cecilia S. Lindestam; Weiskopf, Daniela; Antunes, Ricardo da Silva; Havenar-Daughton, Colin; Reiss, Samantha; Brigger, Matthew; Bothwell, Marcella; Sette, Alessandro; Crotty, Shane

    2016-01-01

    Detection of antigen-specific CD4+ T cells is central to the study of many human infectious diseases, vaccines, and autoimmune diseases. However, such cells are generally rare and heterogeneous in their cytokine profiles. Identification of antigen-specific germinal center (GC) T follicular helper (Tfh) cells by cytokine production has been particularly problematic. The function of a GC Tfh cell is to selectively help adjacent GC B cells via cognate interaction; thus, GC Tfh cells may be ‘stingy’ cytokine producers, fundamentally different than Th1 or Th17 cells in the quantities of cytokines produced. Conventional identification of antigen-specific cells by intracellular cytokine staining (ICS) relies on the ability of the CD4+ T cell to generate substantial amounts of cytokine. To address this problem, we have developed a cytokine-independent activation induced marker (AIM) methodology to identify antigen-specific GC Tfh cells in human lymphoid tissue. Whereas Group A Streptococcus (Strep)-specific GC Tfh cells produced minimal detectable cytokines by ICS, the AIM method identified 85-fold more antigen-specific GC Tfh cells. Intriguingly, these GC Tfh cells consistently expressed programmed death ligand 1 (PD-L1) upon activation. AIM also detected non-Tfh cells in lymphoid tissue. As such, we applied AIM for identification of rare antigen-specific CD4+ T cells in human peripheral blood. Dengue-, tuberculosis-, and pertussis-vaccine-specific CD4+ T cells were readily detectable by AIM. In sum, cytokine assays missed 98% of antigen-specific human GC Tfh cells, reflecting the biology of these cells, which could instead be sensitively identified by co-expression of TCR-dependent activation markers. PMID:27342848

  7. Possible Roles of Proinflammatory and Chemoattractive Cytokines Produced by Human Fetal Membrane Cells in the Pathology of Adverse Pregnancy Outcomes Associated with Influenza Virus Infection

    PubMed Central

    Uchide, Noboru; Ohyama, Kunio; Bessho, Toshio; Takeichi, Makoto; Toyoda, Hiroo

    2012-01-01

    Pregnant women are at an increased risk of influenza-associated adverse outcomes, such as premature delivery, based on data from the latest pandemic with a novel influenza A (H1N1) virus in 2009-2010. It has been suggested that the transplacental transmission of influenza viruses is rarely detected in humans. A series of our study has demonstrated that influenza virus infection induced apoptosis in primary cultured human fetal membrane chorion cells, from which a factor with monocyte differentiation-inducing (MDI) activity was secreted. Proinflammatory cytokines, such as interleukin (IL)-6, tumor necrosis factor (TNF)-α, and interferon (IFN)-β, were identified as a member of the MDI factor. Influenza virus infection induced the mRNA expression of not only the proinflammatory cytokines but also chemoattractive cytokines, such as monocyte chemoattractant protein (MCP)-1, regulated on activation, normal T-cell expressed and secreted (RANTES), macrophage inflammatory protein (MIP)-1β, IL-8, growth-regulated oncogene (GRO)-α, GRO-β, epithelial cell-derived neutrophil-activating protein (ENA)-78, and interferon inducible protein (IP)-10 in cultured chorion cells. These cytokines are postulated to associate with human parturition. This paper, therefore, reviews (1) lessons from pandemic H1N1 2009 in pregnancy, (2) production of proinflammatory and chemoattractive cytokines by human fetal membranes and their functions in gestational tissues, and (3) possible roles of cytokines produced by human fetal membranes in the pathology of adverse pregnancy outcomes associated with influenza virus infection. PMID:22899878

  8. 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.

  9. Human decidual macrophages and NK cells differentially express Toll-like receptors and display distinct cytokine profiles upon TLR stimulation

    PubMed Central

    Duriez, Marion; Quillay, Héloïse; Madec, Yoann; El Costa, Hicham; Cannou, Claude; Marlin, Romain; de Truchis, Claire; Rahmati, Mona; Barré-Sinoussi, Françoise; Nugeyre, Marie-Thérèse; Menu, Elisabeth

    2014-01-01

    Maternofetal pathogen transmission is partially controlled at the level of the maternal uterine mucosa at the fetal implantation site (the decidua basalis), where maternal and fetal cells are in close contact. Toll-like receptors (TLRs) may play an important role in initiating rapid immune responses against pathogens in the decidua basalis, however the tolerant microenvironment should be preserved in order to allow fetal development. Here we investigated the expression and functionality of TLRs expressed by decidual macrophages (dMs) and NK cells (dNKs), the major decidual immune cell populations. We report for the first time that both human dMs and dNK cells express mRNAs encoding TLRs 1-9, albeit with a higher expression level in dMs. TLR2, TLR3, and TLR4 protein expression checked by flow cytometry was positive for both dMs and dNK cells. In vitro treatment of primary dMs and dNK cells with specific TLR2, TLR3, TLR4, TLR7/8, and TLR9 agonists enhanced their secretion of pro- and anti-inflammatory cytokines, as well as cytokines and chemokines involved in immune cell crosstalk. Only dNK cells released IFN-γ, whereas only dMs released IL-1β, IL-10, and IL-12. TLR9 activation of dMs resulted in a distinct pattern of cytokine expression compared to the other TLRs. The cytokine profiles expressed by dMs and dNK cells upon TLR activation are compatible with maintenance of the fetotolerant immune environment during initiation of immune responses to pathogens at the maternofetal interface. PMID:25071732

  10. Cytokine Production Associated with Periportal Fibrosis during Chronic Schistosomiasis Mansoni in Humans

    PubMed Central

    Alves Oliveira, L. F.; Moreno, E. C.; Gazzinelli, G.; Martins-Filho, O. A.; Silveira, A. M. S.; Gazzinelli, A.; Malaquias, L. C. C.; LoVerde, P.; Leite, P. Martins; Correa-Oliveira, R.

    2006-01-01

    Volunteers living in an area where schistosomiasis mansoni is endemic were subjected to ultrasound examination and classified into groups according to the levels of fibrosis diagnosed, namely, absence of indications of fibrosis (group 0), incipient fibrosis (group 1), and moderate/severe fibrosis (group 2). Peripheral blood mononuclear cells (PBMC) collected from the volunteers were stimulated with soluble antigens from adult schistosomes or from schistosome eggs, and the production of the cytokines gamma interferon, tumor necrosis factor alpha, transforming growth factor β (TGF-β), interleukin-4 (IL-4), IL-10, and IL-13 was determined. Potential associations of the level of fibrosis with age, sex, intensity of infection, and cytokine production were investigated between the three groups. Univariate analysis identified associations of age (>50), gender (male), and absence of eggs/g of feces with moderate/severe fibrosis and an association of intensity of infection (>100 eggs) with incipient fibrosis. When cytokine production in PBMC cultures stimulated by soluble egg antigens was categorized as low or high, significant differences in the distribution of IL-13 levels were established between groups 0 and 2. No significant differences were detected between the groups in the cytokines produced by PBMC cultures stimulated with soluble antigens from adult schistosomes. When all variables were tested in multivariate analyses, only IL-13 was strongly associated with fibrosis (odds ratio = 5.8; 95% confidence interval [CI] = 1.1 to 30.5). While high levels of TGF-β appeared to be associated with protection against fibrosis, the strength of the association was low. PMID:16428771

  11. Gene Expression Profile of Human Cytokines in Response to B.pseudomallei Infection

    DTIC Science & Technology

    2017-04-19

    during 68 exposure to soil and contaminated water. Nevertheless inhalation of aerosolized 69 bacteria during extreme weather events such as rainfall and...definitive tests. Bacteria were resuscitated by subculture onto 134 5% blood agar and incubated for 24 h at 37 °C to give single colony growth for all 135...IL17 and other Th17 cytokines are linked to the response against extra-cellular 214 bacteria , pathogenesis of diverse autoimmune and inflammatory

  12. CHARACTERIZATION OF CYTOKINE-INDUCED MYELOID-DERIVED SUPPRESSOR CELLS FROM NORMAL HUMAN PERIPHERAL BLOOD MONONUCLEAR CELLS

    PubMed Central

    Lechner, Melissa G.; Liebertz, Daniel J.; Epstein, Alan L.

    2010-01-01

    Introduction Tumor immune tolerance can derive from the recruitment of suppressor cell populations, including myeloid-derived suppressor cells (MDSC). In cancer patients, increased MDSC correlate with more aggressive disease and a poor prognosis. Experimental Design Expression of 15 immune factors (TGFβ, IL-1β, IL-4, IL-6, IL-10, GM-CSF, M-CSF, IDO, FLT3L, c-kit L, iNOS, ARG-1, TNFα, COX2, VEGF) by MDSC-inducing human solid tumor cell lines was evaluated by RT-PCR. Based upon these data, cytokine mixtures were then tested for their ability to generate suppressive CD33+ cells from healthy donor PBMC in vitro by measuring their ability to inhibit the proliferation of, and IFNγ production by, fresh autologous human T cells after CD3/CD28 stimulation. Induced MDSC were characterized with respect to their morphology, surface phenotype, and gene expression profile. Results MDSC-inducing cancer cell lines demonstrated multiple pathways for MDSC generation, including over-expression of IL-6, IL-1β, COX2, M-CSF, and IDO. CD33+ cells with potent suppressive capacity were best generated in vitro by GM-CSF and IL-6, and secondarily by GM-CSF + IL-1β, PGE2, TNFα, or VEGF. Characterization studies of cytokine-induced suppressive cells revealed CD33+CD11b+CD66b+HLA-DRlowIL-13Rα2int large mononuclear cells with abundant basophilic cytoplasm. Expression of iNOS, TGFβ, NOX2, VEGF, and/or ARG-1 was also up-regulated and transwell studies showed suppression of autologous T cells to be contact dependent. Conclusion Suppressive CD33+ cells generated from PBMC by GM-CSF and IL-6 were consistent with human MDSC. This study suggests that these cytokines are potential therapeutic targets for the inhibition of MDSC induction in cancer patients. PMID:20644162

  13. GADS is Required for TCR-Mediated Calcium Influx and Cytokine Release, but not Cellular Adhesion, in Human T Cells

    PubMed Central

    Bilal, Mahmood Y.; Zhang, Elizabeth Y.; Dinkel, Brittney; Hardy, Daimon; Yankee, Thomas M.; Houtman, Jon C.D.

    2015-01-01

    GRB2 related adaptor protein downstream of Shc (GADS) is a member of the GRB2 family of adaptors and is critical for TCR-induced signaling. The current model is that GADS recruits SLP-76 to the LAT complex, which facilitates the phosphorylation of SLP-76, the activation of PLC-γ1, T cell adhesion and cytokine production. However, this model is largely based on studies of disruption of the GADS/SLP-76 interaction and murine T cell differentiation in GADS deficient mice. The role of GADS in mediating TCR-induced signals in human CD4+ T cells has not been thoroughly investigated. In this study, we have suppressed the expression of GADS in human CD4+ HuT78 T cells. GADS deficient HuT78 T cells displayed similar levels of TCR-induced SLP-76 and PLC-γ1 phosphorylation but exhibited substantial decrease in TCR-induced IL-2 and IFN-γ release. The defect in cytokine production occurred because of impaired calcium mobilization due to reduced recruitment of SLP-76 and PLC-γ1 to the LAT complex. Surprisingly, both GADS deficient HuT78 and GADS deficient primary murine CD8+ T cells had similar TCR-induced adhesion when compared to control T cells. Overall, our results show that GADS is required for calcium influx and cytokine production, but not cellular adhesion, in human CD4+ T cells, suggesting that the current model for T cell regulation by GADS is incomplete. PMID:25636200

  14. Cyclic hydrostatic pressure and particles increase synthesis of 1,25-dihydroxyvitamin D3 by human macrophages in vitro.

    PubMed

    Evans, C E; Mylchreest, S; Mee, A P; Berry, J L; Andrew, J G

    2006-01-01

    1,25-Dihydroxyvitamin D(3) has a pivotal role in bone resorption and osteoclast activity. As activated macrophages are known to synthesise 1,25-dihydroxyvitamin D(3), this study examined whether pressure modulated its synthesis. Pressure and particles have been shown to increase synthesis of pro-resorptive cytokines and other factors by cultured macrophages. Human peripheral blood macrophages were isolated, cultured and exposed to pressure (similar to that found in the human joint) and/or particles. Synthesis of 1,25-dihydroxyvitamin D(3) by macrophages was assayed using high pressure liquid chromatography and in situ hybridization. Synthesis of 1,25-dihydroxyvitamin D(3) but not 24,25-dihydroxyvitamin D(3) was increased in macrophages under pressure. In situ hybridization demonstrated an increase in 1alpha-hydroxylase expression in response to pressure or particles and simultaneous exposure to both stimuli generated higher expression of 1alpha-hydroxylase. In conclusion, this is the first study to demonstrate that mechanical loading, in the form of pressure, stimulates 1,25-dihydroxyvitamin D(3) synthesis in human macrophages. These findings have implications for the in vivo situation, as they suggest that 1,25-dihydroxyvitamin D(3) could be one factor stimulating osteoclastic bone resorption in pathologies, such as arthritis or implant loosening, where intra-articular or intra-osseous pressure is raised or where wear particles interact with macrophages.

  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. Inhibiting inflammatory cytokines.

    PubMed

    Kluth, D C; Rees, A J

    1996-11-01

    Acute glomerulonephritis is a common cause of renal dysfunction and ultimately renal failure. The inflammation involved is a tightly regulated response with pro- and anti-inflammatory cytokines playing key roles. Interleukin-1 (IL-1) and tumor necrosis factor (TNF) are the principal pro-inflammatory cytokines produced by intrinsic cells and infiltrating leukocytes. IL-1 and TNF can be directly antagonized using IL-1 receptor antagonist (IL-1ra) or binding proteins such as soluble receptors or antibodies. Alternatively, cytokines with anti-inflammatory properties can be used to decrease IL-1 and TNF synthesis, increase the production of their natural antagonists and deactivate inflammatory cells such as macrophages. This review will focus on these anti-inflammatory cytokines, principally IL-4, IL-6, IL-10 and IL-13, and highlight recent research of their activities in existing models of renal disease. The results of these experiments offer a promising new avenue of treatment.

  17. DNA polymerases and repair synthesis in NER in human cells.

    PubMed

    Lehmann, Alan R

    2011-07-15

    The late steps of nucleotide excision repair, following incisions to remove the damaged section of DNA, comprise repair synthesis and ligation. In vitro and in vivo studies have shown the size of the repaired patch to be about 30 nucleotides. In vitro studies implicated the replicative polymerases in repair synthesis, but recent in vivo data have shown that several DNA polymerases and ligases are involved in these steps in human cells. Copyright © 2011 Elsevier B.V. All rights reserved.

  18. Curcumin Induces Pro-apoptotic Effects Against Human Melanoma Cells and Modulates The Cellular Response to Immunotherapeutic Cytokines

    PubMed Central

    Bill, Matthew A.; Bakan, Courtney; Benson, Don M.; Fuchs, James; Young, Gregory; Lesinski, Gregory B.

    2009-01-01

    Curcumin has potential as a chemopreventative and chemotherapeutic agent however its interactions with clinically relevant cytokines are poorly characterized. Since cytokine immunotherapy is a mainstay of treatment for malignant melanoma, we hypothesized that curcumin could modulate the cellular responsiveness to interferons and interleukins. As a single agent, curcumin induced a dose-dependent increase in apoptosis of human melanoma cell lines, which was most prominent at doses >10 µM. Immunoblot analysis confirmed that curcumin induced apoptosis and revealed caspase-3 processing, PARP cleavage, reduced Bcl-2 and decreased basal phosphorylated STAT3. Despite its pro-apoptotic effects, curcumin pre-treatment of human melanoma cell lines inhibited the phosphorylation of STAT1 protein and downstream gene transcription following IFN-α and IFN-γ as determined by immunoblot analysis and Real Time PCR, respectively. Pre-treatment of peripheral blood mononuclear cells (PBMCs) from healthy donors with curcumin also inhibited the ability of IFN-α, IFN-γ and IL-2 to phosphorylate STAT proteins critical for their anti-tumor activity (STAT1 and STAT5, respectively) and their respective downstream gene expression as measured by Real Time PCR. Finally, stimulation of natural killer (NK) cells with curcumin reduced the level of IL-12-induced IFN-γ secretion, and production of granzyme b or IFN-γ upon co-culture with A375 melanoma cells or NK sensitive K562 cells as targets. These data demonstrate that although curcumin can induce apoptosis of melanoma cells, it can also adversely affect the responsiveness of immune effector cells to clinically relevant cytokines that possess anti-tumor properties. PMID:19723881

  19. Soluble CD14 acts as a DAMP in human macrophages: origin and involvement in inflammatory cytokine/chemokine production.

    PubMed

    Lévêque, Manuella; Simonin-Le Jeune, Karin; Jouneau, Stéphane; Moulis, Solenn; Desrues, Benoit; Belleguic, Chantal; Brinchault, Graziella; Le Trionnaire, Sophie; Gangneux, Jean-Pierre; Dimanche-Boitrel, Marie-Thérèse; Martin-Chouly, Corinne

    2017-05-01

    The innate immune system is able to detect bacterial LPS through the pattern recognition receptor CD14, which delivers LPS to various TLR signaling complexes that subsequently induce intracellular proinflammatory signaling cascades. In a previous study, we showed the overproduction of the soluble form of CD14 (sCD14) by macrophages from patients with cystic fibrosis (CF). CF is an autosomal recessive disorder that is caused by mutations in the gene that encodes the CFTR protein and is characterized by persistent inflammation. Macrophages play a significant role in the initial stages of this disease due to their inability to act as suppressor cells, leading to chronic inflammation in CF. In this work, we investigated the origin of sCD14 by human macrophages and studied the effect of sCD14 on the production of inflammatory cytokine/chemokine. Our data indicate that sCD14 stimulate proinflammatory cytokine/chemokine production in a manner that is independent of LPS but dependent on the TLR-4/CD14 membrane complex, NF-κB, and the inflammasome. Therefore, sCD14, overproduced by CF macrophages, originates primarily from the endocytosis/exocytosis process and should be considered to be a danger-associated molecular pattern. This elucidation of the origin and inflammation-induced mechanisms associated with sCD14 contributes to our understanding of maintained tissue inflammation.-Lévêque, M., Simonin-Le Jeune, K., Jouneau, S., Moulis, S., Desrues, B., Belleguic, C., Brinchault, G., Le Trionnaire, S., Gangneux, J.-P., Dimanche-Boitrel, M.-T., Martin-Chouly, C. Soluble CD14 acts as a DAMP in human macrophages: origin and involvement in inflammatory cytokine/chemokine production. © FASEB.

  20. Inflammatory Cytokines Induce Expression of Chemokines by Human Retinal Cells: Role in Chemokine Receptor Mediated Age-related Macular Degeneration.

    PubMed

    Nagineni, Chandrasekharam N; Kommineni, Vijay K; Ganjbaksh, Nader; Nagineni, Krishnasai K; Hooks, John J; Detrick, Barbara

    2015-11-01

    Chemokine reeptor-3 (CCR-3) was shown to be associated with choroidal neovascularization (CNV) in age-related macular degeneration (AMD). AMD is a vision threatening retinal disease that affects the aging population world-wide. Retinal pigment epithelium and choroid in the posterior part of the retina are the key tissues targeted in the pathogenesis of CNV in AMD. We used human retinal pigment epithelial (HRPE) and choroidal fibroblast (HCHF) cells, prepared from aged adult human donor eyes, to evaluate the expression of major CCR-3 ligands, CCL-5, CCL -7, CCL-11,CCL-24 and CCL-26. Microarray analysis of gene expression in HRPE cells treated with inflammatory cytokine mix (ICM= IFN-γ+TNF-α+IL-1β) revealed 75 and 23-fold increase in CCL-5 and CCL-7 respectively, but not CCL-11, CCL-24 and CCL-26. Chemokine secretion studies of the production of CCL5 and CCL7 by HRPE corroborated with the gene expression analysis data. When the HRPE cells were treated with either individual cytokines or the ICM, both CCL-5 and CCL-7 were produced in a dose dependent manner. Similar to the gene expression data, the ICM did not enhance HRPE production of CCL-11, CCL-24 and CCL-26. CCL-11 and CCL-26 were increased with IL-4 treatment and this HRPE production was augmented in the presence of TNF-α and IL1β. When HCHF cells were treated with either individual cytokines or the ICM, both CCL-5 and CCL-7 were produced in a dose dependent fashion. IL-4 induced low levels of CCL-11 and CCL-26 in HCHF and this production was significantly enhanced by TNF-α. Under these conditions, neither HRPE nor HCHF were demonstrated to produce CCL-24. These data demonstrate that chronic inflammation triggers CCL-5 and CCL-7 release by HRPE and HCHF and the subsequent interactions with CCR3 may participate in pathologic processes in AMD.

  1. Exposure to wear particles generated from studded tires and pavement induces inflammatory cytokine release from human macrophages.

    PubMed

    Lindbom, John; Gustafsson, Mats; Blomqvist, Göran; Dahl, Andreas; Gudmundsson, Anders; Swietlicki, Erik; Ljungman, Anders G

    2006-04-01

    Health risks associated with exposure to airborne particulate matter (PM) have been shown epidemiologically as well as experimentally, pointing to both respiratory and cardiovascular effects. Lately, wear particles generated from traffic have been recognized to be a major contributing source to the overall particle load, especially in the Nordic countries were studded tires are used. In this work, we investigated the inflammatory effect of PM10 generated from the wear of studded tires on two different types of pavement. As comparison, we also investigated PM10 from a traffic-intensive street, a subway station, and diesel exhaust particles (DEP). Human monocyte-derived macrophages, nasal epithelial cells (RPMI 2650), and bronchial epithelial cells (BEAS-2B) were exposed to the different types of particles, and the secretion of IL-6, IL-8, IL-10, and TNF-alpha into the culture medium was measured. The results show a significant release of cytokines from macrophages after exposure for all types of particles. When particles generated from asphalt/granite pavement were compared to asphalt/quartzite pavement, the granite pavement had a significantly higher capacity to induce the release of cytokines. The granite pavement particles induced cytokine release at the same magnitude as the street particles did, which was higher than what particles from both a subway station and DEP did. Exposure of epithelial cells to PM10 resulted in a significant increase of TNF-alpha secreted from BEAS-2B cells for all types of particles used (DEP was not tested), and the highest levels were induced by subway particles. None of the particle types were able to evoke detectable cytokine release from RPMI 2650 cells. The results indicate that PM10 generated by the wear of studded tires on the street surface is a large contributor to the cytokine-releasing ability of particles in traffic-intensive areas and that the type of pavement used is important for the level of this contribution

  2. Cytokine and adhesion molecule expression in primary human endothelial cells stimulated with fever-range hyperthermia.

    PubMed

    Shah, A; Unger, E; Bain, M D; Bruce, R; Bodkin, J; Ginnetti, J; Wang, W C; Seon, B; Stewart, C C; Evans, S S

    2002-01-01

    Migration of blood-borne lymphocytes into lymphoid tissues and sites of inflammation is initiated by vascular adhesion molecules and proinflammatory cytokines. Previous in vivo studies have shown that febrile temperatures dynamically stimulate adhesion in differentiated high endothelial venules (HEV), which are portals for lymphocyte extravasation. This report examines the direct effect of fever-range hyperthermia on the expression of adhesion molecules and cytokines by primary cultured endothelial cells. In both macrovascular (HUVEC) and microvascular (HMVEC) endothelial cells, fever-range hyperthermia (40 degrees C for 6-12 h) did not affect expression of adhesion molecules (ICAM-1, E-selectin, VCAM-1, P-selectin, PECAM-1, PNAd, MAdCAM-1), cytokine release (IL-1beta, TNF-alpha, IFN-gamma, IL-6, IL-11, IL-12, IL-13), or chemokine secretion (IL-8, RANTES, MCP-1, MIP-1beta, MIG). This is in contrast to the stimulatory effects of TNF-alpha or 43 degrees C heat shock. However, a novel role for fever-range hyperthermia was identified in augmenting actin polymerization in cultured endothelial cells and enhancing the ability of endothelial-derived factors to transactivate the alpha4beta7 integrin lymphocyte homing receptor. These findings provide insight into the tightly regulated effects of fever-range hyperthermia that exclude induction of adhesion in non-activated endothelium of normal blood vessels. Through these mechanisms, it is proposed that febrile temperatures associated with infection or clinical hyperthermia avoid the unproductive exodus of lymphocytes to non-involved extralymphoid tissues while simultaneously promoting lymphocyte delivery to sites of immune activation.

  3. Sulphatide and its precursor galactosylceramide influence the production of cytokines in human mononuclear cells.

    PubMed

    Buschard, K; Diamant, M; Bovin, L E; Månsson, J E; Fredman, P; Bendtzen, K

    1996-12-01

    Sulphatide is expressed in the central and peripheral neural system, in islets of Langerhans, and in tissues affected by late diabetic complications. Autoantibodies to sulphatide are present in patients with insulin-dependent diabetes and the Guillain-Barré syndrome. Cytokines influence these disease processes, and we therefore studied whether sulphatide and its precursor galactosylceramide (gal-cer) influence the in vitro production of cytokines by blood mononuclear cells (MNC) originating from 15 healthy persons. Using lipopolysaccharide (LPS)-stimulated cells, sulphatide increased the IL-2 production (163 +/- 17% of controls without sulphatide, p = 0.02), and gal-cer increased the IL-1 alpha production (145 +/- 13%, p = 0.006), whereas neither gal-cer nor sulphatide had an effect on the production of IL-6, IL-10 or TNF alpha. When stimulating cells with phytohaemagglutinin (PHA), sulphatide decreased the production of IL-6 (88 +/- 5%, p = 0.009), IL-10 (66 +/- 3%, p = 0.000003), and TNF alpha (75 +/- 9% p = 0.02). Gal-cer, however, increased the production of IL-6 (188 +/- 13% p = 0.000006), and decreased the production of TNF beta (80 +/- 6%, p = 0.007). Neither gal-cer nor sulphatide had an effect on the production of IL-2 or IFN gamma from PHA-stimulated cells. Northern blot analysis using an IL-6 probe similarly showed an increased amount of IL-6 mRNA after gal-cer incubation (range 469%-150%, n = 3) of PHA-stimulated control. Thus, sulphatide and gal-cer influence the production of several cytokines thought to be involved in immunoinflammatory disease processes.

  4. Butyrate Regulates the Expression of Inflammatory and Chemotactic Cytokines In Human Acute Leukemic Cells During Apoptosis

    PubMed Central

    Pulliam, Stephanie R.; Pellom, Samuel T.; Shanker, Anil; Adunyah, Samuel E.

    2016-01-01

    Butyrate is a histone deacetylase inhibitor implicated in many studies as a potential therapy for various forms of cancer. High concentrations of butyrate (>1.5 mM) have been shown to activate apoptosis in several cancer cell lines including prostate, breast, and leukemia. Butyrate is also known to influence multiple signaling pathways that are mediators of cytokine production. The purpose of this study was to evaluate the impact of high concentrations of butyrate on the cancer microenvironment vis-à-vis apoptosis, cellular migration, and capacity to modulate cytokine expression in cancer cells. The results indicate that high concentrations of butyrate induced a 2-fold activation of caspase-3 and reduced cell viability by 60% in U937 leukemia cells. Within 24 hours, butyrate significantly decreased the levels of chemokines CCL2 and CCL5 in HL-60 and U937 cells, and decreased CCL5 in THP-1 leukemia cells. Differential effects were observed in treatments with valproic acid for CCL2 and CCL5 indicating butyrate-specificity. Many of the biological effects examined in this study are linked to activation of the AKT and MAPK signaling pathways; therefore, we investigated whether butyrate alters the levels of phosphorylated forms of these signaling proteins and how it correlated with the expression of chemokines. The results show that butyrate may partially regulate CCL5 production via p38 MAPK. The decrease in p-ERK1/2 and p-AKT levels correlated with the decrease in CCL2 production. These data suggest that while promoting apoptosis, butyrate has the potential to influence the cancer microenvironment by inducing differential expression of cytokines. PMID:27253488

  5. 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

  6. 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

  7. Blister fluid and serum cytokine levels in severe sepsis in humans reflect skin dysfunction.

    PubMed

    Koskela, M; Ala-Kokko, T I; Gäddnäs, F; Herzig, K-H; Karhu, T; Oikarinen, A; Koivukangas, V

    2017-01-01

    Knowledge of sepsis-related end-organ inflammation in vivo is limited. We investigated the cytokine response in skin and in serum in sepsis and its relation to multiorgan failure (MOF) and survival. Cytokines were analysed in serum and in suction blister fluid of intact skin of 44 patients with severe sepsis and 15 healthy controls. Blister fluid and serum samples were collected within 48 h of the first sepsis-induced organ failure. This is a substudy of a larger follow-up study on wound healing in sepsis. Cytokine levels were higher in patients with sepsis vs. controls (interleukin [IL]-10, blisters: 65.9 vs. 4.3 pg/ml, P < 0.001, serum: 25.7 vs. 4.5 pg/ml, P = 0.004; IL-6, blisters: 41.9 vs. 0.03 pg/ml, P < 0.001, serum: 45.5 vs. 2.1 pg/ml, P < 0.001). Patients with MOF had higher levels of IL-10 (116.4 vs. 21.3 pg/ml, P = 0.015), IL-4 (0.7 vs. 0.07 pg/ml, P = 0.013) and basic fibroblast growth factor (bFGF) (25.9 vs. 9.5 pg/ml, P = 0.027) in blister fluid than patients without MOF. In blister fluid, survivors had lower levels of IL-10 (43.3 vs. 181.9 pg/ml, P = 0.024) and bFGF (15.8 vs. 31.9 pg/ml, P = 0.006) than non-survivors. In serum, survivors had higher levels of vascular endothelial growth factor (VEGF) (152.2 vs. 14.7 pg/ml, P = 0.012) and lower levels of IL-6 (38.5 vs. 91.1 pg/ml, P = 0.011) than non-survivors. The blister fluid levels of bFGF, TNF and VEGF did not correlate with the serum levels. Cytokine responses in skin blister fluid in patients with sepsis differed from those in healthy controls. © 2016 The Acta Anaesthesiologica Scandinavica Foundation. Published by John Wiley & Sons Ltd.

  8. Proinflammatory cytokine secretion is suppressed by TMEM16A or CFTR channel activity in human cystic fibrosis bronchial epithelia

    PubMed Central

    Veit, Guido; Bossard, Florian; Goepp, Julie; Verkman, A. S.; Galietta, Luis J. V.; Hanrahan, John W.; Lukacs, Gergely L.

    2012-01-01

    Cystic fibrosis (CF) is caused by the functional expression defect of the CF transmembrane conductance regulator (CFTR) chloride channel at the apical plasma membrane. Impaired bacterial clearance and hyperactive innate immune response are hallmarks of the CF lung disease, yet the existence of and mechanism accounting for the innate immune defect that occurs before infection remain controversial. Inducible expression of either CFTR or the calcium-activated chloride channel TMEM16A attenuated the proinflammatory cytokines interleukin-6 (IL-6), IL-8, and CXCL1/2 in two human respiratory epithelial models under air–liquid but not liquid–liquid interface culture. Expression of wild-type but not the inactive G551D-CFTR indicates that secretion of the chemoattractant IL-8 is inversely proportional to CFTR channel activity in cftr∆F508/∆F508 immortalized and primary human bronchial epithelia. Similarly, direct but not P2Y receptor–mediated activation of TMEM16A attenuates IL-8 secretion in respiratory epithelia. Thus augmented proinflammatory cytokine secretion caused by defective anion transport at the apical membrane may contribute to the excessive and persistent lung inflammation in CF and perhaps in other respiratory diseases associated with documented down-regulation of CFTR (e.g., chronic obstructive pulmonary disease). Direct pharmacological activation of TMEM16A offers a potential therapeutic strategy to reduce the inflammation of CF airway epithelia. PMID:22973054

  9. Progesterone promotes maternal–fetal tolerance by reducing human maternal T‐cell polyfunctionality and inducing a specific cytokine profile

    PubMed Central

    Eldershaw, Suzy A.; Inman, Charlotte F.; Coomarasamy, Aravinthan; Moss, Paul A. H.; Kilby, Mark D.

    2015-01-01

    Progesterone is a steroid hormone essential for the maintenance of human pregnancy, and its actions are thought to include promoting maternal immune tolerance of the semiallogenic fetus. We report that exposure of maternal T cells to progesterone at physiological doses induced a unique skewing of the cytokine production profile of CD4+ and CD8+ T cells, with reductions not only in potentially deleterious IFN‐γ and TNF‐α production but also in IL‐10 and IL‐5. Conversely, production of IL‐4 was increased. Maternal T cells also became less polyfunctional, focussing cytokine production toward profiles including IL‐4. This was accompanied by reduced T‐cell proliferation. Using fetal and viral antigen‐specific CD8+ T‐cell clones, we confirmed that this as a direct, nonantigen‐specific effect. Yet human T cells lacked conventional nuclear progesterone receptors, implicating a membrane progesterone receptor. CD4+ and CD8+ T cells responded to progesterone in a dose‐dependent manner, with subtle effects at concentrations comparable to those in maternal blood, but profound effects at concentrations similar to those at the maternal–fetal interface. This characterization of how progesterone modulates T‐cell function is important in understanding the normal biology of pregnancy and informing the rational use of progesterone therapy in pregnancies at risk of fetal loss. PMID:26249148

  10. Evidence supporting a protective role for th9 and th22 cytokines in human and experimental periapical lesions.

    PubMed

    Aranha, Andreza Maria Fabio; Repeke, Carlos Eduardo; Garlet, Thiago Pompermaier; Vieira, Andreia Espindola; Campanelli, Ana Paula; Trombone, Ana Paula Favaro; Letra, Ariadne; Silva, Renato Menezes; Garlet, Gustavo Pompermaier

    2013-01-01

    The development of periapical granulomas is dependent on the host response and involves Th1, Th2, Th17, and Treg-related cytokines. The discovery of new Th9 and Th22 subsets, with important immunomodulatory roles mediated by interleukin (IL)-9 and IL-22, respectively, emphasizes the need for reevaluation of current cytokine paradigms in context of periapical lesions. We investigated the expression of IL-9 and IL-22 in active and stable human granulomas and throughout experimental lesion development in mice. Periapical granulomas (N = 83) and control specimens (N = 24) were evaluated regarding the expression of IL-9 and IL-22 via real-time polymerase chain reaction. Experimental periapical lesions were induced in mice (pulp exposure and bacterial inoculation) and the lesions evolution correlation with IL-9 and IL-22 expression kinetics was evaluated. IL-9 and IL-22 mRNA expression was higher in periapical lesions than in control samples; higher levels of IL-9 and IL-22 were observed in inactive than in active lesions. In the experimental lesions model, increasing levels of IL-9 and IL-22 mRNA were detected in the lesions, and inverse correlations were found between IL-9 and IL-22 and the increase of lesion area in the different time point intervals. Our results suggest that Th9 and Th22 pathways may contribute to human and experimental periapical lesion stability. Copyright © 2013 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

  11. Progesterone promotes maternal-fetal tolerance by reducing human maternal T-cell polyfunctionality and inducing a specific cytokine profile.

    PubMed

    Lissauer, David; Eldershaw, Suzy A; Inman, Charlotte F; Coomarasamy, Aravinthan; Moss, Paul A H; Kilby, Mark D

    2015-10-01

    Progesterone is a steroid hormone essential for the maintenance of human pregnancy, and its actions are thought to include promoting maternal immune tolerance of the semiallogenic fetus. We report that exposure of maternal T cells to progesterone at physiological doses induced a unique skewing of the cytokine production profile of CD4(+) and CD8(+) T cells, with reductions not only in potentially deleterious IFN-γ and TNF-α production but also in IL-10 and IL-5. Conversely, production of IL-4 was increased. Maternal T cells also became less polyfunctional, focussing cytokine production toward profiles including IL-4. This was accompanied by reduced T-cell proliferation. Using fetal and viral antigen-specific CD8(+) T-cell clones, we confirmed that this as a direct, nonantigen-specific effect. Yet human T cells lacked conventional nuclear progesterone receptors, implicating a membrane progesterone receptor. CD4(+) and CD8(+) T cells responded to progesterone in a dose-dependent manner, with subtle effects at concentrations comparable to those in maternal blood, but profound effects at concentrations similar to those at the maternal-fetal interface. This characterization of how progesterone modulates T-cell function is important in understanding the normal biology of pregnancy and informing the rational use of progesterone therapy in pregnancies at risk of fetal loss.

  12. Proinflammatory cytokine secretion is suppressed by TMEM16A or CFTR channel activity in human cystic fibrosis bronchial epithelia.

    PubMed

    Veit, Guido; Bossard, Florian; Goepp, Julie; Verkman, A S; Galietta, Luis J V; Hanrahan, John W; Lukacs, Gergely L

    2012-11-01

    Cystic fibrosis (CF) is caused by the functional expression defect of the CF transmembrane conductance regulator (CFTR) chloride channel at the apical plasma membrane. Impaired bacterial clearance and hyperactive innate immune response are hallmarks of the CF lung disease, yet the existence of and mechanism accounting for the innate immune defect that occurs before infection remain controversial. Inducible expression of either CFTR or the calcium-activated chloride channel TMEM16A attenuated the proinflammatory cytokines interleukin-6 (IL-6), IL-8, and CXCL1/2 in two human respiratory epithelial models under air-liquid but not liquid-liquid interface culture. Expression of wild-type but not the inactive G551D-CFTR indicates that secretion of the chemoattractant IL-8 is inversely proportional to CFTR channel activity in cftr(∆F508/∆F508) immortalized and primary human bronchial epithelia. Similarly, direct but not P2Y receptor-mediated activation of TMEM16A attenuates IL-8 secretion in respiratory epithelia. Thus augmented proinflammatory cytokine secretion caused by defective anion transport at the apical membrane may contribute to the excessive and persistent lung inflammation in CF and perhaps in other respiratory diseases associated with documented down-regulation of CFTR (e.g., chronic obstructive pulmonary disease). Direct pharmacological activation of TMEM16A offers a potential therapeutic strategy to reduce the inflammation of CF airway epithelia.

  13. Single-trial conditioning in a human taste-endotoxin paradigm induces conditioned odor aversion but not cytokine responses.

    PubMed

    Grigoleit, Jan-Sebastian; Kullmann, Jennifer S; Winkelhaus, Anne; Engler, Harald; Wegner, Alexander; Hammes, Florian; Oberbeck, Reiner; Schedlowski, Manfred

    2012-02-01

    Immunological responses to bacterial endotoxin can be behaviorally conditioned in rodents. However, it is unclear whether an acute systemic inflammatory response can be behaviorally conditioned in humans. Thus, in a double-blind placebo-controlled study, 20 healthy, male subjects received either a single injection of lipopolysaccharide (LPS) or saline together with a novel tasting beverage (conditioned stimulus, CS). Five days later, all subjects received a saline injection and were re-exposed to the CS. Blood was drawn prior to as well as 0.5, 1.5, 3, 4, 6, and 24 h after LPS administration or CS re-exposure. Endotoxin administration led to transient increases in plasma concentrations of interleukin (IL)-6, IL-10, and tumor necrosis factor (TNF)-α and to a significant rise in body temperature. Sole presentation of the CS during evocation did induce neither alterations in body temperature nor changes in plasma cytokine levels. However, subjects in the experimental group rated the smell of the CS significantly more aversive compared to the control group. Employing endotoxin as a US in a single trial taste-immune conditioning paradigm in humans shows a behaviorally conditioned smell aversion but no learned alterations in cytokine levels. Copyright © 2011 Elsevier Inc. All rights reserved.

  14. 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.

  15. [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.

  16. Native human interferon-α is a strong inductor of endogenous cytokines involved in the suppression of procollagen type I.

    PubMed

    Santak, G; Santak, M; Forčić, D

    2013-09-01

    Native human interferon-α (nHuIFN-α) has a stronger reductive effect on procollagen type I mRNA expression than recombinant human interferon-α (rHuIFN-α). It is partially due to the additive activity of interleukin-1β (IL-1β), which is present in small concentrations in nHuIFN-α. Here, we show that the reductive effect is also the result of the endogenous cytokines induced by the activity of nHuIFN-α. In the culture of MRC5 fibroblasts, we have further found that nHuIFN-α induces endogenous interferons in higher amounts than rHuIFN-α, measured with PCR. That is more pronounced when interferon-γ (IFN-γ) is measured. This result puts a new light on IFN-γ activity in the nHuIFN-α treatment because its role was neglected due to the loss of its activity during the nHuIFN-α preparation process. The findings lead to the conclusion that endogenous cytokines play a significant role in the nHuIFN-α -mediated reduction of procollagen type I mRNA and are therefore an important factor in potential therapeutic usage. Copyright © 2013 Elsevier Masson SAS. All rights reserved.

  17. Sepsis and cytokines: current status.

    PubMed

    Blackwell, T S; Christman, J W

    1996-07-01

    Sepsis is a constellation of clinical signs and symptoms resulting from excessive systemic host inflammatory response to infection. This inflammatory response is largely mediated by cytokines, which are released into the systemic circulation. Plasma concentrations of specific cytokines, TNF alpha, IL-1 beta, IL-6 and IL-8 are frequently elevated in human sepsis and cytokine concentrations correlate with severity and outcome of sepsis. In addition to pro-inflammatory cytokines, soluble cytokine receptors, cytokine receptor antagonists and counter-inflammatory cytokines are also produced in large quantities in patients with sepsis; however, the specific role of these molecules in sepsis remains undefined. A complex interaction of cytokines and cytokine-neutralizing molecules probably determines the clinical presentation and course of sepsis. Intervening in this sequence of events to modify the host inflammatory responses may prove to be a beneficial treatment strategy for sepsis, but currently tested anticytokine therapies have been largely unsuccessful.

  18. Suppression of human anti-inflammatory plasma cytokines IL-10 and IL-1RA with elevation of proinflammatory cytokine IFN-gamma during the isolation of the Antarctic winter

    NASA Technical Reports Server (NTRS)

    Shearer, William T.; Lee, Bang-Ning; Cron, Stanley G.; Rosenblatt, Howard M.; Smith, E. O'Brian; Lugg, Desmond J.; Nickolls, Peter M.; Sharp, Robert M.; Rollings, Karl; Reuben, James M.

    2002-01-01

    Cellular immune function has been shown to be decreased and latent virus shedding to be increased in human beings isolated during the Antarctic winter, a model used for assessing some effects of space flight. However, the balance of proinflammatory (IFN-gamma) and anti-inflammatory (IL-10 and IL-1RA) cytokines has not previously been evaluated. We therefore sought to determine whether isolation during the Antarctic winter would alter the proinflammatory and anti-inflammatory cytokine balance. Cytokine levels were measured with ELISA in monthly plasma samples from January through September 1999 in 21 study subjects in the Antarctic and 7 control subjects on Macquarie Island. There was a significant time-dependent increase in plasma IFN-gamma (P =.039) as well as decreases in IL-10 (P =.042) and IL-1RA (P =.053) in the study subjects compared with the control subjects. The study subjects also had significantly increased plasma IFN-gamma levels (P < or =.045) but decreased IL-10 and IL-1RA levels (P < or =.036) at individual time points of isolation. Isolation of human beings in the Antarctic appears to shift the plasma cytokine balance toward a proinflammatory profile. These observations are consistent with T-cell activation that might be due to activation of latent viruses, and they could hold importance for determining the risks of space flight.

  19. Suppression of human anti-inflammatory plasma cytokines IL-10 and IL-1RA with elevation of proinflammatory cytokine IFN-gamma during the isolation of the Antarctic winter

    NASA Technical Reports Server (NTRS)

    Shearer, William T.; Lee, Bang-Ning; Cron, Stanley G.; Rosenblatt, Howard M.; Smith, E. O'Brian; Lugg, Desmond J.; Nickolls, Peter M.; Sharp, Robert M.; Rollings, Karl; Reuben, James M.

    2002-01-01

    Cellular immune function has been shown to be decreased and latent virus shedding to be increased in human beings isolated during the Antarctic winter, a model used for assessing some effects of space flight. However, the balance of proinflammatory (IFN-gamma) and anti-inflammatory (IL-10 and IL-1RA) cytokines has not previously been evaluated. We therefore sought to determine whether isolation during the Antarctic winter would alter the proinflammatory and anti-inflammatory cytokine balance. Cytokine levels were measured with ELISA in monthly plasma samples from January through September 1999 in 21 study subjects in the Antarctic and 7 control subjects on Macquarie Island. There was a significant time-dependent increase in plasma IFN-gamma (P =.039) as well as decreases in IL-10 (P =.042) and IL-1RA (P =.053) in the study subjects compared with the control subjects. The study subjects also had significantly increased plasma IFN-gamma levels (P < or =.045) but decreased IL-10 and IL-1RA levels (P < or =.036) at individual time points of isolation. Isolation of human beings in the Antarctic appears to shift the plasma cytokine balance toward a proinflammatory profile. These observations are consistent with T-cell activation that might be due to activation of latent viruses, and they could hold importance for determining the risks of space flight.

  20. Beryllium Alters Lipopolysaccharide-Mediated Intracellular Phosphorylation and Cytokine Release in Human Peripheral Blood Mononuclear Cells

    PubMed Central

    Silva, Shannon; Ganguly, Kumkum; Fresquez, Theresa M.; Gupta, Goutam; McCleskey, T. Mark; Chaudhary, Anu

    2013-01-01

    Beryllium exposure in susceptible individuals leads to the development of chronic beryllium disease, a lung disorder marked by release of inflammatory cytokine and granuloma formation. We have previously reported that beryllium induces an immune response even in blood mononuclear cells from healthy individuals. In this study, we investigate the effects of beryllium on lipopolysaccharide - mediated cytokine release in blood mononuclear and dendritic cells from healthy individuals. We find that in vitro treatment of beryllium sulfate inhibits the secretion of lipopolysaccharide-mediated interleukin 10, while the release of interleukin 1β is enhanced. Additionally, not all lipopolysaccharide - mediated responses are altered, as interleukin 6 release in unaffected upon beryllium treatment. Beryllium sulfate treated cells show altered phosphotyrosine levels upon lipopolysaccharide stimulation. Significantly, beryllium inhibits the phosphorylation of signal transducer and activator of transducer 3, induced by lipopolysaccharide. Finally, inhibitors of phosphoinositide-3 kinase mimic the effects of beryllium in inhibition of interleukin 10 release, while they have no effect on interleukin 1β secretion. This study strongly suggests that prior exposures to beryllium could alter host immune responses to bacterial infections in healthy individuals, by altering intracellular signaling. PMID:19894180

  1. PRDM1/Blimp-1 Controls Effector Cytokine Production in Human NK Cells

    PubMed Central

    Smith, Matthew A.; Maurin, Michelle; Cho, Hyun Il; Becknell, Brian; Freud, Aharon G.; Yu, Jianhua; Wei, Sheng; Djeu, Julie; Celis, Esteban; Caligiuri, Michael A.; Wright, Kenneth L.

    2013-01-01

    NK cells are major effectors of the innate immune response through cytolysis and bridge to the adaptive immune response through cytokine release. The mediators of activation are well studied however little is known about the mechanisms which restrain activation. In this report, we demonstrate that the transcriptional repressor PRDM1 (also known as Blimp-1 or PRDI-BF1) is a critical negative regulator of NK function. Three distinct PRDM1 isoforms are selectively induced in the CD56dim NK population in response to activation. PRDM1 coordinately suppresses release of IFNγ, TNFα and TNFβ through direct binding to multiple conserved regulatory regions. Ablation of PRDM1 expression leads to enhanced production of IFNγ and TNFα but does not alter cytotoxicity, while overexpression blocks cytokine production. Novel PRDM1 response elements are defined at both the IFNG and TNF loci. Collectively, these data demonstrate a key role for PRDM1 in the negative regulation of NK activation and position PRDM1 as a common regulator of both the adaptive and innate immune response. PMID:20944005

  2. Human cytokine responses to coronary artery bypass grafting with and without cardiopulmonary bypass.

    PubMed

    Strüber, M; Cremer, J T; Gohrbandt, B; Hagl, C; Jankowski, M; Völker, B; Rückoldt, H; Martin, M; Haverich, A

    1999-10-01

    Coronary artery bypass grafting (CABG) is associated with a systemic inflammatory response. This has been attributed to cytokine release caused by extracorporeal circulation and myocardial ischemia. This study compares the inflammatory response after CABG with cardiopulmonary bypass and after minimally invasive direct coronary artery bypass grafting (MIDCABG) without cardiopulmonary bypass. Cytokine release and complement activation (interleukin-6 and interleukin-8, soluble tumor necrosis factor receptors 1 and 2, complement factor C3a, and C1 esterase inhibitor) were determined in 24 patients before and after CABG or MIDCABG. The maximum body temperature, chest drainage, and fluid balance were recorded for 24 hours after operation. Release of interleukin-6, interleukin-8, and tumor necrosis factor receptors 1 and 2 was significantly higher (p < or = 0.005) in the CABG group than the MIDCABG group just after operation. After 24 hours, a significant increase in interleukin-6 was also found in the MIDCABG group (p = 0.001) compared with preoperative value. Body temperature and fluid balance were significantly higher after CABG (p < or = 0.001). Minimally invasive direct coronary artery bypass grafting represents a less traumatizing technique of surgical revascularization. The reduction in the inflammatory response may be advantageous for patients with a high degree of comorbidity.

  3. Interferon gamma and interleukin 4 stimulate prolonged expression of inducible nitric oxide synthase in human airway epithelium through synthesis of soluble mediators.

    PubMed Central

    Guo, F H; Uetani, K; Haque, S J; Williams, B R; Dweik, R A; Thunnissen, F B; Calhoun, W; Erzurum, S C

    1997-01-01

    Human respiratory epithelium expresses inducible nitric oxide synthase (iNOS) continuously in vivo, however mechanisms responsible for maintenance of expression are not known. We show that IFNgamma is sufficient for induction of iNOS in primary human airway epithelial cells (HAEC) in vitro, and IL-4 potentiates IFNgamma-induced iNOS expression in HAEC through stabilization of iNOS mRNA. IFNgamma/IL-4- induced iNOS expression in HAEC was delayed in onset and prolonged with expression up to 1 wk. Removal of overlying culture media resulted in loss of expression, while transfer of conditioned media induced iNOS mRNA in other HAEC. IFNgamma and IL-4 stimulation activated STAT1 and STAT6 in HAEC, but conditioned media transfer to HAEC produced even higher levels of STAT1 activation than achieved by direct addition of cytokines. Although cytokine induction of iNOS was dependent on new protein synthesis, conditioned media induction of iNOS in HAEC was not. Further, removal of overlying culture media from cells at different times after cytokine stimulation demonstrated that mediator synthesis and/or secretion important for induction and maintenance of iNOS occurs early after cytokine stimulation. In conclusion, a combination of IFNgamma/ IL-4, which occurs naturally in the lung epithelial lining fluid, leads to maintenance of iNOS expression in human airway epithelium through production of soluble mediators and stabilization of mRNA. PMID:9259582

  4. Inflammatory gene networks in term human decidual cells define a potential signature for cytokine-mediated parturition.

    PubMed

    Ibrahim, Sherrine A; Ackerman, William E; Summerfield, Taryn L; Lockwood, Charles J; Schatz, Frederick; Kniss, Douglas A

    2016-02-01

    Inflammation is a proximate mediator of preterm birth and fetal injury. During inflammation several microRNAs (22 nucleotide noncoding ribonucleic acid (RNA) molecules) are up-regulated in response to cytokines such as interleukin-1β. MicroRNAs, in most cases, fine-tune gene expression, including both up-regulation and down-regulation of their target genes. However, the role of pro- and antiinflammatory microRNAs in this process is poorly understood. The principal goal of the work was to examine the inflammatory genomic profile of human decidual cells challenged with a proinflammatory cytokine known to be present in the setting of preterm parturition. We determined the coding (messenger RNA) and noncoding (microRNA) sequences to construct a network of interacting genes during inflammation using an in vitro model of decidual stromal cells. The effects of interleukin-1β exposure on mature microRNA expression were tested in human decidual cell cultures using the multiplexed NanoString platform, whereas the global inflammatory transcriptional response was measured using oligonucleotide microarrays. Differential expression of select transcripts was confirmed by quantitative real time-polymerase chain reaction. Bioinformatics tools were used to infer transcription factor activation and regulatory interactions. Interleukin-1β elicited up- and down-regulation of 350 and 78 nonredundant transcripts (false discovery rate < 0.1), respectively, including induction of numerous cytokines, chemokines, and other inflammatory mediators. Whereas this transcriptional response included marked changes in several microRNA gene loci, the pool of fully processed, mature microRNA was comparatively stable following a cytokine challenge. Of a total of 6 mature microRNAs identified as being differentially expressed by NanoString profiling, 2 (miR-146a and miR-155) were validated by quantitative real time-polymerase chain reaction. Using complementary bioinformatics approaches, activation

  5. Macrophage inhibitory cytokine MIC-1 is upregulated by short-wavelength light in cultured normal human dermal fibroblasts.

    PubMed

    Akiyama, Masashi; Okano, Keiko; Fukada, Yoshitaka; Okano, Toshiyuki

    2009-03-04

    To better understand dermal response to visible light, we used DNA microarray analysis to search genes induced by blue or near-UV light in normal human dermal fibroblasts. Of about 12800 transcripts analyzed, near-UV light most prominently upregulated the transcript level of Mic-1, a gene encoding a TGF-beta superfamily protein. Quantitative RT-PCR and immunoblot analyses revealed that mRNA and protein levels of Mic-1 were upregulated by both short-wavelength light but not by green or red light. These results suggest that the human dermis is a site for macrophage inhibitory cytokine-1 (MIC-1) production and that visible light activates a dermal transcription cascade. Considering the role of MIC-1 in immune regulation and appetite control, photic MIC-1 regulation is of physiological importance.

  6. Asbestos and erionite prime and activate the NLRP3 inflammasome that stimulates autocrine cytokine release in human mesothelial cells

    PubMed Central

    2013-01-01

    Background Pleural fibrosis and malignant mesotheliomas (MM) occur after exposures to pathogenic fibers, yet the mechanisms initiating these diseases are unclear. Results We document priming and activation of the NLRP3 inflammasome in human mesothelial cells by asbestos and erionite that is causally related to release of IL-1β, IL-6, IL-8, and Vascular Endothelial Growth Factor (VEGF). Transcription and release of these proteins are inhibited in vitro using Anakinra, an IL-1 receptor antagonist that reduces these cytokines in a human peritoneal MM mouse xenograft model. Conclusions These novel data show that asbestos-induced priming and activation of the NLRP3 inflammasome triggers an autocrine feedback loop modulated via the IL-1 receptor in mesothelial cell type targeted in pleural infection, fibrosis, and carcinogenesis. PMID:23937860

  7. Xylosyltransferase-I regulates glycosaminoglycan synthesis during the pathogenic process of human osteoarthritis.

    PubMed

    Venkatesan, Narayanan; Barré, Lydia; Bourhim, Mustapha; Magdalou, Jacques; Mainard, Didier; Netter, Patrick; Fournel-Gigleux, Sylvie; Ouzzine, Mohamed

    2012-01-01

    Loss of glycosaminoglycan (GAG) chains of proteoglycans (PGs) is an early event of osteoarthritis (OA) resulting in cartilage degradation that has been previously demonstrated in both huma and experimental OA models. However, the mechanism of GAG loss and the role of xylosyltransferase-I (XT-I) that initiates GAG biosynthesis onto PG molecules in the pathogenic process of human OA are unknown. In this study, we have characterized XT-I expression and activity together with GAG synthesis in human OA cartilage obtained from different regions of the same joint, defined as "normal", "late-stage" or adjacent to "late-stage". The results showed that GAG synthesis and content increased in cartilage from areas flanking OA lesions compared to cartilage from macroscopically "normal" unaffected regions, while decreased in "late-stage" OA cartilage lesions. This increase in anabolic state was associated with a marked upregulation of XT-I expression and activity in cartilage "next to lesion" while a decrease in the "late-stage" OA cartilage. Importantly, XT-I inhibition by shRNA or forced-expression with a pCMV-XT-I construct correlated with the modulation of GAG anabolism in human cartilage explants. The observation that XT-I gene expression was down-regulated by IL-1β and up-regulated by TGF-β1 indicates that these cytokines may play a role in regulating GAG content in human OA. Noteworthy, expression of IL-1β receptor (IL-1R1) was down-regulated whereas that of TGF-β1 was up-regulated in early OA cartilage. Theses observations may account for upregulation of XT-I and sustained GAG synthesis prior to the development of cartilage lesions during the pathogenic process of OA.

  8. Xylosyltransferase-I Regulates Glycosaminoglycan Synthesis during the Pathogenic Process of Human Osteoarthritis

    PubMed Central

    Venkatesan, Narayanan; Barré, Lydia; Bourhim, Mustapha; Magdalou, Jacques; Mainard, Didier; Netter, Patrick; Fournel-Gigleux, Sylvie; Ouzzine, Mohamed

    2012-01-01

    Loss of glycosaminoglycan (GAG) chains of proteoglycans (PGs) is an early event of osteoarthritis (OA) resulting in cartilage degradation that has been previously demonstrated in both huma and experimental OA models. However, the mechanism of GAG loss and the role of xylosyltransferase-I (XT-I) that initiates GAG biosynthesis onto PG molecules in the pathogenic process of human OA are unknown. In this study, we have characterized XT-I expression and activity together with GAG synthesis in human OA cartilage obtained from different regions of the same joint, defined as “normal”, “late-stage” or adjacent to “late-stage”. The results showed that GAG synthesis and content increased in cartilage from areas flanking OA lesions compared to cartilage from macroscopically “normal” unaffected regions, while decreased in “late-stage” OA cartilage lesions. This increase in anabolic state was associated with a marked upregulation of XT-I expression and activity in cartilage “next to lesion” while a decrease in the “late-stage” OA cartilage. Importantly, XT-I inhibition by shRNA or forced-expression with a pCMV-XT-I construct correlated with the modulation of GAG anabolism in human cartilage explants. The observation that XT-I gene expression was down-regulated by IL-1β and up-regulated by TGF-β1 indicates that these cytokines may play a role in regulating GAG content in human OA. Noteworthy, expression of IL-1β receptor (IL-1R1) was down-regulated whereas that of TGF-β1 was up-regulated in early OA cartilage. Theses observations may account for upregulation of XT-I and sustained GAG synthesis prior to the development of cartilage lesions during the pathogenic process of OA. PMID:22479506

  9. Comparison of the potency of a variety of β-glucans to induce cytokine production in human whole blood

    PubMed Central

    Noss, Ilka; Doekes, Gert; Thorne, Peter S; Heederik, Dick J.J.; Wouters, Inge M.

    2014-01-01

    Beta-glucans are components of fungal cell walls and potent stimulants of innate immunity. The majority of research on biological activities of glucans has focused on β-(1,3)-glucans, which have been implicated in relation with fungal exposure-associated respiratory symptoms, and as important stimulatory agents in anti-fungal immune responses. Fungi - and bacteria and plants - produce a wide variety of glucans with vast differences in proportion and arrangement of their 1,3-, 1,4-, and 1,6-β-glycosidic linkages. Thus far the proinflammatory potential of different β-glucans has not been studied within the same experimental model. Therefore, we compared the potency of 13 different glucan preparations to induce in vitro production of IL1β, IL6, IL8 and TNF-α in human whole blood cultures. The strongest inducers of all cytokines were pustulan (β-(1,6)-glucan), lichenan (β-(1,3)-(1,4)-glucan), xyloglucan (β-(1,4)-glucan), and pullulan (α-(1,4)-(1,6)-glucan). Moderate to strong cytokine production was observed for curdlan (β-(1,3)-glucan), baker’s yeast glucan (β-(1,3)-(1,6)-glucan), and barley glucan (β-(1,3)-(1,4)-glucan), while all other glucan preparations induced only low or no detectable levels of cytokines. We therefore conclude that innate immunity reactions are not exclusively induced by β-(1,3)-glucans, but also by β-(1,6)- and β-(1,4)-structures. Thus, not only β-(1,3)-glucan, but also other β-glucans and particularly β-(1,6)-glucans should be considered in future research. PMID:22653750

  10. Expression and Functional Role of α7 Nicotinic Receptor in Human Cytokine-stimulated Natural Killer (NK) Cells.

    PubMed

    Zanetti, Samanta R; Ziblat, Andrea; Torres, Nicolás I; Zwirner, Norberto W; Bouzat, Cecilia

    2016-08-05

    The homomeric α7 nicotinic receptor (nAChR) is one of the most abundant nAChRs in the central nervous system where it contributes to cognition, attention, and working memory. α7 nAChR is also present in lymphocytes, dendritic cells (DCs), and macrophages and it is emerging as an important drug target for intervention in inflammation and sepsis. Natural killer (NK) cells display cytotoxic activity against susceptible target cells and modulate innate and adaptive immune responses through their interaction with DCs. We here show that human NK cells also express α7 nAChR. α7 nAChR mRNA is detected by RT-PCR and cell surface expression of α7 nAChR is detected by confocal microscopy and flow cytometry using α-bungarotoxin, a specific antagonist. Both mRNA and protein levels increase during NK stimulation with cytokines (IL-12, IL-18, and IL-15). Exposure of cytokine-stimulated NK cells to PNU-282987, a specific α7 nAChR agonist, increases intracellular calcium concentration ([Ca(2+)]i) mainly released from intracellular stores, indicating that α7 nAChR is functional. Moreover, its activation by PNU-282987 plus a specific positive allosteric modulator greatly enhances the Ca(2+) responses in NK cells. Stimulation of NK cells with cytokines and PNU-282987 decreases NF-κB levels and nuclear mobilization, down-regulates NKG2D receptors, and decreases NKG2D-dependent cell-mediated cytotoxicity and IFN-γ production. Also, such NK cells are less efficient to trigger DC maturation. Thus, our results demonstrate the anti-inflammatory role of α7 nAChR in NK cells and suggest that modulation of its activity in these cells may constitute a novel target for regulation of the immune response.

  11. Aluminium, carbonyls and cytokines in human nipple aspirate fluids: Possible relationship between inflammation, oxidative stress and breast cancer microenvironment.

    PubMed

    Mannello, F; Ligi, D; Canale, M

    2013-11-01

    The human breast is likely exposed to Al (aluminium) from many sources including diet and personal care products. Underarm applications of aluminium salt-based antiperspirant provide a possible long-term source of exposure, especially after underarm applications to shaved and abraded skin. Al research in breast fluids likely reflects the intraductal microenvironment. We found increased levels of aluminium in noninvasively collected nipple aspirate fluids (NAF) from 19 breast cancer patients compared with 16 healthy control subjects (268 vs 131 μg/l, respectively; p < 0.0001). In the same NAF samples we found significantly increased levels of protein oxidative carbonyls in cancer patients compared to healthy women (2.35 vs 0.41 nmol/mg prot, respectively; p < 0.0001). Aluminium content and carbonyl levels showed a significant positive linear correlation (r(2) 0.6628, p < 0.0001). In cancer NAF samples (containing higher amounts of aluminium salts) we also found a significantly increased levels of pro-inflammatory cytokines (IL-1β, IL-6, IL-12 p70, and TNF-α) and chemoattractant CC and CXC chemokines (IL-8, MIP-1α and MCP-1). In 12 invasive cancer NAF samples we found a significant positive linear correlation among aluminium, carbonyls and pro-inflammatory IL-6 cytokine (Y = 64.79x-39.63, r(2) 0.8192, p < 0.0005), as well as pro-inflammatory monocyte chemoattractant MCP-1 cytokine (Y = 2026x-866, r(2) 0.9495, p < 0.0001). In addition to emerging evidence, our results support the possible involvement of aluminium ions in oxidative and inflammatory status perturbations of breast cancer microenvironment, suggesting aluminium accumulation in breast microenvironment as a possible risk factor for oxidative/inflammatory phenotype of breast cells. © 2013.

  12. Mycoplasma fermentans and TNF-β interact to amplify immune-modulating cytokines in human lung fibroblasts

    PubMed Central

    Fabisiak, James P.; Gao, Fei; Thomson, Robyn G.; Strieter, Robert M.; Watkins, Simon C.; Dauber, James H.

    2010-01-01

    Mycoplasma can establish latent infections and are associated with arthritis, leukemia, and chronic lung disease. We developed an experimental model in which lung cells are deliberately infected with Mycoplasma fermentans. Human lung fibroblasts (HLF) were exposed to live M. fermentans and immune-modulating cytokine release was assessed with and without known inducers of cytokine production. M. fermentans increased IL-6, IL-8/CXCL8, MCP-1/CCL2, and Gro-α/CXCL1 production. M. fermentans interacted with TNF-β to release more IL-6, CXCL8, and CXCL1 than predicted by the responses to either stimulus alone. The effects of live infection were recapitulated by exposure to M. fermentans-derived macrophage-activating lipopeptide-2 (MALP-2), a Toll-like receptor-2- and receptor-6-specific ligand. The synergistic effect of combined stimuli was more pronounced with prolonged incubations. Preexposure to TNF-β sensitized the cells to subsequent MALP-2 challenge, but preexposure to MALP-2 did not alter the IL-6 response to TNF-β. Exposure to M. fermentans or MALP-2 did not enhance nuclear localization, DNA binding, or transcriptional activity of NF-κB and did not modulate early NF-κB activation in response to TNF-β. Application of specific inhibitors of various MAPKs suggested that p38 and JNK/stress-activated protein kinase were involved in early IL-6 release after exposure to TNF-β and M. fermentans, respectively. The combined response to M. fermentans and TNF-β, however, was uniquely sensitive to delayed application of SP-600125, suggesting that JNK/stress-activated protein kinase contributes to the amplification of IL-6 release. Thus M. fermentans interacts with stimuli such as TNF-β to amplify lung cell production of immune-modulating cytokines. The mechanisms accounting for this interaction can now be dissected with the use of this in vitro model. PMID:16751226

  13. 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. Copyright © 2013 Elsevier Ltd. All rights reserved.

  14. Dehydroepiandrosterone (DHEA) replacement decreases insulin resistance and lowers inflammatory cytokines in aging humans.

    PubMed

    Weiss, Edward P; Villareal, Dennis T; Fontana, Luigi; Han, Dong-Ho; Holloszy, John O

    2011-05-01

    Plasma dehydroepiandrosterone (DHEA) decreases ~80% between ages 25 and 75 yr. In a preliminary study, we found that 6 mo of DHEA replacement improved insulin action in elderly individuals. The purpose of the present larger, randomized double-blind study was to determine whether a longer period of DHEA replacement improves glucose tolerance. Fifty-seven men and 68 women aged 65 to 75 yr were randomly assigned to 50 mg DHEA or placebo once daily. Year one was a randomized, double blind trial. Year 2 was an open label continuation. DHEA replacement improved glucose tolerance in participants who had abnormal GT initially, reduced plasma triglycerides, and the inflammatory cytokines IL6 and TNFα.

  15. Dehydroepiandrosterone (DHEA) replacement decreases insulin resistance and lowers inflammatory cytokines in aging humans

    PubMed Central

    Weiss, Edward P.; Villareal, Dennis T.; Fontana, Luigi; Han, Dong-Ho; Holloszy, John O.

    2011-01-01

    Plasma dehydroepiandrosterone (DHEA) decreases ~80% between ages 25 and 75 yr. In a preliminary study, we found that 6 mo of DHEA replacement improved insulin action in elderly individuals. The purpose of the present larger, randomized double-blind study was to determine whether a longer period of DHEA replacement improves glucose tolerance. Fifty-seven men and 68 women aged 65 to 75 yr were randomly assigned to 50 mg DHEA or placebo once daily. Year one was a randomized, double blind trial. Year 2 was an open label continuation. DHEA replacement improved glucose tolerance in participants who had abnormal GT initially, reduced plasma triglycerides, and the inflammatory cytokines IL6 and TNFα. This trial was registered at clinicaltrials.gov as NCT00182975. PMID:21566261

  16. Cytokines decrease apolipoprotein accumulation in medium from Hep G2 cells.

    PubMed

    Ettinger, W H; Varma, V K; Sorci-Thomas, M; Parks, J S; Sigmon, R C; Smith, T K; Verdery, R B

    1994-01-01

    Cytokines, important biochemical mediators of inflammation, cause a rapid fall in the plasma concentration of cholesterol in vivo. One mechanism by which cytokines may cause acquired hypocholesterolemia is by decreasing the hepatic synthesis and secretion of apolipoproteins. To test this hypothesis, we incubated Hep G2 cells with human recombinant tumor necrosis factor-alpha, interleukin-1 beta, and interleukin-6. Each of the cytokines resulted in a dose-related reduction in the concentrations of apolipoprotein (apo) A-I, apoB, and lecithin:cholesterol acyltransferase (LCAT) activity in the medium after 24 hours of incubation. The effect of cytokines on apolipoprotein accumulation was not affected by preincubation of Hep G2 cells with fatty acids. Cytokines decreased the concentration of cellular apoA-I mRNA in a dose-related fashion but did not affect cellular concentrations of apoB mRNA. The concentrations of triglyceride and cholesterol were also reduced in the medium of cells incubated with cytokines. Total cell sterol synthesis rates were calculated by [14C]acetate incorporation. Cells incubated with interleukin-6 had a 31% increase in sterol synthesis rate but a 41% decrease in sterol secretion. These data suggest that these cytokines can decrease the hepatic synthesis and/or secretion of apolipoproteins and that this may explain, in part, the acquired hypocholesterolemia seen during acute and chronic inflammation.

  17. Postinjury suppression of human neutrophil cytokine production results from the stabilization of inhibitory kappaB.

    PubMed

    Zallen, G; Moore, E E; Johnson, J L; Tamura, D Y; Shames, B; Biffl, W L; Silliman, C C

    1999-02-01

    Postinjury neutrophil (PMN) dysfunction is a well recognized event that may be responsible for increased infections. PMN cytokine production is an important component of their bactericidal capacity. When PMNs are stimulated, inhibitory factor kappaB (IkappaB) is degraded, allowing nuclear factor kappaB (NFkappaB) to translocate to the nucleus and promotes genes for the transcription of the interleukin-8 (IL-8) and tumor necrosis factor (TNF) genes. We hypothesize that similar to their late postinjury depressed superoxide production, postinjury PMNs manifest suppressed cytokine production, which is mediated by stabilization of IkappaB levels. Twelve severely injured patients with an injury severity score (ISS) of 24 (+/-4.6) were studied as well as 10 elective surgical patients as a control. PMNs were isolated and incubated for 24 h in RPMI. PMNs were stimulated with lipopolysaccharide (LPS; 100 ng) or PAF (200 nm) and fMLP (1 microM) and release of IL-8, TNF, and interleukin-1 receptor antagonist (IL-1ra) were measured. Postinjury PMNs were also stimulated with LPS (100 ng), and IkappaB breakdown was measured at 0, 30, and 60 min using gel electrophoresis. Postinjury PMNs displayed a significant suppression of both IL-8 and TNF on postinjury Days 1-3, while the release of IL-1ra was preserved throughout the entire study period. In contrast, elective surgical patients demonstrated no decrease in IL-8 or TNF. Furthermore, IkappaB levels were preserved in the postinjury PMNs as compared with normal control PMNs. Postinjury PMNs have a suppressed release of both IL-8 and TNF following injury that did not occur in elective surgical patients. Furthermore, the NFkappaB/IkappaB-independent IL-1ra did not show suppression of release. In addition, stabilization of IkappaB following severe injury leads to decreased PMN IL-8 and TNF production. This genetic reprogramming may help explain PMN dysfunction and subsequent infections seen in severely injured patients.

  18. The effects of exercise withdrawal on mood and inflammatory cytokine responses in humans.

    PubMed

    Poole, Lydia; Hamer, Mark; Wawrzyniak, Andrew J; Steptoe, Andrew

    2011-07-01

    Mechanisms underlying the relationship between exercise and mood are not well understood. This study sought to investigate the role of pro- and anti-inflammatory cytokines and autonomic balance in determining the impact of exercise withdrawal on negative mood. Healthy men and women who regularly exercised (N = 26, mean age = 25.5 years, SD = 4.5 years) were randomised to exercise withdrawal or exercise maintenance for 2 weeks. Protocol adherence was monitored using accelerometers. Inflammatory markers from plasma (interleukin-6, IL-6; tumour necrosis factor-alpha; interleukin-10; and interleukin-1 receptor antagonist), heart-rate variability (HRV) and measures of mood (General Health Questionnaire-28 (GHQ) and the Profile of Mood States (POMS)) were assessed at study entry and at 2-week follow-up. Exercise withdrawal resulted in significant increases in negative mood over time on both the GHQ (p = 0.028) and the POMS (p = 0.005). Following the intervention, IL-6 concentration was lower in the exercise withdrawal than exercise maintenance condition (p = 0.05). No intervention effects were observed for other cytokines or HRV. The mood changes were significantly related to changes in IL-6 concentration (β = - 0.50, p = 0.011), indicating that reduction in IL-6 was related to increased negative mood. Our results are consistent with positive effects of exercise on mental health, but further research on inflammatory pathways is warranted.

  19. A human monoclonal IgG1 potently neutralizing the pro-inflammatory cytokine GM-CSF.

    PubMed

    Krinner, Eva-Maria; Raum, Tobias; Petsch, Silke; Bruckmaier, Sandra; Schuster, Ioana; Petersen, Laetitia; Cierpka, Ronny; Abebe, Derege; Mølhøj, Michael; Wolf, Andreas; Sørensen, Poul; Locher, Mathias; Baeuerle, Patrick A; Hepp, Julia

    2007-02-01

    The pro-inflammatory cytokine GM-CSF is aberrantly produced in many autoimmune and chronic inflammatory human diseases. GM-CSF neutralization by antibodies has been shown to have a profound therapeutic effect in animal models of rheumatoid arthritis, inflammatory lung diseases, psoriasis and multiple sclerosis. Moreover, the absence of GM-CSF in null mutant mice ameliorates or prevents certain of these diseases. Here we describe the biophysical and biological properties of a human anti-GM-CSF IgG1 antibody designated MT203, which was derived by phage display guided selection. MT203 bound with picomolar affinity to an epitope on human and macaque GM-CSF involved in high-affinity receptor interaction. As a consequence, the antibody potently prevented both GM-CSF-induced proliferation of TF-1 cells with a sub-nanomolar IC50 value and the production of the chemokine IL-8 by U937 cells. MT203 neutralized equally well recombinant (r) human (h) GM-CSF from Escherichia coli and yeast, and also normally glycosylated GM-CSF secreted by human lung epithelial cells in response to IL-1beta stimulation. Furthermore, MT203 significantly reduced both survival and activation of peripheral human eosinophils as may be required for effective treatment of inflammatory lung diseases. The antibody did not show a detectable loss of neutralizing activity after 5 days in human serum at 37 degrees C. Based on its favorable properties, MT203 has been selected for development as a novel anti-inflammatory human monoclonal antibody with therapeutic potential in a multitude of human autoimmune and inflammatory diseases.

  20. Multiplex analysis inflammatory cytokines in human blood, breath condensate, and urine matrices

    EPA Science Inventory

    Scientific evidence suggests that inflammation is associated with human health effects and health endpoints, yet most studies have focused on human populations that are already considered “unhealthy”.  As such, it is pertinent to measure inflammatory biomarkers in human biologica...

  1. Multiplex analysis inflammatory cytokines in human blood, breath condensate, and urine matrices

    EPA Science Inventory

    Scientific evidence suggests that inflammation is associated with human health effects and health endpoints, yet most studies have focused on human populations that are already considered “unhealthy”.  As such, it is pertinent to measure inflammatory biomarkers in human biologica...

  2. Memory and flexibility of cytokine gene expression as separable properties of human T(H)1 and T(H)2 lymphocytes.

    PubMed

    Messi, Mara; Giacchetto, Isabella; Nagata, Kinya; Lanzavecchia, Antonio; Natoli, Gioacchino; Sallusto, Federica

    2003-01-01

    CD4+ T cell priming under T helper type I (T(H)1) or T(H)2 conditions gives rise to polarized cytokine gene expression. We found that in these conditions human naive T cells acquired stable histone hyperacetylation at either the Ifng or Il4 promoter. Effector memory T cells showed polarized cytokine gene acetylation patterns in vivo, whereas central memory T cells had hypoacetylated cytokine genes but acquired polarized acetylation and expression after appropriate stimulation. However, hypoacetylation of the nonexpressed cytokine gene did not lead to irreversible silencing because most T(H)1 and T(H)2 cells acetylated and expressed the alternative gene when stimulated under opposite T(H) conditions. Such cytokine flexibility was absent in a subset of T(H)2 cells that failed to up-regulate T-bet and to express interferon-gamma when stimulated under T(H)1 conditions. Thus, most human CD4+ T cells retain both memory and flexibility of cytokine gene expression.

  3. Human Cytokine Genetic Variants Associated With HBsAg Reverse Seroconversion in Rituximab-Treated Non-Hodgkin Lymphoma Patients

    PubMed Central

    Hsiao, Liang-Tsai; Wang, Hao-Yuan; Yang, Ching-Fen; Chiou, Tzeon-Jye; Gau, Jyh-Pyng; Yu, Yuan-Bin; Liu, Hsiao-Ling; Chang, Wen-Chun; Chen, Po-Min; Tzeng, Cheng-Hwai; Chan, Yu-Jiun; Yang, Muh-Hwa; Liu, Jin-Hwang; Huang, Yi-Hsiang

    2016-01-01

    Abstract Hepatitis B virus (HBV) reactivation has been noted in HBV surface antigen (HBsAg)-seronegative patients with CD20+ B-cell non-Hodgkin lymphoma (NHL) undergoing rituximab treatment. Clinically, hepatitis flares are usually associated with the reappearance of HBsAg (reverse seroconversion of HBsAg, HBV-RS). It is unclear whether human genetic factors are related to rituximab-associated HBV reactivation. Unvaccinated HBsAg-seronegative adults (n = 104) with CD20+ NHL who had received rituximab-containing therapy without anti-HBV prophylaxis were enrolled. Eighty-nine candidate single nucleotide polymorphisms (SNPs) of 49 human cytokine genes were chosen and were analyzed using the iPLEX technique. Competing risk regression was used to identify the factors associated with HBV-RS. Participants had a median age of 66.1 years and 56.7% were male (n = 59). The anti-HBs and anti-HBc positivity rates were 82.4% and 94.1%, respectively, among patients for whom data were available (approximately 81%). A mean of 7.14 cycles of rituximab therapy were administered, and a total of 14 (13.4%) patients developed HBV-RS. Nine SNPs showed significant differences in frequency between patients with or without HBV-RS: CD40 rs1883832, IL4 rs2243248 and rs2243263, IL13 rs1295686, IL18 rs243908, IL20 rs1518108, and TNFSF13B rs12428930 and rs12583006. Multivariate analysis showed that ≥6 cycles of rituximab therapy, IL18 rs243908, and the IL4 haplotype rs2243248∼rs2243263 were independently associated with HBV-RS. The IL4 haplotype rs2243248∼rs2243263 was significantly associated with HBV-RS regardless of anti-HBs status. Polymorphisms in human cytokine genes impact the risk of rituximab-associated HBV-RS. PMID:26986131

  4. 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.

  5. An antibody present in normal human serum inhibits the binding of cytokines to their receptors in an in vitro system.

    PubMed Central

    Mosedale, D E; Grainger, D J

    1999-01-01

    The presence of active transforming growth factor-beta (TGF-beta) in serum has not been widely accepted. In particular, although at least five studies have concluded that active TGF-beta is present in normal human plasma and serum, assays that use the extracellular domain of the TGF-beta type II receptor as a capture agent have given contradictory results. We show that there is an antagonist present in normal human serum which inhibits the binding of active TGF-beta to the extracellular domain of the TGF-beta type II receptor when it is coated on the well of an ELISA plate. This antagonist activity is due to a pool of immunoglobulins of the G2, D and M classes. Moreover, we show that this same pool of immunoglobulins also recognizes the extracellular domain of the platelet-derived growth factor alpha-receptor, insulin-like growth factor-1 receptor and interleukin-3 receptor, by serial transfer of serum over the different receptors. In addition, the same immunoglobulin pool inhibits the binding of platelet-derived growth factor-AA to its receptor, in an analogous way to the inhibition of binding of TGF-beta to its type II receptor. Circumstantial evidence suggests that the pool of immunoglobulins is recognizing a carbohydrate residue that is attached to the protein when it is synthesized by the mouse myeloma cell line, NSO, in which it is made. If the cytokine receptors are similarly glycosylated in vivo, then the presence of these antibodies in normal human serum may modulate physiological cytokine signalling. PMID:10493920

  6. Anandamide suppresses proliferation and cytokine release from primary human T-lymphocytes mainly via CB2 receptors.

    PubMed

    Cencioni, Maria Teresa; Chiurchiù, Valerio; Catanzaro, Giuseppina; Borsellino, Giovanna; Bernardi, Giorgio; Battistini, Luca; Maccarrone, Mauro

    2010-01-14

    Anandamide (AEA) is an endogenous lipid mediator that exerts several effects in the brain as well as in peripheral tissues. These effects are mediated mainly by two types of cannabinoid receptors, named CB(1)R and CB(2)R, making AEA a prominent member of the "endocannabinoid" family. Also immune cells express CB(1) and CB(2) receptors, and possess the whole machinery responsible for endocannabinoid metabolism. Not surprisingly, evidence has been accumulated showing manifold roles of endocannabinoids in the modulation of the immune system. However, details of such a modulation have not yet been disclosed in primary human T-cells. In this investigation we used flow cytometry and ELISA tests, in order to show that AEA suppresses proliferation and release of cytokines like IL-2, TNF-alpha and INF-gamma from activated human peripheral T-lymphocytes. However, AEA did not exert any cytotoxic effect on T-cells. The immunosuppression induced by AEA was mainly dependent on CB(2)R, since it could be mimicked by the CB(2)R selective agonist JWH-015, and could be blocked by the specific CB(2)R antagonist SR144528. Instead the selective CB(1)R agonist ACEA, or the selective CB(1)R antagonist SR141716, were ineffective. Furthermore, we demonstrated an unprecedented immunosuppressive effect of AEA on IL-17 production, a typical cytokine that is released from the unique CD4+ T-cell subset T-helper 17. Overall, our study investigates for the first time the effects of the endocannabinoid AEA on primary human T-lymphocytes, demonstrating that it is a powerful modulator of immune cell functions. In particular, not only we clarify that CB(2)R mediates the immunosuppressive activity of AEA, but we are the first to describe such an immunosuppressive effect on the newly identified Th-17 cells. These findings might be of crucial importance for the rational design of new endocannabinoid-based immunotherapeutic approaches.

  7. Modulatory effects of propolis samples from Latin America (Brazil, Cuba and Mexico) on cytokine production by human monocytes.

    PubMed

    Conti, Bruno J; Santiago, Karina B; Búfalo, Michelle C; Herrera, Yahima F; Alday, Efrain; Velazquez, Carlos; Hernandez, Javier; Sforcin, José M

    2015-10-01

    Propolis has been used in folk medicine in different regions of the world including Latin America. Propolis is a resinous mixture of substances collected by honey bees from several botanical sources, and its composition contains a rich chemical variety, depending on the geographical area and plant sources. Our aim was to compare the modulatory effect of propolis samples from three different countries of Latin America (Brazil, Cuba and Mexico) on pro- and anti-inflammatory cytokine production (tumor necrosis factor (TNF)-α and interleukin (IL)-10, respectively) by human monocytes. Cells were incubated with propolis for 18 h at 37°C. Cell viability was assessed by 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyltetrazolium bromide method, and cytokine production was determined by ELISA. All samples did not affect monocyte viability. Brazilian propolis stimulated both TNF-α and IL-10 production by monocytes. Cuban propolis stimulated TNF-α and inhibited IL-10 production, while Mexican sample exerted the opposite effect, inhibiting TNF-α and stimulating IL-10 production. The major compounds found in Brazilian, Cuban and Mexican propolis samples were artepillin C, isoflavonoids and pinocembrin, respectively. Brazilian, Cuban and Mexican propolis contained different components that may exert pro- and anti-inflammatory activity depending on concentration, what may provide a novel approach to the development of immunomodulatory drugs containing propolis. © 2015 Royal Pharmaceutical Society.

  8. Cytokine expression and ultrastructural alterations in fresh-frozen, freeze-dried and γ-irradiated human amniotic membranes.

    PubMed

    Paolin, Adolfo; Trojan, Diletta; Leonardi, Antonio; Mellone, Stefano; Volpe, Antonio; Orlandi, Augusto; Cogliati, Elisa

    2016-09-01

    The aim of this work was to compare the effects on human amniotic membrane of freeze-drying and γ-irradiation at doses of 10, 20 and 30 kGy, with freezing. For this purpose, nine cytokines (interleukin 10, platelet-derived growth factor-AA, platelet-derived growth factor-BB, basic fibroblast growth factor, epidermal growth factor, transforming growth factor beta 1, and tissue inhibitors of metalloproteinase-1, -2, and -4) were titrated in 5 different preparations for each of 3 amniotic membranes included in the study. In addition, the extracellular matrix structure of each sample was assessed by transmission electron microscopy. While freeze-drying did not seem to affect the biological structure or cytokine content of the different amniotic membrane samples, γ-irradiation led to a significant decrease in the tissue inhibitors of metalloproteinase-4, basic fibroblast growth factor and epidermal growth factor, and induced structural damage to the epithelium, basement membrane and lamina densa. The higher the irradiation dose the more severe the damage to the amniotic membrane structure. In conclusion, the Authors recommend processing amniotic membrane under sterile conditions to guarantee safety at every step rather than final sterilization with γ-irradiation, thereby avoiding alteration to the biological characteristics of the amniotic membrane.

  9. Phagocytosis and cytokine response to rough and smooth colony variants of Mycobacterium abscessus by human peripheral blood mononuclear cells.

    PubMed

    Jönsson, Bodil; Ridell, Malin; Wold, Agnes E

    2013-01-01

    Mycobacterium abscessus is a non-tuberculous mycobacteria able to cause opportunistic infections in selected patient groups. During the last decades it has emerged as a cause of chronic pulmonary infection in patients with cystic fibrosis (CF). M. abscessus strains exhibit either smooth or rough colony morphology. Strains exhibiting the rough phenotype more often cause pulmonary infections in CF patients than did the smooth ones. Here, we examined phagocytosis and production of cytokines by human peripheral blood mononuclear cells, in response to M. abscessus strains with smooth and rough colony phenotype. The rough isolates all formed multicellular cords, similar to what is observed in Mycobacterium tuberculosis. Monocytes were generally unable to internalize these rough cord isolates, in contrast with the smooth ones. Furthermore, the rough M. abscessus strains induced a distinct cytokine profile differing from that induced by the smooth ones. Rough isolates induced significantly less IL-10 and tumour necrosis factor compared to smooth strains, but more IL-1β. Both varieties induced equal amounts of IFN-γ, IL-17, IL-23, IL-6, IL-8 and equally little IL-12. The ability to withstand phagocytosis might be a virulence factor contributing to the capacity of rough M. abscessus strains to give persistent pulmonary infections. © 2012 The Authors APMIS © 2012 APMIS.

  10. Effects of airborne World Trade Center dust on cytokine release by primary human lung cells in vitro.

    PubMed

    Payne, J P; Kemp, S J; Dewar, A; Goldstraw, P; Kendall, M; Chen, L C; Tetley, T D

    2004-05-01

    There are continuing concerns regarding the respiratory health effects of airborne particulate matter (PM) after the destruction of the World Trade Centre (WTC). We examined cytokine (interleukin [IL]-8, IL-6, tumor necrosis factor-alpha) release by primary human lung alveolar macrophages (AM) and type II epithelial cells after exposure to WTC PM2.5 (indoor and outdoor), PM10-2.5 (indoor), and PM53-10 (outdoor), fractionated from settled dusts within 2 months of the incident. There was an increase in AM cytokine/chemokine release at 5 and/or 50 microg/well WTC PM, which fell at 500 microg/well. Type II cells did not release tumor necrosis factor-alpha, and the increase in IL-8 and IL-6, although significant, was lower than that of AM. Respirable PM generated by the WTC collapse stimulates inflammatory mediator release by lung cells, which may contribute to the increased incidence of respiratory illness since September 11th 2001.

  11. Human bronchial epithelial cells injury and cytokine production induced by Tityus serrulatus scorpion venom: An in vitro study.

    PubMed

    Rigoni, Vera Lucia Silva; Kwasniewski, Fabio H; Vieira, Rodolfo Paula; Linhares, Ingrid Sestrem; da Silva, Joelmir Lucena Veiga; Nogueira-Pedro, Amanda; Zamuner, Stella Regina

    2016-09-15

    Tityus serrulatus is the scorpion specie responsible for the majority of scorpion sting accidents in Brazil. Symptoms of envenomation by Tityus serrulatus range from local pain to severe systemic reactions such as cardiac dysfunction and pulmonary edema. Thus, this study has evaluated the participation of bronchial epithelial cells in the pulmonary effects of Tityus serrulatus scorpion venom (Tsv). Human bronchial epithelial cell line BEAS-2B were utilized as a model target and were incubated with Tsv (10 or 50 μg/mL) for 1, 3, 6 and 24 h. Effects on cellular response of venom-induce cytotoxicity were examined including cell viability, cell integrity, cell morphology, apoptosis/necrosis as well as cell activation through the release of pro-inflammatory cytokines IL-1β, IL-6 and IL-8. Tsv caused a decrease in cell viability at 10 and 50 μg/mL, which was confirmed by lactate dehydrogenase (LDH) measurement. Flow cytometry analyses revealed necrosis as the main cell death pathway caused by Tsv. Furthermore, Tsv induced the release of IL-1β, IL-6 and IL-8. Altogether, these results demonstrate that Tsv induces cytotoxic effects on bronchial epithelial cells, involving necrosis and release of pro-inflammatory cytokines, suggesting that bronchial epithelial cells may play a role in the pulmonary injury caused by Tsv. Copyright © 2016 Elsevier Ltd. All rights reserved.

  12. ProNGF, a cytokine induced after myocardial infarction in humans, targets pericytes to promote microvascular damage and activation

    PubMed Central

    Siao, Chia-Jen; Lorentz, Christina U.; Kermani, Pouneh; Marinic, Tina; Carter, John; McGrath, Kelly; Padow, Victoria A.; Mark, Willie; Falcone, Domenick J.; Cohen-Gould, Leona; Parrish, Diana C.; Habecker, Beth A.; Nykjaer, Anders; Ellenson, Lora H.; Tessarollo, Lino

    2012-01-01

    Treatment of acute cardiac ischemia focuses on reestablishment of blood flow in coronary arteries. However, impaired microvascular perfusion damages peri-infarct tissue, despite arterial patency. Identification of cytokines that induce microvascular dysfunction would provide new targets to limit microvascular damage. Pro–nerve growth factor (NGF), the precursor of NGF, is a well characterized cytokine in the brain induced by injury. ProNGF activates p75 neurotrophin receptor (p75NTR) and sortilin receptors to mediate proapoptotic responses. We describe induction of proNGF by cardiomyocytes, and p75NTR in human arterioles after fatal myocardial infarction, but not with unrelated pathologies. After mouse cardiac ischemia-reperfusion (I-R) injury, rapid up-regulation of proNGF by cardiomyocytes and p75NTR by microvascular pericytes is observed. To identify proNGF actions, we generated a mouse expressing a mutant Ngf allele with impaired processing of proNGF to mature NGF. The proNGF-expressing mouse exhibits cardiac microvascular endothelial activation, a decrease in pericyte process length, and increased vascular permeability, leading to lethal cardiomyopathy in adulthood. Deletion of p75NTR in proNGF-expressing mice rescues the phenotype, confirming the importance of p75NTR-expressing pericytes in the development of microvascular injury. Furthermore, deficiency in p75NTR limits infarct size after I-R. These studies identify novel, nonneuronal actions for proNGF and suggest that proNGF represents a new target to limit microvascular dysfunction. PMID:23091165

  13. ProNGF, a cytokine induced after myocardial infarction in humans, targets pericytes to promote microvascular damage and activation.

    PubMed

    Siao, Chia-Jen; Lorentz, Christina U; Kermani, Pouneh; Marinic, Tina; Carter, John; McGrath, Kelly; Padow, Victoria A; Mark, Willie; Falcone, Domenick J; Cohen-Gould, Leona; Parrish, Diana C; Habecker, Beth A; Nykjaer, Anders; Ellenson, Lora H; Tessarollo, Lino; Hempstead, Barbara L

    2012-11-19

    Treatment of acute cardiac ischemia focuses on reestablishment of blood flow in coronary arteries. However, impaired microvascular perfusion damages peri-infarct tissue, despite arterial patency. Identification of cytokines that induce microvascular dysfunction would provide new targets to limit microvascular damage. Pro-nerve growth factor (NGF), the precursor of NGF, is a well characterized cytokine in the brain induced by injury. ProNGF activates p75 neurotrophin receptor (p75(NTR)) and sortilin receptors to mediate proapoptotic responses. We describe induction of proNGF by cardiomyocytes, and p75(NTR) in human arterioles after fatal myocardial infarction, but not with unrelated pathologies. After mouse cardiac ischemia-reperfusion (I-R) injury, rapid up-regulation of proNGF by cardiomyocytes and p75(NTR) by microvascular pericytes is observed. To identify proNGF actions, we generated a mouse expressing a mutant Ngf allele with impaired processing of proNGF to mature NGF. The proNGF-expressing mouse exhibits cardiac microvascular endothelial activation, a decrease in pericyte process length, and increased vascular permeability, leading to lethal cardiomyopathy in adulthood. Deletion of p75(NTR) in proNGF-expressing mice rescues the phenotype, confirming the importance of p75(NTR)-expressing pericytes in the development of microvascular injury. Furthermore, deficiency in p75(NTR) limits infarct size after I-R. These studies identify novel, nonneuronal actions for proNGF and suggest that proNGF represents a new target to limit microvascular dysfunction.

  14. Cytokine Profiles in Human Metapneumovirus Infected Children: Identification of Genes Involved in the Antiviral Response and Pathogenesis

    PubMed Central

    Malmo, Jostein; Moe, Nina; Krokstad, Sidsel; Ryan, Liv; Loevenich, Simon; Johnsen, Ingvild B.; Espevik, Terje; Nordbø, Svein Arne; Døllner, Henrik; Anthonsen, Marit W.

    2016-01-01

    Human metapneumovirus (hMPV) causes severe airway infection in children that may be caused by an unfavorable immune response. The nature of the innate immune response to hMPV in naturally occurring infections in children is largely undescribed, and it is unknown if inflammasome activation is implicated in disease pathogenesis. We examined nasopharynx aspirates and blood samples from hMPV-infected children without detectable co-infections. The expression of inflammatory and antiviral genes were measured in nasal airway secretions by relative mRNA quantification while blood plasma proteins were determined by a multiplex immunoassay. Several genes were significantly up-regulated at mRNA and protein level in the hMPV infected children. Most apparent was the expression of the chemokine IP-10, the pro-inflammatory cytokine IL-18 in addition to the interferon inducible gene ISG54. Interestingly, children experiencing more severe disease, as indicated by a severity index, had significantly more often up-regulation of the inflammasome-associated genes IL-1β and NLRP3. Overall, our data point to cytokines, particularly inflammasome-associated, that might be important in hMPV mediated lung disease and the antiviral response in children with severe infection. Our study is the first to demonstrate that inflammasome components are associated with increased illness severity in hMPV-infected children. PMID:27171557

  15. Human intestinal epithelial cells produce proinflammatory cytokines in response to infection in a SCID mouse-human intestinal xenograft model of amebiasis.

    PubMed Central

    Seydel, K B; Li, E; Swanson, P E; Stanley, S L

    1997-01-01

    The protozoan parasite Entamoeba histolytica causes amebic dysentery and amebic liver abscess, diseases associated with significant morbidity and mortality worldwide. E. histolytica infection appears to involve the initial attachment of amebic trophozoites to intestinal epithelial cells, followed by lysis of these cells and subsequent invasion into the submucosa. A recent in vitro study (L. Eckmann, S. L. Reed, J. R. Smith, and M. F. Kagnoff, J. Clin. Invest. 96:1269-1279, 1995) demonstrated that incubation of E. histolytica trophozoites with epithelial cell lines results in epithelial cell production of inflammatory cytokines, including interleukin-1 (IL-1) and IL-8, suggesting that intestinal epithelial cell production of cytokines might play a role in the inflammatory response and tissue damage seen in intestinal amebiasis. To determine whether intestinal epithelial cell production of IL-1 and IL-8 occurs in response to E. histolytica infection in vivo and as an approach to studying the specific interactions between amebic trophozoites and human intestine, we used a SCID mouse-human intestinal xenograft (SCID-HU-INT) model of disease, where human intestinal xenografts were infected with virulent E. histolytica trophozoites. Infection of xenografts with E. histolytica trophozoites resulted in extensive tissue damage, which was associated with the development of an early inflammatory response composed primarily of neutrophils. Using oligonucleotide primers that specifically amplify human IL-1beta and IL-8, we could demonstrate by reverse transcription PCR that mRNA for both IL-1beta and IL-8 is produced by human intestinal xenografts in response to amebic infection. The increase in human intestinal IL-1beta and IL-8 in response to invasive amebiasis was confirmed by enzyme-linked immunosorbent assays specific for human IL-1beta and IL-8. Using immunohistochemistry, we confirmed that human intestinal epithelial cells were the source of IL-8 in infected xenografts

  16. 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.

  17. A novel, dual cytokine-secretion assay for the purification of human Th22 cells that do not co-produce IL-17A.

    PubMed

    Wawrzyniak, M; Ochsner, U; Wirz, O; Wawrzyniak, P; van de Veen, W; Akdis, C A; Akdis, M

    2016-01-01

    Interleukin-22 is produced by certain T helper cells subsets (Th17, Th22) and at lower levels by γ-δ T cells, NKT and innate lymphoid cells. Th22 cells are unique immune cells that regulate tissue responses by IL-22 production. The exact discrimination between Th17 cells that co-produce IL-22 and single IL-22-producing Th22 cells has not been possible until the present study. Isolation of pure Th22 cells without co-expression of cytokines of other T-cell subsets is essential to better understand their function in humans. The aim of this study is the isolation and characterization of viable, human IL-22-producing CD4+ T cells that do not produce IL-17A. Isolation of viable Th22 cells was performed with the combination of two cytokine secretion assays detecting IL-17A- and IL-22-producing cells in a single purification step. The newly developed cytokine secretion assay consists of anti-IL-22 and anti-IL-17A catch antibodies, which via biotin-streptavidin interaction are bound to the biotinylated surface of the target cell, and anti-IL-22 and IL-17A detection antibody labelled with a fluorescent dye, which detects cytokines bound to these catch antibodies. A unique population of human Th22 cells, which do not produce IL-17A, was sorted, and cytokine expression pattern was confirmed by quantitative PCR analysis and ELISA. The presented technique allows the detection and isolation of pure human Th22 cells. This technique may allow the purification of any single cytokine-producing cell subset, and the combination of several different cytokine secretion assays can be used to purify and characterize novel and unique cell subsets. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  18. Involvement of Reactive Oxygen Species in Brominated Diphenyl Ether-47-induced Inflammatory Cytokine Release from Human Extravillous Trophoblasts in vitro

    PubMed Central

    Park, Hae-Ryung; Kamau, Patricia W.; Loch-Caruso, Rita

    2014-01-01

    Polybrominated diphenyl ethers (PBDEs) are widely used flame retardant compounds. Brominated diphenyl ether (BDE)-47 is one of the most prevalent PBDE congeners found in human breast milk, serum and placenta. Despite the presence of PBDEs in human placenta, effects of PBDEs on placental cell function are poorly understood. The present study investigated BDE-47-induced reactive oxygen species (ROS) formation and its role in BDE-47-stimulated proinflammatory cytokine release in a first trimester human extravillous trophoblast cell line, HTR-8/SVneo. Exposure of HTR-8/SVneo cells for 4 h to 20 μM BDE-47 increased ROS generation 1.7 fold as measured by the dichlorofluorescein (DCF) assay. Likewise, superoxide anion production increased approximately 5 fold at 10 and 15 μM and 9 fold at 20 μM BDE-47 with a 1-h exposure, as measured by cytochrome c reduction. BDE-47 (10, 15 and 20 μM) decreased the mitochondrial membrane potential by 47–64.5% at 4, 8 and 24 h as assessed with the fluorescent probe Rh123. Treatment with 15 and 20 μM BDE-47 stimulated cellular release and mRNA expression of IL-6 and IL-8 after 12 and 24 h exposures: the greatest increases were a 35-fold increased mRNA expression at 12 h and a 12-fold increased protein concentration at 24 h for IL-6. Antioxidant treatments (deferoxamine mesylate, (±)α-tocopherol, or tempol) suppressed BDE-47-stimulated IL-6 release by 54.1%, 56.3% and 37.7%, respectively, implicating a role for ROS in regulation of inflammatory pathways in HTR-8/SVneo cells. Solvent (DMSO) controls exhibited statistically significantly decreased responses compared with non-treated controls for IL-6 release and IL-8 mRNA expression, but these responses were not consistent across experiments and times. Nonetheless, it is possible that DMSO (used to dissolve BDE-47) may have attenuated the stimulatory actions of BDE-47 on cytokine responses. Because abnormal activation of proinflammatory responses can disrupt trophoblast functions

  19. [The effect of kefir consumption on human immune system: a cytokine study].

    PubMed

    Adiloğlu, Ali Kudret; Gönülateş, Nurettin; Işler, Mehmet; Senol, Altuğ

    2013-04-01

    The systemic effects of bioactive peptides which are produced by the fermentation of milk via the microorganisms found in kefir have been the subject of interest in recent years. Bioactive peptides activate innate immunity by stimulating macrophages, increasing phagocytosis, augmenting NO and cytokine production and boosting the lumen levels of IgG and IgA+ B-lymphocytes. The aim of the present study was to determine the serum cytokine profiles of healthy volunteers after kefir consumption to evaluate helper T (TH) cell polarization and to bring out the effects on native and allergic immune responses. The study was designed as a prospective and self-controlled study. A total of 18 healthy volunteers (age range: 20-40 yrs, mean age: 35.5 ± 7.38 yrs) from a university hospital staff were recruited to the study, with the approval of ethical board and informed consent. The body mass indices of all participants were between normal range (20.10-25.70 kg/m2). After two weeks of a diet free from fermented products, the participants consumed 200 mL kefir daily, for six weeks. Kefir product was prepared by using kefir starter culture (Danisco Biolacta Sp - 05223B 10001, Poland) which contains Lactobacillus spp., Leuconostoc spp., Lactococcus lactis ssp. lactis and Streptococcus termophilus, an overnight incubation at 26°C, and consumed freshly. Fasting blood samples of subjects were collected just before kefir use (0th week), at the end of the 3rd and 6th weeks of kefir consumption, and three weeks after cessation of kefir usage (9th week). Serum TNF-a, IL-1, IL-5, IL-8 and TGF-β levels were measured by using commercial ELISA kits (BioSource, Belgium and Invitrogen, USA). Hemoglobin, serum creatinine and ALT levels of all subjects were also determined for follow-up. All volunteers completed the study period without any problem and declared no complaint. Hemoglobin, creatinine and ALT levels did not change with kefir consumption. Serum IL-8 levels were decreased at 3rd and

  20. Teaching and Learning Children's Human Rights: A Research Synthesis

    ERIC Educational Resources Information Center

    Brantefors, Lotta; Quennerstedt, Ann

    2016-01-01

    The study presented in this paper is a research synthesis examining how issues relating to the teaching and learning of children's human rights have been approached in educational research. Drawing theoretically on the European Didaktik tradition, the purpose of the paper is to map and synthesise the educational interest in children's rights…

  1. Differences in gene expression and cytokine production by crystalline vs. amorphous silica in human lung epithelial cells

    PubMed Central

    2012-01-01

    Background Exposure to respirable crystalline silica particles, as opposed to amorphous silica, is associated with lung inflammation, pulmonary fibrosis (silicosis), and potentially with lung cancer. We used Affymetrix/GeneSifter microarray analysis to determine whether gene expression profiles differed in a human bronchial epithelial cell line (BEAS 2B) exposed to cristobalite vs. amorphous silica particles at non-toxic and equal surface areas (75 and 150 × 106μm2/cm2). Bio-Plex analysis was also used to determine profiles of secreted cytokines and chemokines in response to both particles. Finally, primary human bronchial epithelial cells (NHBE) were used to comparatively assess silica particle-induced alterations in gene expression. Results Microarray analysis at 24 hours in BEAS 2B revealed 333 and 631 significant alterations in gene expression induced by cristobalite at low (75) and high (150 × 106μm2/cm2) amounts, respectively (p < 0.05/cut off ≥ 2.0-fold change). Exposure to amorphous silica micro-particles at high amounts (150 × 106μm2/cm2) induced 108 significant gene changes. Bio-Plex analysis of 27 human cytokines and chemokines revealed 9 secreted mediators (p < 0.05) induced by crystalline silica, but none were induced by amorphous silica. QRT-PCR revealed that cristobalite selectively up-regulated stress-related genes and cytokines (FOS, ATF3, IL6 and IL8) early and over time (2, 4, 8, and 24 h). Patterns of gene expression in NHBE cells were similar overall to BEAS 2B cells. At 75 × 106μm2/cm2, there were 339 significant alterations in gene expression induced by cristobalite and 42 by amorphous silica. Comparison of genes in response to cristobalite (75 × 106μm2/cm2) revealed 60 common, significant gene alterations in NHBE and BEAS 2B cells. Conclusions Cristobalite silica, as compared to synthetic amorphous silica particles at equal surface area concentrations, had comparable effects on the viability of human bronchial epithelial cells

  2. Effect of pro-inflammatory cytokine stimulation on human breast cancer: implications of chemokine receptor expression in cancer metastasis.

    PubMed

    Valdivia-Silva, Julio E; Franco-Barraza, Janusz; Silva, Ana Luisa Esparza; Pont, Gisela Du; Soldevila, Gloria; Meza, Isaura; García-Zepeda, Eduardo A

    2009-10-08

    Interactions between tumour cells and microenvironments may affect their growth and metastasis formation. In search for a better understanding of the role of cellular mediators in the progression of cancer, we investigated the effect of pro-inflammatory cytokines IL-1, IL-6, TNF-alpha and IFN-gamma on the regulation of expression of chemokine receptors CXCR4, CXCR2, CX3CR1, CCR9, and CCR5 in the human breast cancer cell line MCF-7. Our results showed that IL-1 increased CXCR4 expression whereas TNF-alpha increased CX3CR1, CCR9 and CCR5. Interestingly, this regulation was not homogeneous, emphasizing the inherent heterogeneity in cancer that may be responsive to specific inflammatory microenvironments.

  3. Impact of antigen-presenting cells on cytokine profiles of human Th clones established after stimulation with Mycobacterium tuberculosis antigens.

    PubMed Central

    Conradt, P; Kaufmann, S H

    1995-01-01

    Human T cells reactive with mycobacterial antigens are generally considered to correlate with a Th1 cytokine profile. Our data show that, in addition, Th0 and Th2 clones develop in bulk culture with appropriate antigen-presenting cells before cloning. CD4+ blasts activated by mycobacterial antigens were cloned, and their mRNA patterns for the interleukins (IL) IL-2, IL-4, IL-5, IL-6, and IL-10 and gamma interferon were characterized by reverse-transcribed PCR. Nonadherent, nonrosetting, enriched peripheral blood mononuclear cells promoted development of Th0; after further depletion of monocytes and natural killer cells, Th2 clones were also found. Epstein-Barr virus-transformed B cells, with specificity for the stimulating antigen, increased the proportion of Th2 clones. PMID:7729923

  4. Cytokines, Chemokines, and Growth Factors in Banked Human Donor Milk for Preterm Infants

    PubMed Central

    Groer, Maureen; Duffy, Allyson; Morse, Shannon; Kane, Bradley; Zaritt, Judy; Roberts, Shari; Ashmeade, Terri

    2014-01-01

    Background There has been a recent increase in availability of banked donor milk for feeding of preterm infants. This milk is pooled from donations to milk banks from carefully screened lactating women. The milk is then pasteurized by the Holder method to remove all microbes. The processed milk is frozen, banked, and sold to neonatal intensive care units (NICUs). The nutrient bioavailability of banked donor milk has been described, but little is known about preservation of immune components such as cytokines, chemokines, and growth factors (CCGF). Objective The objective was to compare CCGF in banked donor milk with mother's own milk (MOM). Methods Aliquots (0.5 mL) were collected daily from MOM pumped by 45 mothers of NICU-admitted infants weighing < 1500 grams at birth. All daily aliquots of each mother's milk were pooled each week during 6 weeks of an infant's NICU stay or for as long as the mother provided MOM. The weekly pooled milk was measured for a panel of CCGF through multiplexing using magnetic beads and a MAGPIX instrument. Banked donor milk samples (n = 25) were handled and measured in the same way as MOM. Results Multiplex analysis revealed that there were levels of CCGF in banked donor milk samples comparable to values obtained from MOM after 6 weeks of lactation. Conclusion These data suggest that many important CCGF are not destroyed by Holder pasteurization. PMID:24663954

  5. Cytoskeletal reorganizations in human umbilical vein endothelial cells as a result of cytokine exposure

    SciTech Connect

    Molony, L.; Armstrong, L. )

    1991-09-01

    Treatment of HUVECs in culture with several cytokines and phorbol esters caused reorganizations of the actin and microtubule networks, as well as a redistribution of focal contract proteins. However, expression of the cytoskeletal proteins which link cells, via integrins, to the substrate, was not significantly affected. Indirect immunofluorescence microscopy of endothelial cells after treatment with interleukin-1 alpha and beta, gamma-interferon, tumor necrosis factor (TNF), phorbol 12-myristate 13-acetate, and phorbol 12,13-dibutyrate allowed us to observe reductions in the areas of cell-cell contact, redistribution of the stress fiber network, and concomitant changes in focal contacts. Microtubule arrays in TNF-treated cells became bundled. Phorbol esters induced formation of microtubule organizing centers not seen in resting or TNF-treated HUVECs. Talin was distributed along stress fibers and not exclusively in focal contacts. Vitronectin receptor was observed in focal contacts, occasionally at cell-cell contacts, and in vesicular structures close to the lumenal surface, after both types of treatment. Although these morphological changes were easily observed by indirect immunofluorescence, no quantitative differences in specific cytoskeletal proteins were detected by immunoblots and (35S)cysteine metabolic labeling experiments.

  6. Acidic sphingomyelinase induced by electrophiles promotes proinflammatory cytokine production in human bladder carcinoma ECV-304 cells.

    PubMed

    Kumagai, Takeshi; Ishino, Tomohiro; Nakagawa, Yasuhito

    2012-03-01

    Electrophiles in environmental pollutants or cigarette smoke are high risk factors for various diseases caused by cell injuries such as apoptosis and inflammation. Here we show that electrophilic compounds such as diethyl malate (DEM), methyl mercury and cigarette smoke extracts significantly enhanced the expression of acidic sphingomyelinase (ASMase). ASMase activity and the amount of ceramide of DEM-treated cells were approximately 6 times and 4 times higher than these of non-treated cells, respectively. Moreover, we found that DEM pretreatment enhanced the production of IL-6 induced by TNF-α. Knockdown of ASMase attenuated the enhancement of TNF-α-dependent IL-6 production. On the other hand, enhancement of TNF-α-induced IL-6 production was observed in ASMase-overexpressing cells without DEM. Fractionation of the lipid raft revealed that the TNF receptor 1 (TNFR1) was migrated into the lipid raft in DEM-treated cells or ASMase-overexpressing cells. The TNF-α-induced IL-6 expression required the clustering of TNFR1 since IL-6 expression were decreased by the destruction of the lipid raft with filipin. These results demonstrated a new role for ASMase in the acceleration of the production of TNF-induced IL-6 as a pro-inflammatory cytokine and indicated that electrophiles could potentiate inflammation response by up-regulating of ASMase expression following formation of lipid rafts. Copyright © 2012 Elsevier Inc. All rights reserved.

  7. 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.

  8. Pathogenesis of Streptococcus urinary tract infection depends on bacterial strain and β-hemolysin/cytolysin that mediates cytotoxicity, cytokine synthesis, inflammation and virulence

    PubMed Central

    Leclercq, Sophie Y.; Sullivan, Matthew J.; Ipe, Deepak S.; Smith, Joshua P.; Cripps, Allan W.; Ulett, Glen C.

    2016-01-01

    Streptococcus agalactiae can cause urinary tract infection (UTI) including cystitis and asymptomatic bacteriuria (ABU). The early host-pathogen interactions that occur during S. agalactiae UTI and subsequent mechanisms of disease pathogenesis are poorly defined. Here, we define the early interactions between human bladder urothelial cells, monocyte-derived macrophages, and mouse bladder using uropathogenic S. agalactiae (UPSA) 807 and ABU-causing S. agalactiae (ABSA) 834 strains. UPSA 807 adhered, invaded and killed bladder urothelial cells more efficiently compared to ABSA 834 via mechanisms including low-level caspase-3 activation, and cytolysis, according to lactate dehydrogenase release measures and cell viability. Severe UPSA 807-induced cytotoxicity was mediated entirely by the bacterial β-hemolysin/cytolysin (β-H/C) because an β-H/C-deficient UPSA 807 isogenic mutant, UPSA 807ΔcylE, was not cytotoxic in vitro; the mutant was also significantly attenuated for colonization in the bladder in vivo. Analysis of infection-induced cytokines, including IL-8, IL-1β, IL-6 and TNF-α in vitro and in vivo revealed that cytokine and chemokine responses were dependent on expression of β-H/C that also elicited severe bladder neutrophilia. Thus, virulence of UPSA 807 encompasses adhesion to, invasion of and killing of bladder cells, pro-inflammatory cytokine/chemokine responses that elicit neutrophil infiltration, and β-H/C-mediated subversion of innate immune-mediated bacterial clearance from the bladder. PMID:27383371

  9. Simvastatin inhibits the expression of inflammatory cytokines and cell adhesion molecules induced by LPS in human dental pulp cells.

    PubMed

    Jung, J Y; Woo, S M; Kim, W J; Lee, B N; Nör, J E; Min, K S; Choi, C H; Koh, J T; Lee, K J; Hwang, Y C

    2017-04-01

    To investigate the effect of simvastatin on lipopolysaccharide (LPS)-stimulated inflammatory cytokines, cell adhesion molecules and nuclear factor-κB (NF-κB) transcription factors in human dental pulp cells (HDPCs). The effect of LPS and simvastatin on human dental pulp cell (HDPCs) viability was measured using a 3-[4, 5-dimethylthiazol-2-yl]-2, 5 diphenyltetrazolium bromide (MTT) assay. The expression of inflammatory cytokines and cell adhesion molecules was evaluated by reverse-transcription polymerase chain reaction (RT-PCR), enzyme-linked immunosorbent assay (ELISA) and Western blot analysis. NF-κB transcription factors were evaluated by Western blot analysis. Statistical analysis was performed with analysis of variance (anova). The viability of cells exposed to different concentrations of E. coli LPS, P. gingivalis LPS and simvastatin was not significantly different compared with that of control cells (P > 0.05). LPS significantly increased interleukin (IL)-1β (P < 0.05) and IL-6 mRNA expression (P < 0.05) and vascular cell adhesion molecule-1 (VCAM-1) (P < 0.05) and intercellular adhesion molecule-1 (ICAM-1) protein expression (P < 0.05) in HDPCs. Treatment with simvastatin significantly attenuated LPS-stimulated production of IL-1β, IL-6, VCAM-1 and ICAM-1 (P < 0.05). Treatment with simvastatin decreased LPS-induced expression of p65 and phosphorylation of IκB and also significantly decreased the phosphorylation of p65 and IκB in the cytoplasm and the level of p65 in the nucleus (P < 0.05). Simvastatin has a suppressing effect on LPS-induced inflammatory cytokine, cell adhesion molecules and NF-κB transcription factors in HDPCs. Therefore, simvastatin might be a useful candidate as a pulp-capping agent in vital pulp therapy. © 2016 International Endodontic Journal. Published by John Wiley & Sons Ltd.

  10. β2-Adrenoceptor agonists inhibit release of eosinophil-activating cytokines from human airway smooth muscle cells

    PubMed Central

    Hallsworth, Matthew P; Twort, Charles H C; Lee, Tak H; Hirst, Stuart J

    2001-01-01

    Airway smooth muscle (ASM) is a potential source of multiple pro-inflammatory cytokines during airway inflammation. β-Adrenoceptor agonist hyporesponsiveness is a characteristic feature of asthma, and interleukin (IL)-1β and tumour necrosis factor (TNF)-α are implicated in its cause. Here, the capacity of β-adrenoceptor agonists to prevent release of GM-CSF, RANTES, eotaxin and IL-8, elicited by IL-1β or TNFα, was examined in human ASM cells. Isoprenaline (∼EC50 150 nM), a non-selective β-adrenoceptor agonist, and salbutamol (∼EC50 25 nM), a selective β2-adrenoceptor agonist, attenuated release of GM-CSF, RANTES and eotaxin, but not IL-8 (EC50 >1 μM). The maximum extent of attenuation was RANTES ⩾ eotaxin > GM-CSF >> IL-8, and was prevented by either propranolol (1 μM), a non-selective β-adrenoceptor antagonist, or ICI 118511 (IC50 15 nM), a selective β2-adrenoceptor antagonist. The cyclic AMP-elevating agents, dibutyryl cyclic AMP (∼EC50 135 μM), forskolin (∼EC50 530 nM) and cholera toxin (∼EC50 575 pg ml−1) abolished IL-1β-induced release of GM-CSF, RANTES and eotaxin, but not IL-8. IL-1β (1 ng ml−1) attenuated early increases (up to 1 h) in cyclic AMP formation induced by salbutamol (1 μM), but not by forskolin (10 μM). The cyclo-oxygenase inhibitor, indomethacin (1 μM) prevented later increases (3 – 12 h) in IL-1β-stimulated cyclic AMP content, but did not prevent the attenuation by salbutamol of IL-1β-induced cytokine release. We conclude in human ASM cells that activation of β2-adrenoceptors and generation of cyclic AMP is negatively-linked to the release, elicited by IL-1β or TNFα, of eosinophil-activating cytokines such as GM-CSF, RANTES and eotaxin, but not IL-8. PMID:11159726

  11. Inflammatory gene regulatory networks in amnion cells following cytokine stimulation: translational systems approach to modeling human parturition.

    PubMed

    Li, Ruth; Ackerman, William E; Summerfield, Taryn L; Yu, Lianbo; Gulati, Parul; Zhang, Jie; Huang, Kun; Romero, Roberto; Kniss, Douglas A

    2011-01-01

    A majority of the studies examining the molecular regulation of human labor have been conducted using single gene approaches. While the technology to produce multi-dimensional datasets is readily available, the means for facile analysis of such data are limited. The objective of this study was to develop a systems approach to infer regulatory mechanisms governing global gene expression in cytokine-challenged cells in vitro, and to apply these methods to predict gene regulatory networks (GRNs) in intrauterine tissues during term parturition. To this end, microarray analysis was applied to human amnion mesenchymal cells (AMCs) stimulated with interleukin-1β, and differentially expressed transcripts were subjected to hierarchical clustering, temporal expression profiling, and motif enrichment analysis, from which a GRN was constructed. These methods were then a