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Sample records for human cytokine synthesis

  1. 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.

  2. 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

  3. Leptin enhances ICAM-1 expression, induces migration and cytokine synthesis, and prolongs survival of human airway epithelial cells.

    PubMed

    Suzukawa, Maho; Koketsu, Rikiya; Baba, Shintaro; Igarashi, Sayaka; Nagase, Hiroyuki; Yamaguchi, Masao; Matsutani, Noriyuki; Kawamura, Masafumi; Shoji, Shunsuke; Hebisawa, Akira; Ohta, Ken

    2015-10-15

    There is rising interest in how obesity affects respiratory diseases, since epidemiological findings indicate a strong relationship between the two conditions. Leptin is a potent adipokine produced mainly by adipocytes. It regulates energy storage and expenditure and also induces inflammation. Previous studies have shown that leptin is able to activate inflammatory cells such as lymphocytes and granulocytes, but little is known about its effect on lung structural cells. The present study investigated the effects of leptin on human airway epithelial cells by using human primary airway epithelial cells and a human airway epithelial cell line, BEAS-2B. Flow cytometry showed enhanced ICAM-1 expression by both of those cells in response to leptin, and that effect was abrogated by dexamethasone or NF-κB inhibitor. Flow cytometry and quantitative PCR showed that airway epithelial cells expressed leptin receptor (Ob-R), whose expression level was downregulated by leptin itself. Multiplex cytokine analysis demonstrated enhanced production of CCL11, G-CSF, VEGF, and IL-6 by BEAS-2B cells stimulated with leptin. Furthermore, transfection of Ob-R small interference RNA decreased the effect of leptin on CCL11 production as assessed by quantitative PCR. Finally, leptin induced migration of primary airway epithelial cells toward leptin, suppressed BEAS-2B apoptosis induced with TNF-α and IFN-γ, and enhanced proliferation of primary airway epithelial cells. In summary, leptin was able to directly activate human airway epithelial cells by binding to Ob-R and by NF-κB activation, resulting in upregulation of ICAM-1 expression, induction of CCL11, VEGF, G-CSF, and IL-6 synthesis, induction of migration, inhibition of apoptosis, and enhancement of proliferation. PMID:26276826

  4. Prostaglandin (PG) F2 Alpha Synthesis in Human Subcutaneous and Omental Adipose Tissue: Modulation by Inflammatory Cytokines and Role of the Human Aldose Reductase AKR1B1

    PubMed Central

    Michaud, Andréanne; Lacroix-Pépin, Nicolas; Pelletier, Mélissa; Veilleux, Alain; Noël, Suzanne; Bouchard, Céline; Marceau, Picard; Fortier, Michel A.; Tchernof, André

    2014-01-01

    Introduction PGF2α may be involved in the regulation of adipose tissue function. Objectives 1) To examine PGF2α release by primary preadipocytes, mature adipocytes and whole tissue explants from the subcutaneous and omental fat compartments; 2) To assess which PGF synthase is the most relevant in human adipose tissue. Methods Fat samples were obtained by surgery in women. PGF2α release by preadipocytes, adipocytes and explants under stimulation by TNF-α, IL-1β or both was measured. Messenger RNA expression levels of AKR1B1 and AKR1C3 were measured by RT-PCR in whole adipose tissue and cytokine-treated preadipocytes. The effect of AKR1B1 inhibitor ponalrestat on PGF2α synthesis was investigated. Results PGF2α release was significantly induced in response to cytokines compared to control in omental (p = 0.01) and to a lesser extent in subcutaneous preadipocytes (p = 0.02). Messenger RNA of COX-2 was significantly higher in omental compared to subcutaneous preadipocytes in response to combined TNF-α and IL-1β (p = 0.01). Inflammatory cytokines increased AKR1B1 mRNA expression and protein levels (p≤0.05), but failed to increase expression levels of AKR1C3 in cultured preadipocytes. Accordingly, ponalrestat blunted PGF2α synthesis by preadipocytes in basal and stimulated conditions (p≤0.05). Women with the highest PGF2α release by omental adipocytes had a higher BMI (p = 0.05), waist circumference (p≤0.05) and HOMAir index (p≤0.005) as well as higher mRNA expression of AKR1B1 in omental (p<0.10) and subcutaneous (p≤0.05) adipose tissue compared to women with low omental adipocytes PGF2α release. Positive correlations were observed between mRNA expression of AKR1B1 in both compartments and BMI, waist circumference as well as HOMAir index (p≤0.05 for all). Conclusion PGF2α release by omental mature adipocytes is increased in abdominally obese women. Moreover, COX-2 expression and PGF2α release is particularly responsive to

  5. 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.

  6. Human Bladder Uroepithelial Cells Synergize with Monocytes to Promote IL-10 Synthesis and Other Cytokine Responses to Uropathogenic Escherichia coli

    PubMed Central

    Duell, Benjamin L.; Carey, Alison J.; Dando, Samantha J.; Schembri, Mark A.; Ulett, Glen C.

    2013-01-01

    Urinary tract infections are a major source of morbidity for women and the elderly, with Uropathogenic Escherichia coli (UPEC) being the most prevalent causative pathogen. Studies in recent years have defined a key anti-inflammatory role for Interleukin-10 (IL-10) in urinary tract infection mediated by UPEC and other uropathogens. We investigated the nature of the IL-10-producing interactions between UPEC and host cells by utilising a novel co-culture model that incorporated lymphocytes, mononuclear and uroepithelial cells in histotypic proportions. This co-culture model demonstrated synergistic IL-10 production effects between monocytes and uroepithelial cells following infection with UPEC. Membrane inserts were used to separate the monocyte and uroepithelial cell types during infection and revealed two synergistic IL-10 production effects based on contact-dependent and soluble interactions. Analysis of a comprehensive set of immunologically relevant biomarkers in monocyte-uroepithelial cell co-cultures highlighted that multiple cytokine, chemokine and signalling factors were also produced in a synergistic or antagonistic fashion. These results demonstrate that IL-10 responses to UPEC occur via multiple interactions between several cells types, implying a complex role for infection-related IL-10 during UTI. Development and application of the co-culture model described in this study is thus useful to define the degree of contact dependency of biomarker production to UPEC, and highlights the relevance of histotypic co-cultures in studying complex host-pathogen interactions. PMID:24155979

  7. 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.

  8. Depressed type 1 cytokine synthesis by superantigen-activated CD4+ T cells of women with human papillomavirus-related high-grade squamous intraepithelial lesions.

    PubMed

    Lee, Bang-Ning; Follen, Michele; Shen, De-Yu; Malpica, Anais; Adler-Storthz, Karen; Shearer, William T; Reuben, James M

    2004-03-01

    Carcinoma of the cervix is causally related to infection with the human papillomavirus (HPV), and T cells play a pivotal role in the immune response of the host to rid itself of HPV infection. Therefore, we assessed the T-cell function of women with HPV-related cervical neoplasia against a superantigen, Staphylococcus enterotoxin B (SEB). Each woman provided a cervical brush specimen for HPV DNA testing and Papanicolaou (Pap) smears for the staging of cervical lesions. They also provided a blood specimen for determination of the ability of CD4(+) T and CD8(+) T cells to synthesize Th1 (interleukin-2 [IL-2], gamma interferon [IFN-gamma], and tumor necrosis factor alpha [TNF-alpha]) and Th2 (IL-10) cytokines in response to activation with SEB. Compared with control subjects with self-attested negative Pap smears, women with high-grade squamous intraepithelial lesions (HSIL) had significantly lower percentages of activated CD4(+) T cells that produced IL-2 (P = 0.045), IFN-gamma (P = 0.040), and TNF-alpha (P = 0.015) and a significantly lower percentage of activated CD8(+) T cells that produced IL-2 (P < 0.01). These data indicate that women with HPV-related cervical HSIL show a decrease in Th1 cytokine production by activated CD4(+) T cells and suggested that compromised T-helper functions may negatively impact the function of cytotoxic CD8(+) T cells. PMID:15013969

  9. Inhibitory effects of bisbenzylisoquinolines on synthesis of the inflammatory cytokines interleukin-1 and tumour necrosis factor-alpha

    PubMed Central

    Seow, W. Kim; Nakamura, Kazuhiro; Sugimura, Yukio; Sugimoto, Yukihiro; Yamada, Yasuyuki; Fairlie, David P.

    1993-01-01

    Synthesis of IL-1β and TNFα by human monocytesmacrophages was significantly inhibited by eleven bisbenzylisoquinolines and one half-molecule (benzylisoquinoline), with IC50 values in the μM range. The results indicate that these compounds may have value in the therapy of human diseases where these inflammatory cytokines have a central role in pathogenesis. PMID:18475522

  10. Cytokine-mediated PGE2 expression in human colonic fibroblasts.

    PubMed

    Kim, E C; Zhu, Y; Andersen, V; Sciaky, D; Cao, H J; Meekins, H; Smith, T J; Lance, P

    1998-10-01

    We investigated prostanoid biogenesis in human colonic fibroblasts (CCD-18Co and 5 primary fibroblast cultures) and epithelial cell lines (NCM460, T84, HT-29, and LS 174T) and the effect of PGE2 on fibroblast morphology. Cytokine-stimulated PGE2 production was measured. PGH synthase-1 and -2 (PGHS-1 and -2) protein and mRNA expression were evaluated. Basal PGE2 levels were low in all cell types (0.15-6.47 ng/mg protein). Treatment for 24 h with interleukin-1beta (IL-1beta; 10 ng/ml) or tumor necrosis factor-alpha (50 ng/ml), respectively, elicited maximal 25- and 6-fold inductions of PGE2 synthesis in CCD-18Co cultures and similar results in primary fibroblast cultures; maximal inductions with IL-1beta in colonic epithelial cell lines were from zero to fivefold. Treatment of CCD-18Co fibroblasts with IL-1beta caused maximal 21- and 53-fold increases, respectively, in PGHS-2 protein and mRNA levels without altering PGHS-1 expression. PGE2 (0.1 micromol/l) elicited a dramatic shape change in selected fibroblasts. Colonic fibroblasts are potentially important as cytokine targets and a source of and target for colonic prostanoids in vivo. PMID:9755052

  11. Regulation of sucrase-isomaltase gene expression in human intestinal epithelial cells by inflammatory cytokines.

    PubMed

    Ziambaras, T; Rubin, D C; Perlmutter, D H

    1996-01-12

    Using metabolic labeling techniques in human intestinal epithelial cell lines in tissue culture and in situ hybridization techniques in normal and inflamed (Crohn's) intestine, recent studies have shown that there is synthesis of acute phase proteins in enterocytes. Moreover, these studies have shown that acute phase protein biosynthesis in enterocytes is regulated by inflammatory cytokines in a manner characteristic of the physiologic acute phase response. In the course of these studies it was noticed that one inflammatory cytokine, interleukin-6 (IL-6), mediated selective down-regulation of the enterocyte-specific, differentiation-dependent integral membrane protein sucrase-isomaltase (SI) in the Caco2 intestinal epithelial cell line. In the current study we examined the effect of several other inflammatory cytokines interleukin-1 (IL-1 beta), tumor necrosis factor alpha (TNF alpha), and interferon gamma (IFN gamma) on synthesis of SI in Caco2 cells, examined the possibility that inflammatory cytokines affect the synthesis of other enterocyte integral membrane proteins using lactase as a prototype, and examined the possibility that SI gene expression was down-regulated in villous enterocytes in vivo during the local inflammatory response of Crohn's disease. The results show that IL-6 and IFN gamma each mediate a decrease and TNF alpha mediates an increase in synthesis of SI in Caco2 cells. The magnitude of down-regulation by IL-6 and IFN gamma is significantly greater than the up-regulation by TNF alpha. IL-1 beta has no effect on synthesis of SI. Synthesis of lactase is not affected by any of the cytokines. There is a marked specific decrease in SI gene expression in villous enterocytes in acutely inflamed Crohn's ileum as compared to adjacent uninflamed ileum and normal ileum. Taken together, these data show that inflammatory cytokines have specific and selective effects on the expression of the brush border hydrolase SI in tissue culture and in vivo and

  12. 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...

  13. Cytokine disbalance in common human cancers.

    PubMed

    Culig, Zoran

    2011-02-01

    Interleukin (IL)-6, -4, and -8 levels have been elevated in most patients suffering from prostate, breast, or colon cancer. There is a large body of evidence suggesting that chronic inflammation is one of the etiologic factors in these tumors. IL-6 is a multifunctional cytokine which is known to influence proliferation, apoptosis, and angiogenesis in cancer. Its transcription factor STAT3 is known as an oncogene that is constitutively phosphorylated in these malignancies. However, IL-6-induced STAT3 phosphorylation may result in growth arrest. IL-6 activation of androgen receptor in prostate cancer may yield either tumor cell proliferation or differentiation. Prolonged treatment with IL-6 results in generation of sublines which express a more malignant phenotype. Therapy options against IL-6 have been established and the antibody siltuximab has been applied in preclinical and clinical studies. Recently, investigations of the role of suppressors of cytokine signaling have been carried out. IL-4 and -8 are implicated in regulation of apoptosis, migration, and angiogenesis in cancers associated with chronic inflammation. All cytokines mentioned above regulate cellular events in stem cells. These cells could not be targeted by most conventional cancer therapies. PMID:21167870

  14. Regulation of proinflammatory cytokines in human lung epithelial cells infected with Mycoplasma pneumoniae.

    PubMed

    Yang, Jun; Hooper, W Craig; Phillips, Donald J; Talkington, Deborah F

    2002-07-01

    Mycoplasma pneumoniae is a small bacterium without a cell wall that causes tracheobronchitis and atypical pneumonia in humans. It has also been associated with chronic conditions, such as arthritis, and extrapulmonary complications, such as encephalitis. Although the interaction of mycoplasmas with respiratory epithelial cells is a critical early phase of pathogenesis, little is known about the cascade of events initiated by infection of respiratory epithelial cells by mycoplasmas. Previous studies have shown that M. pneumoniae can induce proinflammatory cytokines in several different study systems including cultured murine and human monocytes. In this study, we demonstrate that M. pneumoniae infection also induces proinflammatory cytokine expression in A549 human lung carcinoma cells. Infection of A549 cells resulted in increased levels of interleukin-8 (IL-8) and tumor necrosis factor alpha mRNA, and both proteins were secreted into culture medium. IL-1 beta mRNA also increased after infection and IL-1 beta protein was synthesized, but it remained intracellular. In contrast, levels of IL-6 and gamma interferon mRNA and protein remained unchanged or undetectable. Using protease digestion and antibody blocking methods, we found that M. pneumoniae cytoadherence is important for the induction of cytokines. On the other hand, while M. pneumoniae protein synthesis and DNA synthesis do not appear to be prerequisites for the induction of cytokine gene expression, A549 cellular de novo protein synthesis is responsible for the increased cytokine protein levels. These results suggest a novel role for lung epithelial cells in the pathogenesis of M. pneumoniae infection and provide a better understanding of M. pneumoniae pathology at the cellular level. PMID:12065506

  15. Cytokine induced expression of programmed death ligands in human neutrophils

    PubMed Central

    Bankey, Paul E.; Banerjee, Sanjib; Zucchiatti, Andrea; De, Mita; Sleem, Rami W.; Lin, Chuen-Fu L.; Miller-Graziano, Carol L.; De, Asit K.

    2010-01-01

    1. Summary Recent evidence indicates that human neutrophils can serve as non-professional antigen presenting cells (APC). Although expression of MHC class II and co-stimulatory molecules on human neutrophils is limited, these molecules can be significantly induced following in vitro exposure to the cytokines IFNγ and GM-CSF. Since professional APCs such as dendritic cells express both co-stimulatory and co-inhibitory molecules for activation and regulation of adaptive immunity, we determined whether cytokines induce increased expression of specific co-signaling molecules on human neutrophils. We report here that circulating human neutrophils express co-inhibitory molecules such as immunoglobulin–like transcript (ILT) 4 and 5, and also comparatively low and highly variable levels of ILT2 and 3, but the expression of these ILTs was not significantly changed by cytokine treatment. In contrast, we demonstrate for the first time that human peripheral blood neutrophils, although do not express the co-inhibitory molecule, programmed death ligand (PD-L) 1 on their surface, can express this molecule at moderate levels following cytokine exposure. Although moderate PD-L1 levels on healthy volunteers’ neutrophils were not inhibitory to T cells, our findings do not exclude a possible robust increase in neutrophil PD-L1 expression in pathological conditions with immunosuppressive functions. These results suggest a possible immunoregulatory role for human neutrophils in adaptive immunity. PMID:20123111

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

    PubMed Central

    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.

    1996-01-01

    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. Images Figure 1 Figure 2 Figure 3 Figure 4 PMID:8952534

  17. Are cytokines associated with neuropsychiatric syndromes in humans?

    PubMed

    Hickie, I; Lloyd, A

    1995-08-01

    Traditional aetiological models in neuropsychiatry have placed little emphasis on the abnormal behavioural responses (decreased psychomotor activity, anorexia, weight loss, decreased social exploration and sexual behaviour, impaired cognitive function and increased somnolence) that are common to both psychiatric syndromes, notably depression, and the illness behaviour of sick animals. In recent years, the possible role of cytokines, as mediators of not only the immunological and metabolic responses to infection and inflammation but also a co-ordinated behavioural response, has been described. Further, a range of possible mechanisms for these effects has been postulated, notably involving corticotropin releasing factor (CRF) and prostaglandins of the E series (PgE) with the central nervous system (CNS). Here we outline a series of human clinical conditions where neuropsychiatric syndromes co-occur with a host response to infection or inflammation. These may be characterized by cytokine production (e.g. acute, recurrent and chronic viral illness, systemic autoimmune diseases and chronic fatigue syndrome). Other clinical situations characterized by exposure to or in vivo production of cytokines (e.g. treatment of chronic infections and malignancies, progression and/or recurrence of malignancies) are also discussed. We postulate that the stereotyped behavioural repertoire observed is mediated by cytokine-dependent mechanisms within the CNS. Systematic studies of the behavioural responses of such patient groups are suggested, noting specifically correlations between the time course and severity of immune and neuroendocrine and behavioural responses and dose-response effects. PMID:8847162

  18. Cytokines and Mycobacterium leprae induce apoptosis in human Schwann cells.

    PubMed

    Oliveira, Rosane B; Sampaio, Elizabeth P; Aarestrup, Fernando; Teles, Rosane M B; Silva, Tatiana P; Oliveira, Ariane L; Antas, Paulo R Z; Sarno, Euzenir N

    2005-10-01

    The development of deformities during the course of leprosy disease is a major public health concern worldwide. It is possible that cytokine production and apoptosis of Schwann cells (SCs) directly affect nerve degeneration and regeneration leading to injury of the myelin sheath and axon. In the present study, the expression of TNFalpha, TGFbeta, and their receptors, in addition to cell death triggered by cytokines or whole Mycobacterium leprae were investigated in a human SC line. The results showed the presence of TNF-Rs and TGF-RII on the SC membrane and the shedding of TNF-Rs during the culture period. Evaluation of cell death was performed through TUNEL and flow cytometry techniques. TNFalpha/TGFbeta combination as well as M. leprae infection triggered an increase in the apoptosis rate in the cultured SC. Moreover, reverse transcriptase-polymerase chain reaction assay revealed that M. leprae upregulated the expression of such cytokines and their receptors on the SC line. Despite the detection of TNFalpha mRNA, no protein was found in the culture supernatants. The data indicate that induction of SC death after cell interaction with M. leprae may, in fact, be implicated in the pathogenesis of nerve damage, which can most likely be modulated by in vivo cytokine production. PMID:16215460

  19. Titanium surface hydrophilicity modulates the human macrophage inflammatory cytokine response.

    PubMed

    Alfarsi, Mohammed A; Hamlet, Stephen M; Ivanovski, Saso

    2014-01-01

    Increased titanium surface hydrophilicity has been shown to accelerate dental implant osseointegration. Macrophages are important in the early inflammatory response to surgical implant placement and influence the subsequent healing response. This study investigated the modulatory effect of a hydrophilic titanium surface on the inflammatory cytokine expression profile in a human macrophage cell line (THP-1). Genes for 84 cytokines, chemokines, and their receptors were analyzed following exposure to (1) polished (SMO), (2) micro-rough sand blasted, acid etched (SLA), and (3) hydrophilic-modified SLA (modSLA) titanium surfaces for 1 and 3 days. By day 3, the SLA surface elicited a pro-inflammatory response compared to the SMO surface with statistically significant up-regulation of 16 genes [Tumor necrosis factor (TNF) Interleukin (IL)-1β, Chemokine (C-C motif) ligand (CCL)-1, 2, 3, 4, 18, 19, and 20, Chemokine (C-X-C motif) ligand (CXCL)-1, 5, 8 and 12, Chemokine (C-C motif) receptor (CCR)-7, Lymphotoxin-beta (LTB), and Leukotriene B4 receptor (LTB4R)]. This effect was countered by the modSLA surface, which down-regulated the expression of 10 genes (TNF, IL-1α and β, CCL-1, 3, 19 and 20, CXCL-1 and 8, and IL-1 receptor type 1), while two were up-regulated (osteopontin and CCR5) compared to the SLA surface. These cytokine gene expression changes were confirmed by decreased levels of corresponding protein secretion in response to modSLA compared to SLA. These results show that a hydrophilic titanium surface can modulate human macrophage pro-inflammatory cytokine gene expression and protein secretion. An attenuated pro-inflammatory response may be an important molecular mechanism for faster and/or improved wound healing. PMID:23595995

  20. Synthesis Approaches to (-)-Cytoxazone, a Novel Cytokine Modulator, and Related Structures.

    PubMed

    Miranda, Izabel L; Lopes, Ítala K B; Diaz, Marisa A N; Diaz, Gaspar

    2016-01-01

    (-)-Cytoxazone, originally isolated from cultures of a Streptomyces species has an oxazolidin-2-one 4,5-disubstituted ring. It is known that this natural product presents a cytokine modulator effect through the signaling pathway of Th2 cells (type 2 cytokines), which are involved in the process of growth and differentiation of cells. From this, the interest in the development of research aimed at the total synthesis of this molecule and its analogs has remained high, which can be confirmed by the large number of publications on the topic, more than 30 to date. This review focuses on the various creative methods for the synthesis of (-)-cytoxazone and its congeners. The assessment of the preparation of this oxazolidinone and related structures serves as a treatise on the efforts made in the synthesis of this important class of compound from its first total synthesis in 1999. PMID:27608004

  1. P-body formation limits proinflammatory cytokine synthesis in endotoxin tolerant monocytes and murine septic macrophages

    PubMed Central

    McClure, Clara; Brudecki, Laura; Yao, Zhi Q.; McCall, Charles E.; Gazzar, Mohamed El

    2015-01-01

    An anti-inflammatory phenotype with pronounced immunosuppression develops during sepsis, during which time neutrophils and monocyte/macrophages limit their toll-like receptor 4 responses to bacterial lipopolysaccharide (LPS/endotoxin). We previously reported that during this endotoxin tolerant state, distinct signaling pathways differentially repress transcription and translation of proinflammatory cytokines such as TNFα and IL-6. Sustained endotoxin tolerance contributes to sepsis mortality. While transcription repression requires chromatin modifications, a translational repressor complex of Ago2 and RBM4, which bind the 3’ UTR of TNFα and IL-6 mRNA, limits protein synthesis. Here, we show that Dcp1 supports the assembly of Ago2 and RBM4 repressor complex into cytoplasmic p-bodies in endotoxin-tolerant THP-1 human monocytes following stimulation with LPS, resulting in translational repression and limiting protein synthesis. Importantly, this translocation process is reversed by Dcp1 knockdown, which restores TNFα and IL-6 protein levels. We also find this translational repression mechanism in primary macrophages of septic mice. Because p-body formation is a critical step in mRNA translation repression, we conclude that Dcp1 is a major component of the translational repression machinery of endotoxin tolerance and may contribute to sepsis outcome. PMID:25998849

  2. Human immunodeficiency virus type 1 infection of human macrophages modulates the cytokine response to Pneumocystis carinii.

    PubMed Central

    Kandil, O; Fishman, J A; Koziel, H; Pinkston, P; Rose, R M; Remold, H G

    1994-01-01

    The present studies examined production of the cytokines tumor necrosis factor alpha (TNF-alpha), interleukin-1 beta (IL-1 beta), and IL-6 by human monocyte-derived macrophages exposed to Pneumocystis carinii in vitro and the impact of concurrent macrophage infection with human immunodeficiency virus type 1 (HIV-1) on these cytokine responses. Macrophages were infected with the HIV-1 BaL monocytotropic strain for 10 to 14 days and then exposed to P. carinii. At various times following P. carinii treatment, culture supernatants were harvested to assess the cytokine profile. Addition of P. carinii to HIV-uninfected macrophages resulted in augmented production of IL-6, TNF-alpha, and IL-1 beta protein. By contrast, in HIV-infected macrophages exposed to P. carinii, only the release of IL-6 was increased compared with that for HIV-uninfected macrophages, while the levels of TNF-alpha and IL-1 beta decreased. This altered response was confirmed at the molecular level for TNF-alpha mRNA. Preventing physical contact between P. carinii and macrophages by a membrane filter inhibited all cytokine release. Substituting P. carinii with a preparation of P. carinii 95- to 115-kDa major membrane glycoprotein A yielded a response similar to that obtained by addition of intact P. carinii. These results suggest that HIV-1 infection of human macrophages modulates cytokine responses to P. carinii. Images PMID:8300221

  3. 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.

  4. Original inhibition method of excessive synthesis of pro-inflammatory cytokine of tumour necrosis factor α

    PubMed Central

    Zinchuk, AleXander; Holubovska, Olga; Shkurba, Andrij; Hrytsko, Roman; Vorozhbyt, Olga; Richniak, Mykhailo

    2015-01-01

    Influence on pro- and anti-inflammatory cytokines of an ill person is an urgent aspect of treatment of many diseases. For inhibition of synthesis of a high level of pro-inflammatory cytokines, medications which are recombinant monoclonal antibodies, especially to tumour necrosis factor α (TNF-α), are used. However, these methods of treatment require further improvement by elaborating new approaches with a wider spectrum of influence on the immune system. A completely new method of reduction in high activity of TN F-α with the method of intradermal autoleukocyte immunization is presented in the article. Investigation was performed in a group of patients with psoriasis (24) with a high level of TNF-α in the blood (over 30 pg/ml). Simultaneously such investigation was performed on patients with psoriasis (9) without TNF-α detected (0 pg/ml). As a result of immunization, a significant reduction in TNF-α occurred in all patients with its high level, in 16 (66.7%) from 24 patients – to 0-5 pg/ml. The level of reduction and duration of the achieved effect was of an individual character and requires further investigation. However, the achieved results prove the expediency of administration of this immunization method for patients requiring reduction of TNF-α synthesis. However, the content of TNF-α in blood serum could not be detected in most patients with a low level of cytokine (in 6 from 9) after immunization (as well as before immunization), but an increase in its level from 0 to 5-8 pg/ml was observed in 3 patients. On the basis of the conducted research, the authors suggest that the influence of immunization on cytokine synthesis depends on the condition of immune cells and correlation of pro- and anti-inflammatory cytokines in a patient's skin. PMID:26648779

  5. 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

  6. Cytokine factors present in dengue patient sera induces alterations of junctional proteins in human endothelial cells.

    PubMed

    Appanna, Ramapraba; Wang, Seok Mui; Ponnampalavanar, Sasheela A; Lum, Lucy Chai See; Sekaran, Shamala Devi

    2012-11-01

    Plasma leakage in severe dengue has been postulated to be associated with skewed cytokine immune responses. In this study, the association of cytokines with vascular permeability in dengue patients was investigated. Human serum samples collected from 48 persons (13 with dengue fever, 29 with dengue hemorrhagic fever, and 6 healthy) were subjected to cytokines analysis by using Luminex Multiplex Technology. Selected serum samples from patients with dengue hemorrhagic fever sera and recombinant human cytokines were then tested for roles on inducing vascular permeability by treatment of human umbilical vein endothelial cells. Confocal immunofluorescence staining indicated morphologic alteration of human umbilical vein endothelial cells treated with serum samples from patients with dengue hemorrhagic fever compared with serum samples from healthy persons. The findings suggest that cytokines produced during dengue hemorrhagic infections could induce alterations in the vascular endothelium, which may play a fundamental role in the pathophysiology of dengue. PMID:22987650

  7. 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

  8. 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

  9. TGN1412 Induces Lymphopenia and Human Cytokine Release in a Humanized Mouse Model

    PubMed Central

    Weißmüller, Sabrina; Kronhart, Stefanie; Kreuz, Dorothea; Schnierle, Barbara; Kalinke, Ulrich; Kirberg, Jörg; Hanschmann, Kay-Martin; Waibler, Zoe

    2016-01-01

    Therapeutic monoclonal antibodies (mAbs) such as the superagonistic, CD28-specific antibody TGN1412, or OKT3, an anti-CD3 mAb, can cause severe adverse events including cytokine release syndrome. A predictive model for mAb-mediated adverse effects, for which no previous knowledge on severe adverse events to be expected or on molecular mechanisms underlying is prerequisite, is not available yet. We used a humanized mouse model of human peripheral blood mononuclear cell-reconstituted NOD-RAG1-/-Aβ-/-HLADQ(tg+ or tg-)IL-2Rγc-/- mice to evaluate its predictive value for preclinical testing of mAbs. 2–6 hours after TGN1412 treatment, mice showed a loss of human CD45+ cells from the peripheral blood and loss of only human T cells after OKT3 injection, reminiscent of effects observed in mAb-treated humans. Moreover, upon OKT3 injection we detected selective CD3 downmodulation on T cells, a typical effect of OKT3. Importantly, we detected release of human cytokines in humanized mice upon both OKT3 and TGN1412 application. Finally, humanized mice showed severe signs of illness, a rapid drop of body temperature, and succumbed to antibody application 2–6 hours after administration. Hence, the humanized mouse model used here reproduces several effects and adverse events induced in humans upon application of the therapeutic mAbs OKT3 and TGN1412. PMID:26959227

  10. TGN1412 Induces Lymphopenia and Human Cytokine Release in a Humanized Mouse Model.

    PubMed

    Weißmüller, Sabrina; Kronhart, Stefanie; Kreuz, Dorothea; Schnierle, Barbara; Kalinke, Ulrich; Kirberg, Jörg; Hanschmann, Kay-Martin; Waibler, Zoe

    2016-01-01

    Therapeutic monoclonal antibodies (mAbs) such as the superagonistic, CD28-specific antibody TGN1412, or OKT3, an anti-CD3 mAb, can cause severe adverse events including cytokine release syndrome. A predictive model for mAb-mediated adverse effects, for which no previous knowledge on severe adverse events to be expected or on molecular mechanisms underlying is prerequisite, is not available yet. We used a humanized mouse model of human peripheral blood mononuclear cell-reconstituted NOD-RAG1-/-Aβ-/-HLADQ(tg+ or tg-)IL-2Rγc-/- mice to evaluate its predictive value for preclinical testing of mAbs. 2-6 hours after TGN1412 treatment, mice showed a loss of human CD45+ cells from the peripheral blood and loss of only human T cells after OKT3 injection, reminiscent of effects observed in mAb-treated humans. Moreover, upon OKT3 injection we detected selective CD3 downmodulation on T cells, a typical effect of OKT3. Importantly, we detected release of human cytokines in humanized mice upon both OKT3 and TGN1412 application. Finally, humanized mice showed severe signs of illness, a rapid drop of body temperature, and succumbed to antibody application 2-6 hours after administration. Hence, the humanized mouse model used here reproduces several effects and adverse events induced in humans upon application of the therapeutic mAbs OKT3 and TGN1412. PMID:26959227

  11. Enhanced cytokine production and collagen synthesis of gingival fibroblasts from patients with denture fibromatosis.

    PubMed

    Nakao, K; Yoneda, K; Osaki, T

    1995-04-01

    The mechanisms of denture-induced gingival hypertrophy remain to be explored. Since fibroblast proliferation and bone resorption characterize this disorder, the possible involvement of cytokines was investigated. Gingival fibroblasts were obtained from six patients with denture fibromatosis (Den-Fb) and six healthy persons (Nor-Fb). Cells were compared for proliferation, collagen synthesis, and cytokine production. Incorporation of [3H]thymidine (TdR) was increased in 3 Den-Fb and 3 Nor-Fb lines in the presence of interleukin-1-beta (IL-1 beta) (10 U/mL) and tumor necrosis factor-alpha (TNF-alpha) (from 10 to 100 U/mL). Proline incorporation in Den-Fb was higher than that in Nor-Fb, and the mean collagen synthesis level in Den-Fb was significantly higher than that in Nor-Fb. Although there was no difference between the up-regulation of protein synthesis in Den-Fb and Nor-Fb induced by IL-1 beta or TNF-alpha, the receptors for these cytokines were expressed at higher levels in cell lines which exhibited higher protein synthesis. Between Nor-Fb and Den-Fb, there was no difference in the generation of granulocyte-macrophage colony-stimulating factor (GM-CSF) or interleukin-6 (IL-6). However, most Den-Fb produced more GM-CSF and IL-6 in the presence of TNF-alpha. Enhancement of IL-6 generation by GM-CSF was also more prominent in Den-Fb. GM-CSF and IL-6 were synergistically generated after co-culture of the fibroblasts with gingival keratinocytes. GM-CSF and IL-6 generation of Den-Fb was markedly enhanced by co-culture of Den-Fb with peripheral blood mononuclear cells (PBMC), especially PBMC from patients.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:7782537

  12. Mutational Separation of Aminoacylation and Cytokine Activities of Human Tyrosyl-tRNA Synthetase

    PubMed Central

    Kapoor, Mili; Otero, Francella J.; Slike, Bonnie M.; Ewalt, Karla L.; Yang, Xiang-Lei

    2009-01-01

    SUMMARY Aminoacyl-tRNA synthetases are known for catalysis of aminoacylation. Significantly, some mammalian synthetases developed cytokine functions possibly linked to disease-causing mutations in tRNA synthetases. Not understood is how epitopes for cytokine signaling were introduced into catalytic scaffolds without disturbing aminoacylation. Here we investigate human tyrosyl-tRNA synthetase, where a catalytic-domain surface helix—next to the active site—was recruited for IL-8-like cytokine signaling. Taking advantage of our high-resolution structure, the reciprocal impact of rational mutations designed to disrupt aminoacylation or cytokine signaling was investigated with multiple assays. The collective analysis demonstrated a protective fine–structure separation of aminoacylation from cytokine activities within the conserved catalytic domain. As a consequence, disease-causing mutations affecting cell signaling can arise without disturbing aminoacylation. These results with TyrRS also predict the previously unknown binding conformation of IL-8-like CXC cytokines. PMID:19477417

  13. Mutational separation of aminoacylation and cytokine activities of human tyrosyl-tRNA synthetase.

    PubMed

    Kapoor, Mili; Otero, Francella J; Slike, Bonnie M; Ewalt, Karla L; Yang, Xiang-Lei

    2009-05-29

    Aminoacyl tRNA synthetases are known for catalysis of aminoacylation. Significantly, some mammalian synthetases developed cytokine functions possibly linked to disease-causing mutations in tRNA synthetases. Not understood is how epitopes for cytokine signaling were introduced into catalytic scaffolds without disturbing aminoacylation. Here we investigate human tyrosyl-tRNA synthetase, where a catalytic-domain surface helix, next to the active site, was recruited for interleukin-8-like cytokine signaling. Taking advantage of our high resolution structure, the reciprocal impact of rational mutations designed to disrupt aminoacylation or cytokine signaling was investigated with multiple assays. The collective analysis demonstrated a protective fine-structure separation of aminoacylation from cytokine activities within the conserved catalytic domain. As a consequence, disease-causing mutations affecting cell signaling can arise without disturbing aminoacylation. These results with TyrRS also predict the previously unknown binding conformation of interleukin-8-like CXC cytokines. PMID:19477417

  14. Food contaminant zearalenone and its metabolites affect cytokine synthesis and intestinal epithelial integrity of porcine cells.

    PubMed

    Marin, Daniela E; Motiu, Monica; Taranu, Ionelia

    2015-06-01

    The intestinal epithelium is the first barrier against food contaminants. Zearalenone (ZEN) is an estrogenic mycotoxin that was identified as a common contaminant of cereal grains and food and feedstuffs. In the present study, we have investigated the in vitro effects of ZEN and some of its metabolites (α-ZOL, β-ZOL) in concentrations of 10-100 µM on a swine epithelial cell line: Intestinal porcine epithelial cells (IPEC-1). We demonstrated that both ZEN metabolites were more toxic for IPEC cells as resulted from the XTT test, while for doses lower than 10 µM, only β-ZOL showed a more pronounced cytotoxicity versus epithelial cells as resulted from neutral red assay. ZEN has no effect on TER values, while α-ZOL significantly decreased the TER values, starting with day 4 of treatment. β-ZOL had a dual effect, firstly it induced a significant increase of TER, and then, starting on day 6, it induced a dramatic decrease of TER values as compared with on day 0. Concerning the cytokine synthesis, our results showed that ZEN has a tendency to increase the synthesis of IL-8 and IL-10. By contrast, α- and β-ZOL decreased the expression of both IL-8 and IL-10, in a dose dependent manner. In conclusion, our results showed that ZEN and its metabolites differently affected porcine intestinal cell viability, transepithelial resistance and cytokine synthesis with important implication for gut health. PMID:26035492

  15. Food Contaminant Zearalenone and Its Metabolites Affect Cytokine Synthesis and Intestinal Epithelial Integrity of Porcine Cells

    PubMed Central

    Marin, Daniela E.; Motiu, Monica; Taranu, Ionelia

    2015-01-01

    The intestinal epithelium is the first barrier against food contaminants. Zearalenone (ZEN) is an estrogenic mycotoxin that was identified as a common contaminant of cereal grains and food and feedstuffs. In the present study, we have investigated the in vitro effects of ZEN and some of its metabolites (α-ZOL, β-ZOL) in concentrations of 10–100 µM on a swine epithelial cell line: Intestinal porcine epithelial cells (IPEC-1). We demonstrated that both ZEN metabolites were more toxic for IPEC cells as resulted from the XTT test, while for doses lower than 10 µM, only β-ZOL showed a more pronounced cytotoxicity versus epithelial cells as resulted from neutral red assay. ZEN has no effect on TER values, while α-ZOL significantly decreased the TER values, starting with day 4 of treatment. β-ZOL had a dual effect, firstly it induced a significant increase of TER, and then, starting on day 6, it induced a dramatic decrease of TER values as compared with on day 0. Concerning the cytokine synthesis, our results showed that ZEN has a tendency to increase the synthesis of IL-8 and IL-10. By contrast, α- and β-ZOL decreased the expression of both IL-8 and IL-10, in a dose dependent manner. In conclusion, our results showed that ZEN and its metabolites differently affected porcine intestinal cell viability, transepithelial resistance and cytokine synthesis with important implication for gut health. PMID:26035492

  16. Direct and indirect effects of retinoic acid on human Th2 cytokine and chemokine expression by human T lymphocytes

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Vitamin A (VA) deficiency induces a type 1 cytokine response and exogenously provided retinoids can induce a type 2 cytokine response both in vitro and in vivo. The precise mechanism(s) involved in this phenotypic switch are inconsistent and have been poorly characterized in humans. In an effort t...

  17. 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. PMID:26948901

  18. Systemic Inflammation Affects Human Osteocyte-Specific Protein and Cytokine Expression.

    PubMed

    Pathak, Janak L; Bakker, Astrid D; Luyten, Frank P; Verschueren, Patrick; Lems, Willem F; Klein-Nulend, Jenneke; Bravenboer, Nathalie

    2016-06-01

    Bone remodeling can be disturbed in active rheumatoid arthritis (RA), possibly as a result of elevated levels of circulating inflammatory cytokines. Osteocyte-specific proteins and cytokines play a vital role in bone remodeling by orchestrating bone formation and/or bone resorption. Therefore, we aimed to investigate the effect of RA-serum or inflammatory cytokines on expression of human osteocyte-specific proteins and cytokines. Human trabecular bone chips were cultured with RA-serum or inflammatory cytokines for 7-days. Live-dead staining was performed to assess cell viability. Gene expression of osteocyte-specific proteins and cytokines was analyzed by qPCR. Immuno-staining was performed for osteocyte-specific markers. Approximately 60 % of the osteocytes on the bone chips were alive at day-7. Cells in or on the bone chips did express the gene for osteocyte markers SOST, FGF23, DMP1, and MEPE, and the cytokines IL-1β, IL-6, and TNFα at day 0 and 7. Active RA-serum treatment enhanced IL-1β, TNFα, SOST, and DKK1 gene expression. IL-1β treatment enhanced IL-1β, TNFα, IL-6, IL-8, FGF23, and SOST gene expression. TNFα treatment enhanced IL-1β, TNFα, IL-6, IL-8, and FGF23 gene expression. IL-8 treatment enhanced TNFα, IL-8, and FGF23 gene expression. A combination of IL-1β, IL-6, and TNFα treatment synergistically upregulated IL-1β, IL-6, and IL-8 gene expression, as well as enhanced TNFα, OPG, SOST, and FGF23, and inhibited DKK1 gene expression. In conclusion, gene expression of human osteocyte-specific proteins and cytokines was affected by RA-serum, and exogenous recombinant cytokines treatment suggesting that osteocytes could provide a new target to prevent systemic inflammation-induced bone loss in RA. PMID:26887974

  19. Control of Cytokine Production by Human Fc Gamma Receptors: Implications for Pathogen Defense and Autoimmunity

    PubMed Central

    Vogelpoel, Lisa T. C.; Baeten, Dominique L. P.; de Jong, Esther C.; den Dunnen, Jeroen

    2015-01-01

    Control of cytokine production by immune cells is pivotal for counteracting infections via orchestration of local and systemic inflammation. Although their contribution has long been underexposed, it has recently become clear that human Fc gamma receptors (FcγRs), which are receptors for the Fc region of immunoglobulin G (IgG) antibodies, play a critical role in this process by controlling tissue- and pathogen-specific cytokine production. Whereas individual stimulation of FcγRs does not evoke cytokine production, FcγRs cell-type specifically interact with various other receptors for selective amplification or inhibition of particular cytokines, thereby tailoring cytokine responses to the immunological context. The physiological function of FcγR-mediated control of cytokine production is to counteract infections with various classes of pathogens. Upon IgG opsonization, pathogens are simultaneously recognized by FcγRs as well as by various pathogen-sensing receptors, leading to the induction of pathogen class-specific immune responses. However, when erroneously activated, the same mechanism also contributes to the development of autoimmune diseases such as rheumatoid arthritis and systemic lupus erythematosus. In this review, we discuss control of cytokine production as a novel function of FcγRs in human innate immune cells in the context of homeostasis, infection, and autoimmunity and address the possibilities for future therapeutic exploitation. PMID:25759693

  20. 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.

  1. Interleukin-6: a multifunctional targetable cytokine in human prostate cancer.

    PubMed

    Culig, Zoran; Puhr, Martin

    2012-09-01

    Several cytokines are involved in regulation of cellular events in prostate cancer. Interleukin-6 (IL-6) was frequently investigated in prostate cancer models because of its increased expression in cancer tissue at early stages of the disease. In patients with metastatic prostate cancer, it is well-known that IL-6 levels increase in serum. High levels of IL-6 were measured in the supernatants of cells which do not respond to androgenic stimulation. IL-6 expression in prostate cancer increases due to enhanced expression of transforming growth factor-beta, and members of the activating protein-1 complex, and loss of the retinoblastoma tumour suppressor. IL-6 activation of androgen receptor (AR) may contribute to progression of a subgroup of prostate cancers. Results obtained with two prostate cancer cell lines, LNCaP and MDA PCa 2b, indicate that IL-6 activation of AR may cause either stimulatory or inhibitory responses on proliferation. Interestingly, prolonged treatment with IL-6 led to establishment of an IL-6 autocrine loop, suppressed signal transducer and activator of transcription (STAT)3 activation, and increased mitogen-activated protein kinase phosphorylation. In several cell lines IL-6 acts as a survival molecule through activation of the signalling pathway of phosphotidylinositol 3-kinase. Expression of suppressors of cytokine signalling (SOCS) has been studied in prostate cancer. SOCS-3 prevents phosphorylation of STAT3 and is an important anti-apoptotic factor in AR-negative prostate cancer cells. Experimental therapy against IL-6 in prostate cancer is based on the use of the monoclonal antibody siltuximab which may be used for personalised therapy coming in the future. PMID:21664423

  2. 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

  3. Heat stress and/or endotoxin effects on cytokine expression by human whole blood.

    PubMed

    DuBose, David A; Balcius, James; Morehouse, David

    2002-03-01

    Immune system cytokines induce vascular shock. Tumor necrosis factor-alpha (TNF-alpha), interleukin 1beta (IL-1beta), and bacterial endotoxin (E) circulate in human heatstroke to suggest that E release from a heat-damaged gut may stimulate cytokines that contribute to hypovolemia. However, immune activation by heat-induced tissue necrosis might stimulate cytokine generation in the absence of E. To evaluate this potential and heat stress effects on the anti-inflammatory cytokines, IL-1 receptor antagonist (IL-1ra) and IL-1 soluble receptor II (IL-1srII), a human whole blood (HWB) model was employed in which the presence or absence of E could be controlled. Using thermoelectric technology to regulate the HWB heat exposures, the temperature modulations of lethal heatstroke were precisely replicated (maximum temperature = 42.4 degrees C +/- 0.04 degrees C; thermal area = 52.3 degrees C +/- 1.5 degrees C per min). Cytokine and mRNA measurements employed enzyme-linked immunosorbant-based assay systems. Significant elevations in TNF-alpha, IL-1beta, interleukin 6 (IL-6), and IL-1ra resulted when HWB was exposed to E concentrations (10 ng/ml) reported to circulate in heatstroke. While E-stimulated IL-1ra was significantly decreased by the presence of prior heat stress (PPHS), E-stimulated IL-1beta, TNF-alpha, and IL-6 were not significantly altered by PPHS, but tended to be elevated. IL-1srII expression was unchanged by PPHS and/or E. PPHS in the absence of E did not induce cytokine responses, nor were there elevations in TNF-alpha or IL-1beta mRNA. Thus, some factor normally absent under in vitro conditions, like endotoxin, was required to provoke HWB cytokine expressions and the heat stress and E conditions that characterize heatstroke affected HWB cytokine metabolism to favor a proinflammatory environment. PMID:11900341

  4. Suppressors of cytokine signalling-3 and -1 in human carcinogenesis.

    PubMed

    Culig, Zoran

    2013-01-01

    The role of suppressors of cytokine signaling (SOCS)-3 and -1 has been investigated in various cancers. These proteins have been identified as endogenous controllers of activation of Janus kinase/signal transducers and activators of transcription factors pathway factors under physiological conditions and in disease. SOCS-3 expression is lost in several cancers due to epigenetic mechanisms, mostly promoter methylation. In liver, lung, and squamous head and neck cancer, and several hematological malignancies SOCS-3 acts as a classic tumor suppressor. In prostate cancer, SOCS-3 effects are cell type-dependent. It prevents apoptosis in androgen receptor-negative cells. However, in androgen-sensitive cells, it could act as a negative feedback factor for androgenic regulation. Melanoma cells which overexpress SOCS-3 confer a growth advantage. SOCS-1 is in most cancers a tumor suppressor which may inhibit expression of cyclin-dependent kinases and cyclins. In general, the mechanisms responsible for the different effects of SOCS in cancer cell lines have to be further investigated. The results discussed in the present review may have an impact on personalized approaches in cancer medicine. PMID:23277051

  5. 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.

  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. 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

  8. Response to stimulation with recombinant cytokines and synthesis of cytokines by murine intestinal macrophages infected with the Mycobacterium avium complex.

    PubMed Central

    Hsu, N; Young, L S; Bermudez, L E

    1995-01-01

    Current evidence suggests that the gut is the chief portal of entry for organisms of the Mycobacterium avium complex (MAC) in AIDS patients. Bacterial invasion of intestinal mucosa presumably occurs through epithelial cells, and M cells in the Peyer's patches, where the bacteria have contact with immunocompetent cells such as macrophages and T and B lymphocytes. As mucosal macrophages are probably the first line of defense against MAC, we examined their ability to inhibit intracellular growth of MAC when properly stimulated. Mouse intestinal macrophages were purified, infected with MAC 101, serovar 1, and MAC 86-2686, serovar 16, and subsequently stimulated with recombinant tumor necrosis factor alpha (TNF-alpha), gamma interferon (IFN-gamma), granulocyte-macrophage colony-stimulating factor (GM-CSF), or macrophage colony-stimulating factor (M-CSF). Viable intracellular bacteria were quantitated at 24 h after infection and again after 4 days of infection. Stimulation with TNF-alpha, IFN-gamma, and GM-CSF, but not M-CSF, was associated with mycobacteriostatic and/or mycobactericidal activity in macrophages. Treatment with 10(3) U of TNF-alpha, GM-CSF, and IFN-gamma per ml at 24 h prior to infection with MAC resulted in a significant enhancement in killing of MAC at 4 days after infection, compared with that observed for macrophages exposed to cytokines after infection. When stimulated with lipopolysaccharide or live MAC, intestinal macrophages had produced significantly less TNF-alpha and transforming growth factor beta than had splenic and peritoneal macrophages, although the levels of production of interleukin 6 and interleukin 10 among the three populations of cells were similar. Intestinal macrophages can be stimulated with cytokines to inhibit the intracellular growth of MAC, but they have differentiated abilities to produce cytokines which can modulate the anti-MAC immune response. PMID:7822018

  9. 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...

  10. Dharmendra antigen but not integral M. leprae is an efficient inducer of immunostimulant cytokine production by human monocytes, and M. leprae lipids inhibit the cytokine production.

    PubMed

    Nakamura, C; Fukutomi, Y; Kashiwabara, Y; Oomoto, Y; Kojima, M; Hayashi, H; Onozaki, K

    1997-03-01

    Killed integral Mycobacterium leprae, Mitsuda antigen, and chloroform-treated M. leprae, Dharmendra antigen (Dh-Ag), have been used for the classification of leprosy patients based on cell-mediated immunity. Heat-killed M. leprae also were used as a component of the Convit vaccine. Human blood monocytes were stimulated with M. leprae or Dh-Ag and their cytokine-inducing ability was compared. Monocytes were cultured in the presence of fresh human serum because of the efficiency of cytokine induction and the phagocytosis of M. leprae have been shown to be optimal in the presence of fresh serum. M. leprae and Dh-Ag were equally phagocytosed by monocytes. Dh-Ag was more potent than M. leprae in the induction of immunostimulatory/proinflammatory cytokines, interleukin-1 (IL-1), IL-6 and tumor necrosis factor (TNF). In contrast, a comparable level of IL-1ra, an immunosuppressive cytokine, was induced by M. leprae and Dh-Ag. The lipids extracted from M. leprae induced none of these cytokines by monocytes. Nevertheless, when monocytes were pretreated with the lipids followed by stimulation with Dh-Ag, productions of IL-1, IL-6 and TNF were all inhibited in a dose-dependent manner. However, the lipids did not inhibit the cytokine production induced by other stimuli including BCG and lipopolysaccharide. Moreover the lipids did not affect the production of IL-1ra. These results suggest that the lipids from M. leprae are responsible for the poor cytokine-inducing ability of M. leprae, thus favoring their infection. These results also suggest that Dh-Ag rather than integral M. leprae may be useful as a vaccine candidate because Dh-Ag is able to induce a large amount of cytokines from monocytes. PMID:9207755

  11. Influenza Vaccination Generates Cytokine-Induced Memory-like NK Cells: Impact of Human Cytomegalovirus Infection.

    PubMed

    Goodier, Martin R; Rodriguez-Galan, Ana; Lusa, Chiara; Nielsen, Carolyn M; Darboe, Alansana; Moldoveanu, Ana L; White, Matthew J; Behrens, Ron; Riley, Eleanor M

    2016-07-01

    Human NK cells are activated by cytokines, immune complexes, and signals transduced via activating ligands on other host cells. After vaccination, or during secondary infection, adaptive immune responses can enhance both cytokine-driven and Ab-dependent NK cell responses. However, induction of NK cells for enhanced function after in vitro exposure to innate inflammatory cytokines has also been reported and may synergize with adaptive signals to potentiate NK cell activity during infection or vaccination. To test this hypothesis, we examined the effect of seasonal influenza vaccination on NK cell function and phenotype in 52 previously unvaccinated individuals. Enhanced, IL-2-dependent, NK cell IFN-γ responses to Influenza A/California/7/2009 virus were detected up to 4 wk postvaccination and higher in human CMV (HCMV)-seronegative (HCMV(-)) individuals than in HCMV-seropositive (HCMV(+)) individuals. By comparison, robust NK cell degranulation responses were observed both before and after vaccination, due to high titers of naturally occurring anti-influenza Abs in human plasma, and did not differ between HCMV(+) and HCMV(-) subjects. In addition to these IL-2-dependent and Ab-dependent responses, NK cell responses to innate cytokines were also enhanced after influenza vaccination; this was associated with proliferation of CD57(-) NK cells and was most evident in HCMV(+) subjects. Similar enhancement of cytokine responsiveness was observed when NK cells were cocultured in vitro with Influenza A/California/7/2009 virus, and this was at least partially dependent upon IFN-αβR2. In summary, our data indicate that attenuated or live viral vaccines promote cytokine-induced memory-like NK cells and that this process is influenced by HCMV infection. PMID:27233958

  12. Influenza Vaccination Generates Cytokine-Induced Memory-like NK Cells: Impact of Human Cytomegalovirus Infection

    PubMed Central

    Goodier, Martin R.; Rodriguez-Galan, Ana; Lusa, Chiara; Nielsen, Carolyn M.; Darboe, Alansana; Moldoveanu, Ana L.; White, Matthew J.; Behrens, Ron

    2016-01-01

    Human NK cells are activated by cytokines, immune complexes, and signals transduced via activating ligands on other host cells. After vaccination, or during secondary infection, adaptive immune responses can enhance both cytokine-driven and Ab-dependent NK cell responses. However, induction of NK cells for enhanced function after in vitro exposure to innate inflammatory cytokines has also been reported and may synergize with adaptive signals to potentiate NK cell activity during infection or vaccination. To test this hypothesis, we examined the effect of seasonal influenza vaccination on NK cell function and phenotype in 52 previously unvaccinated individuals. Enhanced, IL-2–dependent, NK cell IFN-γ responses to Influenza A/California/7/2009 virus were detected up to 4 wk postvaccination and higher in human CMV (HCMV)-seronegative (HCMV−) individuals than in HCMV-seropositive (HCMV+) individuals. By comparison, robust NK cell degranulation responses were observed both before and after vaccination, due to high titers of naturally occurring anti-influenza Abs in human plasma, and did not differ between HCMV+ and HCMV− subjects. In addition to these IL-2–dependent and Ab-dependent responses, NK cell responses to innate cytokines were also enhanced after influenza vaccination; this was associated with proliferation of CD57− NK cells and was most evident in HCMV+ subjects. Similar enhancement of cytokine responsiveness was observed when NK cells were cocultured in vitro with Influenza A/California/7/2009 virus, and this was at least partially dependent upon IFN-αβR2. In summary, our data indicate that attenuated or live viral vaccines promote cytokine-induced memory-like NK cells and that this process is influenced by HCMV infection. PMID:27233958

  13. Human asthma phenotypes: from the clinic, to cytokines, and back again

    PubMed Central

    Bhakta, Nirav R.; Woodruff, Prescott G.

    2012-01-01

    Summary A large body of experimental evidence supports the hypothesis that T-helper 2 (Th2) cytokines orchestrate allergic airway inflammation in animal models. However, human asthma is heterogeneous with respect to clinical features, cellular sources of inflammation, and response to common therapies. This disease heterogeneity has been investigated using sputum cytology as well as unbiased clustering approaches using cellular and clinical data. Important differences in cytokine-driven inflammation may underlie this heterogeneity, and studies in human subjects with asthma have begun to elucidate these molecular differences. This molecular heterogeneity may be assessed by existing biomarkers (induced sputum evaluation or exhaled nitric oxide testing) or may require novel biomarkers. Effective testing and application of emerging therapies that target Th2 cytokines will depend on accurate and easily obtained biomarkers of this molecular heterogeneity in asthma. Furthermore, whether other non-Th2 cytokine pathways underlie airway inflammation in specific subsets of patients with asthma is an unresolved question and an important goal of future research using both mouse models and human studies. PMID:21682748

  14. Proinflammatory cytokines and HIV-1 synergistically enhance CXCL10 expression in human astrocytes.

    PubMed

    Williams, Rachel; Dhillon, Navneet K; Hegde, Sonia T; Yao, Honghong; Peng, Fuwang; Callen, Shannon; Chebloune, Yahia; Davis, Randall L; Buch, Shilpa J

    2009-05-01

    HIV encephalitis (HIVE), the pathologic correlate of HIV-associated dementia (HAD) is characterized by astrogliosis, cytokine/chemokine dysregulation, and neuronal degeneration. Increasing evidence suggests that inflammation is actively involved in the pathogenesis of HAD. In fact, the severity of HAD/HIVE correlates more closely with the presence of activated glial cells than with the presence and amount of HIV-infected cells in the brain. Astrocytes, the most numerous cell type within the brain, provide an important reservoir for the generation of inflammatory mediators, including interferon-gamma inducible peptide-10 (CXCL10), a neurotoxin and a chemoattractant, implicated in the pathophysiology of HAD. Additionally, the proinflammatory cytokines, IFN-gamma and TNF-alpha, are also markedly increased in CNS tissues during HIV-1 infection. In this study, we hypothesized that the interplay of host cytokines and HIV-1 could lead to enhanced expression of the toxic chemokine, CXCL10. Our findings demonstrate a synergistic induction of CXCL10 mRNA and protein in human astrocytes exposed to HIV-1 and the proinflammatory cytokines. Signaling molecules, including JAK, STATs, MAPK (via activation of Erk1/2, AKT, and p38), and NF-kappaB were identified as instrumental in the synergistic induction of CXCL10. Understanding the mechanisms involved in HIV-1 and cytokine-mediated up-regulation of CXCL10 could aid in the development of therapeutic modalities for HAD. PMID:18985732

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

    PubMed

    Burns, Ariel; Ciborowski, Pawel

    2016-02-01

    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

  16. Interleukin-18 Increases TLR4 and Mannose Receptor Expression and Modulates Cytokine Production in Human Monocytes

    PubMed Central

    Dias-Melicio, Luciane Alarcão; Fernandes, Reginaldo Keller; Rodrigues, Daniela Ramos; Golim, Marjorie Assis; Soares, Angela Maria Victoriano Campos

    2015-01-01

    Interleukin-18 is a proinflammatory cytokine belonging to the interleukin-1 family of cytokines. This cytokine exerts many unique biological and immunological effects. To explore the role of IL-18 in inflammatory innate immune responses, we investigated its impact on expression of two toll-like receptors (TLR2 and TLR4) and mannose receptor (MR) by human peripheral blood monocytes and its effect on TNF-α, IL-12, IL-15, and IL-10 production. Monocytes from healthy donors were stimulated or not with IL-18 for 18 h, and then the TLR2, TLR4, and MR expression and intracellular TNF-α, IL-12, and IL-10 production were assessed by flow cytometry and the levels of TNF-α, IL-12, IL-15, and IL-10 in culture supernatants were measured by ELISA. IL-18 treatment was able to increase TLR4 and MR expression by monocytes. The production of TNF-α and IL-10 was also increased by cytokine treatment. However, IL-18 was unable to induce neither IL-12 nor IL-15 production by these cells. Taken together, these results show an important role of IL-18 on the early phase of inflammatory response by promoting the expression of some pattern recognition receptors (PRRs) that are important during the microbe recognition phase and by inducing some important cytokines such as TNF-α and IL-10. PMID:25873755

  17. Gene-Specific Repression of Proinflammatory Cytokines in Stimulated Human Macrophages by Nuclear IκBα

    PubMed Central

    Ghosh, Chandra C.; Ramaswami, Sitharam; Juvekar, Ashish; Vu, Hai-Yen; Galdieri, Luciano; Davidson, Dennis; Vancurova, Ivana

    2011-01-01

    We have previously shown that increased nuclear accumulation of IκBα inhibits NF-κB activity and induces apoptosis in human leukocytes. In this study, we wanted to explore the possibility that the nucleocytoplasmic distribution of IκBα can be used as a therapeutic target for the regulation of NF-κB–dependent cytokine synthesis. Treatment of LPS-stimulated human U937 macrophages with an inhibitor of chromosome region maintenance 1-dependent nuclear export, leptomycin B, resulted in the increased nuclear accumulation of IκBα and inhibition of NF-κB DNA binding activity, caused by the nuclear IκBα-p65 NF-κB interaction. Surprisingly, however, whereas mRNA expression and cellular release of TNF-α, the β form of pro-IL-1 (IL-1β), and IL-6 were inhibited by the leptomycin B-induced nuclear IκBα, IL-8 mRNA expression and cellular release were not significantly affected. Analysis of in vivo recruitment of p65 NF-κB to NF-κB–regulated promoters by chromatin immunoprecipitation in U937 cells and human PBMCs indicated that although the p65 recruitment to TNF-α, IL-1β, and IL-6 promoters was inhibited by the nuclear IκBα, p65 recruitment to IL-8 promoter was not repressed. Chromatin immunoprecipitation analyses using IκBα and S536 phosphospecific p65 NF-κB Abs demonstrated that although the newly synthesized IκBα induced by postinduction repression is recruited to TNF-α, IL-1β, and IL-6 promoters but not to the IL-8 promoter, S536-phosphorylated p65 is recruited to IL-8 promoter, but not to TNF-α, IL-1β, or IL-6 promoters. Together, these data indicate that the inhibition of NF-κB–dependent transcription by nuclear IκBα in LPS-stimulated macrophages is gene specific and depends on the S536 phosphorylation status of the recruited p65 NF-κB. PMID:20696864

  18. 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

  19. Kinetic study of cytokines production by human peripheral blood mononuclear cells in response to Brucella DNA.

    PubMed

    Lashkarbolouki, Taghi; Ardestani, Sussan K; Kariminia, Amina; Ziaee, Abed-Ali; Torkabadi, Ebrahim; Ebrahimi, Mohammad

    2008-01-01

    In spite of reports on cytokines induction by the Brucella DNA in murine model, there is no comparison between pathogenic and appropriate vaccine strains in human. We investigated the cytokines profile of human peripheral blood mononuclear cells (PBMCs) induced by DNA extracted from pathogenic isolates of Brucella melitensis and B. abortus as well as Rev1 and S19; the appropriate vaccine strains. It was observed that despite differential induction of Interleukin(IL)-12 and IL-10 production, identical IL-12/IL-10 concentration ratio was obtained by all Brucella strains DNAs that was 2 after 24 h and 4 after 5 days of incubation. In addition, IL-2 and Interferon(IFN)-gamma production were profoundly increased compared to the medium at day 3 and 5 respectively but IFN-alpha was not induced. Therefore, Brucella strains DNAs are Th1 inducing component with similar pattern in human PBMCs. PMID:17008080

  20. Shikonin Inhibits Inflammatory Cytokine Production in Human Periodontal Ligament Cells.

    PubMed

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

    2016-06-01

    Shikonin, which is derived from Lithospermum erythrorhizon, a herb used in traditional medicine, has long been considered to be a useful treatment for various diseases in traditional oriental medicine. Shikonin has recently been reported to have several pharmacological properties, e.g., it has anti-microbial, anti-tumor, and anti-inflammatory effects. The aim of this study was to examine whether shikonin is able to influence the production of interleukin (IL)-6, IL-8, and/or chemokine C-C motif ligand (CCL)20, which contribute to the pathogenesis of periodontal disease, in human periodontal ligament cells (HPDLC). The production levels of IL-6, IL-8, and CCL20 in HPDLC were determined using an ELISA. Western blot analysis was used to detect nuclear factor kappa B (NF-κB) pathway activation in HPDLC. Shikonin prevented IL-1β- or tumor necrosis factor (TNF)-α-mediated IL-6, IL-8, and CCL20 production in HPDLC. Moreover, we found that shikonin suppressed the phosphorylation and degradation of inhibitor of kappa B-alpha (IκB-α) in IL-1β- or TNF-α-stimulated HPDLC. These findings suggest that shikonin could have direct beneficial effects against periodontal disease by reducing IL-6, IL-8, and CCL20 production in periodontal lesions. PMID:27072015

  1. Heterogeneity of cytokine and growth factor gene expression in human melanoma cells with different metastatic potentials.

    PubMed

    Singh, R K; Gutman, M; Radinsky, R

    1995-01-01

    The purpose of this study was to determine the mRNA expression level of multiple cytokine and growth factor genes in human malignant melanoma. Melanoma cells were isolated from several surgical specimens, adapted to growth in culture, characterized for their ability to produce experimental metastases in nude mice, and assessed for cytokine and growth factor steady-state gene expression. Highly metastatic in vivo- and in vitro-derived variants isolated from a single melanoma, A375, were also analyzed. Northern blot analyses revealed that all melanomas analyzed constitutively expressed steady-state mRNA transcripts for the growth and angiogenic factors, basic fibroblast growth factor (bFGF), and transforming growth factor alpha (TGF-alpha), which correlated with metastatic propensity. Only one highly metastatic melanoma, TXM-1, originally isolated from a lymph node metastasis, expressed mRNA transcripts specific for monocyte chemotactic and activating factor (MCAF) and granulocyte-macrophage colony-stimulating factor (GM-CSF). Similarly, of the nine melanomas examined, only TXM-1 expressed interleukin (IL)-1 alpha, IL-1 beta, and IL-6, important immunomodulatory cytokines. These data demonstrate the differential and heterogeneous expression of cytokine and growth factor genes in human malignant melanoma. PMID:7648437

  2. Epigenetic regulation of inflammatory cytokines and associated genes in human malignancies.

    PubMed

    Yasmin, Rehana; Siraj, Sami; Hassan, Amjad; Khan, Abdul Rehman; Abbasi, Rashda; Ahmad, Nafees

    2015-01-01

    Inflammation is a multifaceted defense response of immune system against infection. Chronic inflammation has been implicated as an imminent threat for major human malignancies and is directly linked to various steps involved in tumorigenesis. Inflammatory cytokines, interleukins, interferons, transforming growth factors, chemokines, and adhesion molecules have been associated with chronic inflammation. Numerous cytokines are reported to be aberrantly regulated by different epigenetic mechanisms like DNA methylation and histone modifications in tumor tissues, contributing to pathogenesis of tumor in multiple ways. Some of these cytokines also work as epigenetic regulators of other crucial genes in tumor biology, either directly or indirectly. Such regulations are reported in lung, breast, cervical, gastric, colorectal, pancreatic, prostate, and head and neck cancers. Epigenetics of inflammatory mediators in cancer is currently subject of extensive research. These investigations may help in understanding cancer biology and to develop effective therapeutic strategies. The purpose of this paper is to have a brief view of the aberrant regulation of inflammatory cytokines in human malignancies. PMID:25814785

  3. Epigenetic Regulation of Inflammatory Cytokines and Associated Genes in Human Malignancies

    PubMed Central

    Yasmin, Rehana; Hassan, Amjad; Khan, Abdul Rehman; Abbasi, Rashda; Ahmad, Nafees

    2015-01-01

    Inflammation is a multifaceted defense response of immune system against infection. Chronic inflammation has been implicated as an imminent threat for major human malignancies and is directly linked to various steps involved in tumorigenesis. Inflammatory cytokines, interleukins, interferons, transforming growth factors, chemokines, and adhesion molecules have been associated with chronic inflammation. Numerous cytokines are reported to be aberrantly regulated by different epigenetic mechanisms like DNA methylation and histone modifications in tumor tissues, contributing to pathogenesis of tumor in multiple ways. Some of these cytokines also work as epigenetic regulators of other crucial genes in tumor biology, either directly or indirectly. Such regulations are reported in lung, breast, cervical, gastric, colorectal, pancreatic, prostate, and head and neck cancers. Epigenetics of inflammatory mediators in cancer is currently subject of extensive research. These investigations may help in understanding cancer biology and to develop effective therapeutic strategies. The purpose of this paper is to have a brief view of the aberrant regulation of inflammatory cytokines in human malignancies. PMID:25814785

  4. Pro-inflammatory cytokines downregulate Hsp27 and cause apoptosis of human retinal capillary endothelial cells

    PubMed Central

    Nahomi, Rooban B.; Palmer, Allison; Roth, Katelyn E.; Fort, Patrice E.; Nagaraj, Ram H.

    2013-01-01

    The formation of acellular capillaries in the retina, a hallmark feature of diabetic retinopathy, is caused by apoptosis of endothelial cells and pericytes. The biochemical mechanism of such apoptosis remains unclear. Small heat shock proteins play an important role in the regulation of apoptosis. In the diabetic retina, pro-inflammatory cytokines are upregulated. In this study, we investigated the effects of pro-inflammatory cytokines on small heat shock protein 27 (Hsp27) in human retinal endothelial cells (HREC). In HREC cultured in the presence of cytokine mixtures (CM), a significant downregulation of Hsp27 at the protein and mRNA level occurred, with no effect on HSF-1, the transcription factor for Hsp27. The presence of high glucose (25 mM) amplified the effects of cytokines on Hsp27. CM activated indoleamine 2,3-dioxygenase (IDO) and enhanced the production of kynurenine and ROS. An inhibitor of IDO, 1-methyl tryptophan (MT), inhibited the effects of CM on Hsp27. CM also upregulated NOS2 and, consequently, nitric oxide (NO). A NOS inhibitor, L-NAME, and a ROS scavenger blocked the CM-mediated Hsp27 downregulation. While a NO donor in the culture medium did not decrease the Hsp27 content, a peroxynitrite donor and exogenous peroxynitrite did. The cytokines and high glucose-induced apoptosis of HREC were inhibited by MT and L-NAME. Downregulation of Hsp27 by a siRNA treatment promoted apoptosis in HREC. Together, these data suggest that pro-inflammatory cytokines induce the formation of ROS and NO, which, through the formation of peroxynitrite, reduce the Hsp27 content and bring about apoptosis of retinal capillary endothelial cells. PMID:24252613

  5. A comparative analysis of cytokine production and tolerance induction by bacterial lipopeptides, lipopolysaccharides and Staphyloccous aureus in human monocytes.

    PubMed Central

    Kreutz, M; Ackermann, U; Hauschildt, S; Krause, S W; Riedel, D; Bessler, W; Andreesen, R

    1997-01-01

    Monocytes (MO) and macrophages (MAC) are important producers of cytokines involved in the pathophysiology of bacterial sepsis. Most studies concentrate on the effects of bacterial lipopolysaccharides (LPS) regarding the induction of cytokine gene expression and secretion in MO/MAC. Here we report that besides LPS, the synthetic lipoprotein analogue lipopeptide N-palmitoyl-S-(2,3-bis(palmitoyl)-(2RS)-propyl)-(R)-cysteinyl-alanyl- glycine (Pam3-Cys-Ala-Gly), another component of the outer membrane of Gram-negative bacteria, as well as heat-killed Staphyloccocus aureus (S. aureus/SAC) are potent stimuli for cytokines in human MO. For all three investigated stimuli we found an individual pattern of cytokine induction: LPS was most potent in inducing interleukin-6 (IL-6) synthesis, whereas for tumour necrosis factor-alpha (TNF-alpha) secretion SAC was the best stimulus. Comparable amounts of IL-8 were induced by either LPS or Pam3-Cys-Ala-Gly, with SAC being less effective even at higher concentrations. The addition of serum led to an increase in LPS-, SAC- and Pam3-Cys-Ala-Gly-stimulated TNF-alpha secretion, indicating that the presence of serum is critical not just for LPS stimulation. Furthermore, as is known for LPS, Pam3-Cys-Ala-Gly and SAC rendered MO refractory to a second bacterial stimulus. Pam3-Cys-Ala-Gly and SAC induced tolerance for itself, but LPS could partially overcome this effect. As the CD14 molecule is discussed as a common receptor for different bacterial components, we investigated whether the TNF-alpha response of MO could be blocked by anti-CD14 antibodies. MY4, a CD14 antibody, selectively blocked the TNF-alpha secretion induced by LPS but not by Pam3-Cys-Ala-Gly or SAC. In summary, we conclude that besides LPS, lipopeptide Pam3-Cys-Ala-Gly and SAC are potent stimuli for human MO, while the mechanisms of activation seem to be partially different from LPS. Images Figure 2 PMID:9486114

  6. 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. PMID:26921773

  7. 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. PMID:26661249

  8. [Effects of cytokines on somatostatin in nude mice bearing human renal cell carcinoma].

    PubMed

    Li, G; Cao, G; Huo, J

    1997-06-01

    We studied the relationship between the production of SS and treatment with cytokines and a new method for the treatment of renal cell carcinoma. 4.4 x 10(6)RCC94616 cells were injected subcutaneously into the back of nude mice. Five groups with TNF, IL-2, rIFN, TNF + IL-2, TNF + rIFN and controls were randomly divided according to the mean diameter of experimental tumor. After the last injection of cytokines, 0.5-0.8 ml blood, 1g tumor tissue, para-tissue and normal tissue were havested respectively. Contents of SS were tested by radioimmunoassay. In the treatment groups with cytokines, the concentration of SS was changed, siginificantly increased in the TNF + IL-2 group (P < 0.01). The effect on distribution of SS by cytokines may also be mediated by the regulation of human immunity and antitumor activity. It may be suggested that the method of TNF + IL-2 + SS is best to treat renal cell carcinoma. PMID:10374465

  9. 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. PMID:27521270

  10. Stretch-induced human myometrial cytokines enhance immune cell recruitment via endothelial activation

    PubMed Central

    Lee, Yu-Hui; Shynlova, Oksana; Lye, Stephen J

    2015-01-01

    Spontaneous term labour is associated with amplified inflammatory events in the myometrium including cytokine production and leukocyte infiltration; however, potential mechanisms regulating such events are not fully understood. We hypothesized that mechanical stretch of the uterine wall by the growing fetus facilitates peripheral leukocyte extravasation into the term myometrium through the release of various cytokines by uterine myocytes. Human myometrial cells (hTERT-HM) were subjected to static mechanical stretch; stretch-conditioned media was collected and analysed using 48-plex Luminex assay and ELISA. Effect of stretch-conditioned media on cell adhesion molecule expression of human uterine microvascular endothelial cells (UtMVEC-Myo) was detected by quantitative polymerase chain reaction (qPCR) and flow cytometry; functional assays testing leukocyte–endothelial interactions: adhesion of leukocytes to endothelial cells and transendothelial migration of calcein-labelled primary human neutrophils as well as migration of THP-1 monocytic cells were assessed by fluorometry. The current in vitro study demonstrated that mechanical stretch (i) directly induces secretion of multiple cytokines and chemokines by hTERT-HM cells (IL-6, CXCL8, CXCL1, migration inhibitory factor (MIF), VEGF, G-CSF, IL-12p70, bFGF and platelet-derived growth factor subunit B (PDGF-bb), P<0.05); stretch-induced cytokines (ii) enhance leukocyte adhesion to the endothelium of the surrounding uterine microvasculature by (iii) inducing the expression of endothelial cell adhesion molecules and (iv) directing the transendothelial migration of peripheral leukocytes. (vi) Chemokine-neutralizing antibodies and broad-spectrum chemokine inhibitor block leukocyte migration. Our data provide a proof of mechanical regulation for leukocyte recruitment from the uterine blood vessels to the myometrium, suggesting a putative mechanism for the leukocyte infiltrate into the uterus during labour and postpartum

  11. 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

  12. Kinetics of cytokine expression during primary human immunodeficiency virus type 1 infection.

    PubMed Central

    Graziosi, C; Gantt, K R; Vaccarezza, M; Demarest, J F; Daucher, M; Saag, M S; Shaw, G M; Quinn, T C; Cohen, O J; Welbon, C C; Pantaleo, G; Fauci, A S

    1996-01-01

    In the present study, we have determined the kinetics of constitutive expression of a panel of cytokines [interleukin (IL) 2, IL-4, IL-6, IL-10, interferon gamma (IFN-gamma), and tumor necrosis factor alpha (TNF-alpha)] in sequential peripheral blood mononuclear cell samples from nine individuals with primary human immunodeficiency virus infection. Expression of IL-2 and IL-4 was barely detected in peripheral blood mononuclear cells. However, substantial levels of IL-2 expression were found in mononuclear cells isolated from lymph node. Expression of IL-6 was detected in only three of nine patients, and IL-6 expression was observed when transition from the acute to the chronic phase had already occurred. Expression of IL-10 and TNF-alpha was consistently observed in all patients tested, and levels of both cytokines were either stable or progressively increased over time. Similar to IL-10 and TNF-alpha, IFN-gamma expression was detected in all patients; however, in five of nine patients, IFN-gamma expression peaked very early during primary infection. The early peak in IFN-gamma expression coincided with oligoclonal expansions of CD8+ T cells in five of six patients, and CD8+ T cells mostly accounted for the expression of this cytokine. These results indicate that high levels of expression of proinflammatory cytokines are associated with primary infection and that the cytokine response during this phase of infection is strongly influenced by oligoclonal expansions of CD8+ T cells. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 PMID:8633076

  13. 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

  14. 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. PMID:22548238

  15. Cytokine secretion and NK cell activity in human ADAM17 deficiency

    PubMed Central

    Chavkin, Maor; Schmiedel, Dominik; Wong, Eitan; Werner, Marion; Yaacov, Barak; Averbuch, Diana; Molho-Pessach, Vered; Stepensky, Polina; Kaynan, Noa; Bar-On, Yotam; Seidel, Einat; Yamin, Rachel; Sagi, Irit; Elpeleg, Orly; Mandelboim, Ofer

    2015-01-01

    Genetic deficiencies provide insights into gene function in humans. Here we describe a patient with a very rare genetic deficiency of ADAM17. We show that the patient's PBMCs had impaired cytokine secretion in response to LPS stimulation, correlating with the clinical picture of severe bacteremia from which the patient suffered. ADAM17 was shown to cleave CD16, a major NK killer receptor. Functional analysis of patient's NK cells demonstrated that his NK cells express normal levels of activating receptors and maintain high surface levels of CD16 following mAb stimulation. Activation of individual NK cell receptors showed that the patient's NK cells are more potent when activated directly by CD16, albeit no difference was observed in Antibody Depedent Cytotoxicity (ADCC) assays. Our data suggest that ADAM17 inhibitors currently considered for clinical use to boost CD16 activity should be cautiously applied, as they might have severe side effects resulting from impaired cytokine secretion. PMID:26683521

  16. Senescence of human skeletal muscle impairs the local inflammatory cytokine response to acute eccentric exercise.

    PubMed

    Hamada, Koichiro; Vannier, Edouard; Sacheck, Jennifer M; Witsell, Alice L; Roubenoff, Ronenn

    2005-02-01

    The impact of aging on the cytokine response of human skeletal muscle to exercise-induced injury remains poorly understood. We enrolled physically active, young (23-35 years old, n=15) and old (66-78 years old, n=15) men to perform 45 min of downhill running (16% descent) at 75% VO2max. Biopsies of vastus lateralis were obtained 24 h before and 72 h after acute eccentric exercise. Transcripts for inflammatory (TNF-alpha, IL-1beta) and anti-inflammatory cytokines (IL-6, TGF-beta1) were quantified by real-time PCR. Before exercise, cytokine transcripts did not differ with age. At old age, exercise induced a blunted accumulation of transcripts encoding the pan-leukocyte surface marker CD18 (young: 10.1-fold increase, P<0.005; old: 4.7-fold increase, P=0.02; young vs. old: P<0.05). In both age groups, CD18 transcript accumulation strongly correlated with TNF-alpha (young, r=0.87, P<0.001; old, r=0.72, P=0.002) and TGF-beta1 transcript accumulation (young, r=0.80, P<0.001; old, r=0.64, P=0.008). At old age, there was no correlation between IL-1beta and CD18 transcript accumulation. Furthermore, exercise induced IL-6 transcript accumulation in young (3.6-fold, P=0.057) but not in old men. Our results suggest that aging impairs the adaptive response of human skeletal muscle to eccentric exercise by differential modulation of a discrete set of inflammatory and anti-inflammatory cytokine genes. PMID:15556970

  17. 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.

  18. [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. PMID:19353953

  19. Neutrophil-mediated damage to human vascular endothelium. Role of cytokine activation.

    PubMed Central

    Westlin, W. F.; Gimbrone, M. A.

    1993-01-01

    Cytokine activation of cultured human vascular endothelial cells renders them hyperadhesive for blood leukocytes. Co-incubation of freshly isolated, unstimulated human blood neutrophils with confluent cytokine-activated human endothelial monolayers for 90 minutes results in extensive endothelial detachment and destruction of monolayer integrity. In contrast, unactivated endothelial monolayers remain intact. Using this in vitro model, we have explored the neutrophil-effector mechanisms involved in this injury. Coincubation in the presence of a serine protease inhibitor (phenylmethylsulfonyl fluoride) or specific elastase inhibitors (Ala-Ala-Pro-Val-chloromethyl ketone or alpha-1-protease inhibitor) markedly diminished injury. In contrast, scavengers or inhibitors of oxygen-derived free radicals (superoxide dismutase, catalase, mannitol, or sodium azide) were not protective. Purified human neutrophil elastase mimicked the effect of the neutrophils suggesting a key role for elastase in the neutrophil-mediated injury in this model. Interfering with direct neutrophil-endothelial cell contact by interposing a microporous barrier insert prevented endothelial cell detachment. Furthermore, this neutrophil-mediated detachment could be inhibited with interleukin-8, an action correlated with a decrease in neutrophil adhesion to activated endothelial monolayers. By defining the role of endothelial activation in neutrophil-mediated injury, this in vitro model may provide useful insights into potential therapeutic interventions designed to prevent disruption of the endothelial barrier function. Images Figure 1 Figure 6 PMID:8424450

  20. Differential Inhibition of T Lymphocyte Proliferation and Cytokine Synthesis by [6]-Gingerol, [8]-Gingerol, and [10]-Gingerol.

    PubMed

    Bernard, Megan; Furlong, Suzanne J; Power Coombs, Melanie R; Hoskin, David W

    2015-11-01

    [6]-Gingerol, [8]-gingerol, and [10]-gingerol are pungent components of fresh ginger, extracts of which inhibit various components of the inflammatory response. Because little is known regarding the effect of gingerols with different unbranched alkyl side chain lengths on the activation and effector function of T lymphocytes, we compared the effects of [6]-gingerol, [8]-gingerol, and [10]-gingerol on murine T lymphocyte proliferation, expression of CD25 and CD69 activation markers, cytokine synthesis, and interleukin (IL)-2 receptor signaling. All three gingerols inhibited DNA synthesis by T lymphocytes, as well as interferon-γ synthesis. In contrast, only [8]-gingerol and [10]-gingerol inhibited CD25 and CD69 expression, and IL-2 synthesis. None of the gingerols affected IL-4 synthesis. Exogenous IL-2 enhanced T lymphocyte proliferation in the presence of [6]-gingerol but did not significantly increase T lymphocyte proliferation in the presence of [8]-gingerol or [10]-gingerol. In line with this finding, [8]-gingerol and [10]-gingerol impaired IL-2-induced proliferation of CTLL-2 cells, but constitutive CD25 expression was unaffected, indicating inhibition of IL-2 receptor signaling. In general, [10]-gingerol and [8]-gingerol were more potent inhibitors of T lymphocytes than [6]-gingerol. Suppression of T lymphocyte responses by gingerols suggests that these phytochemicals may be beneficial in chronic inflammatory conditions associated with excessive or inappropriate T lymphocyte activation. PMID:26178781

  1. Modulation of the Cytokine Response in Human Monocytes by Mycobacterium leprae Phenolic Glycolipid-1

    PubMed Central

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

    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. PMID:21981546

  2. 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

  3. Multiplexed femtomolar quantitation of human cytokines in a fluoropolymer microcapillary film.

    PubMed

    Castanheira, Ana P; Barbosa, Ana I; Edwards, Alexander D; Reis, Nuno M

    2015-08-21

    Sensitive quantitation of multiple cytokines can provide important diagnostic information during infection, inflammation and immunopathology. In this study sensitive immunoassay detection of human cytokines IL-1β, IL-6, IL-12p70 and TNFα is shown for singleplex and multiplex formats using a novel miniaturized ELISA platform. The platform uses a disposable plastic multi-syringe aspirator (MSA) integrating 8 disposable fluoropolymer microfluidic test strips, each containing an array of ten 200 μm mean i.d. microcapillaries coated with a set of monoclonal antibodies. Each MSA device thus performs 10 tests on 8 samples, delivering 80 measurements. Unprecedented levels of sensitivity were obtained with the novel fluoropolymer microfluidic material and simple colorimetric detection in a flatbed scanner. The limit of detection for singleplex detection ranged from 2.0 to 15.0 pg ml(-1), i.e. 35 and 713 femtomolar for singleplex cytokine detection, and the intra- and inter-assay coefficient of variation (CV) remained within 10%. In addition, a triplex immunoassay was developed for measuring IL-1β, IL-12p70 and TNFα simultaneously from a given sample in the pg ml(-1) range. These assays permit high sensitivity measurement with rapid <15 min assay or detection from undiluted blood serum. The portability, speed and low-cost of this system are highly suited to point-of-care testing and field diagnostics applications. PMID:26120601

  4. Immunomodulatory capacity of fungal proteins on the cytokine production of human peripheral blood mononuclear cells.

    PubMed

    Jeurink, Prescilla V; Noguera, Cristina Lull; Savelkoul, Huub F J; Wichers, Harry J

    2008-08-01

    Immunomodulation by fungal compounds can be determined by the capacity of the compounds to influence the cytokine production by human peripheral blood mononuclear cells (hPBMC). These activities include mitogenicity, stimulation and activation of immune effector cells. Eight mushroom strains (Agaricus blazei, Coprinus comatus, Flammulina velutipes, Ganoderma lucidum, Grifola frondosa, Volvariella volvacea, Lentinus edodes, and Pleurotus ostreatus) were tested for the immunomodulating activity of the isolated protein fractions and polysaccharides fractions present in mycelia and culture liquid. The fungal proteins and polysaccharides have been investigated for their in vitro effect on the cytokine profile (IFN-gamma, IL-4, IL-10, IL-12 and TNF-alpha) of unstimulated or hPBMC stimulated with the polyclonal stimulations PMA/Ca-I, ConA or LPS. In addition to their influence on the cytokine profile, the hemagglutination activity of the fungal proteins on rabbit red blood cells was determined. Proteins from V. volvacea and G. lucidum showed immunomodulating activity without the presence of any mitogen, however, neither of them decreased the production of IL-4 and IFN-gamma in combination with a stimulus. All used stimuli resulted in an induction of IL-12 in the presence of the protein extracts, suggesting a direct effect on monocytes. This effect might lead to the indirect immunomodulation of T cell activation and cytokine production. In addition, both protein extracts showed more hemagglutination activity after trypsin treatment of the rabbit red blood cells, indicating the presence of carbohydrate-binding proteins, like lectins and FIPs. In conclusion, the protein extracts of V. volvacea and G. lucidum contain immunomodulating activity by acting directly on monocytes and thereby modulating T cell activation. Further purification of the fungal extracts is needed to clarify whether there are FIPs or lectins present that are responsible for this immunomodulating activity

  5. Leukotriene E4 activates human Th2 cells for exaggerated proinflammatory cytokine production in response to prostaglandin D2.

    PubMed

    Xue, Luzheng; Barrow, Anna; Fleming, Vicki M; Hunter, Michael G; Ogg, Graham; Klenerman, Paul; Pettipher, Roy

    2012-01-15

    PGD(2) exerts a number of proinflammatory responses through a high-affinity interaction with chemoattractant receptor-homologous molecule expressed on Th2 cells (CRTH2) and has been detected at high concentrations at sites of allergic inflammation. Because cysteinyl leukotrienes (cysLTs) are also produced during the allergic response, we investigated the possibility that cysLTs may modulate the response of human Th2 cells to PGD(2). PGD(2) induced concentration-dependent Th2 cytokine production in the absence of TCR stimulation. Leukotrienes D(4) and E(4) (LTE(4)) also stimulated the cytokine production but were much less active than PGD(2). However, when combined with PGD(2), cysLTs caused a greater than additive enhancement of the response, with LTE(4) being most effective in activating Th2 cells. LTE(4) enhanced calcium mobilization in response to PGD(2) in Th2 cells without affecting endogenous PGD(2) production or CRTH2 receptor expression. The effect of LTE(4) was inhibited by montelukast but not by the P2Y(12) antagonist methylthioadenosine 5'-monophosphate. The enhancing effect was also evident with endogenous cysLTs produced from immunologically activated mast cells because inhibition of cysLT action by montelukast or cysLT synthesis by MK886, an inhibitor of 5-lipoxygenase-activating protein, reduced the response of Th2 cells to the levels produced by PGD(2) alone. These findings reveal that cysLTs, in particular LTE(4), have a significant proinflammatory impact on T cells and demonstrate their effects on Th2 cells are mediated by a montelukast-sensitive receptor. PMID:22174450

  6. Activation of natural killer cells and cytokine production in humans by bacterial extracts (OM-85 BV).

    PubMed

    Wybran, J; Libin, M; Schandene, L

    1990-01-01

    The influence of Broncho-Vaxom (BV) on different immune parameters was investigated in vitro on human peripheral blood mononuclear cells (PBMC). It was found that BV enhances the natural killer (NK) activity of PBMC and increases their spontaneous and phytohemagglutin (PHA)-induced production of tumor-necrosis factor--alpha and interleukin-2 as well as the PHA-stimulated production of interferon-gamma. These immunostimulating actions of BV on NK activity and cytokine production can contribute to the understanding of the enhancement of the body's defense mechanisms against respiratory tract infections. PMID:2117183

  7. T(H)2 cytokines modulate the IL-9R expression on human neutrophils.

    PubMed

    Dragon, Stéphane; Takhar, Manrit Kaur; Shan, Lianyu; Hayglass, Kent T; Simons, F Estelle; Gounni, Abdelilah S

    2009-06-26

    Interleukin (IL)-9 is associated with key pathological features of asthma such as airway hyperresponsiveness, bronchoconstriction and mucus production. Inflammatory responses mediated by IL-9 rely on the expression of the IL-9R which has been reported on lung epithelial cells, T lymphocytes and recently on airway granulocyte infiltrates. In this study, we assessed the regulatory and constitutive cell surface expression of the IL-9Ralpha in unfractionated and purified human neutrophils from atopic asthmatics, atopic non-asthmatics and healthy normal controls. We demonstrate that T(H)2 cytokines (IL-4 or IL-13) and granulocyte macrophage-colony stimulating factor (GM-CSF) up-regulated mRNA and cell surface expression levels of the IL-9Ralpha in primary human and HL-60 differentiated neutrophils. Pharmacological inhibition of NF-kappaB did not affect T(H)2-mediated IL-9Ralpha expression in human neutrophils although IFN-gamma and IL-10 down-regulated IL-9Ralpha expression when co-incubated with IL-4, IL-13 or GM-CSF. Collectively, our results reveal a regulatory function for IFN-gamma and IL-10 on modulating the inducible IL-9Ralpha expression levels on peripheral blood neutrophils by T(H)2 cytokines. PMID:19401191

  8. 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. PMID:26084699

  9. 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

  10. 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

    2015-10-01

    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

  11. In vivo expression of immunosuppressive cytokines in human papillomavirus-transformed cervical cancer cells.

    PubMed

    Alcocer-González, Juan Manuel; Berumen, Jaime; Taméz-Guerra, Reyes; Bermúdez-Morales, Víctor; Peralta-Zaragoza, Oscar; Hernández-Pando, Rogelio; Moreno, José; Gariglio, Patricio; Madrid-Marina, Vicente

    2006-01-01

    Genital human Papillomavirus infection is common and only a minor fraction of infected subjects develop progressing cervical epithelial lesions or cancer. Bypassing local immune responses is important for the development of cervical cancer. In this work we determined the cytokine pattern in samples from patients with cervical cancer. Thus, we examined the local mRNA expression profile of helper T cell type 1 (Th1), Th2, and Th3 cytokines in HPV-positive cervical cancer biopsies by reverse transcription-polymerase chain reaction. Our data indicate that 80% of the tumors expressed low levels of CD4 mRNA, with all of them expressing higher CD8 mRNA levels. Most tumors expressed interleukin (IL)-4 and IL-10 mRNAs and, most importantly, all of them expressed transforming growth factor (TGF)-beta1 and interferon gamma mRNA. None of the tumors studied expressed IL-12, IL-6, or tumor necrosis factor (TNF) mRNA. Immunohistochemical analysis identified IL-10 only in tumor cells and koilocytic cells, but not in tumor-infiltrating lymphocytes, suggesting that IL-10-producing cells are those transformed by HPV. We found a correlation between immunostaining for IL-10 protein and the level of IL-10 mRNA expression. Moreover, supernatants from HPV-transformed cell cultures contained IL-10 and TGF- beta1. Our findings indicate a predominant expression of immunosuppressive cytokines, which might help downregulate tumor-specific immune responses in the microenvironment of the tumor. This information may be useful for cervical cancer immunotherapies or for therapeutic vaccine design against Human Papillomavirus. PMID:16987066

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

    PubMed

    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

  13. 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

  14. 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. PMID:27529696

  15. 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

  16. An in vitro model for the study of human bone marrow angiogenesis: role of hematopoietic cytokines.

    PubMed

    Pelletier, L; Regnard, J; Fellmann, D; Charbord, P

    2000-04-01

    This study describes a human bone marrow endothelial cell culture in which endothelial cells are organized into capillary tubes. These endothelial cells were positive for von Willebrand Factor, expressed CD34, CD31, and L-fucose residues, took up acetylated low-density lipoproteins, contained Weibel-Palade bodies, and were ensheathed in a basal lamina (which included laminin beta1, EDa+ and EDb+ fibronectin, and collagen type iv). Pericytes expressing alpha-smooth muscle (alpha-SM) actin were spatially associated with the capillary tubes and there was a highly significant correlation between the number of capillary tubes and pericytes. In this model, basal angiogenesis was found to be vascular endothelial growth factor (VEGF)-dependent, because neutralization of endogenous VEGF induced a dramatic regression in the number of tubes. However, the presence of alpha-SM actin-expressing pericytes in the linings of endothelial tubes partially prevented the VEGF-neutralized tube regression. We also observed that nitric oxide production contributed to basal angiogenesis and that upregulation of nitric oxide increased the number of tubes. Tube numbers also decreased when antibodies neutralizing the integrin alphavbeta5 were applied to the cultures. Moreover, addition of any of the hematopoietic cytokines, erythropoietin, stem cell factor, granulocytic colony stimulating factor, or granulomonocytic colony stimulating factor induced a highly significant increase in tube formation. When erythropoietin and granulocytic colony stimulating factor were added, this increase was larger than the maximum increase observed with VEGF. Thus, we have described an in vitro model for human bone marrow angiogenesis in which pericytes and basal lamina matrix were associated with endothelial cells and formed fully organized capillary tubes. In this model, cytokines known to regulate hematopoiesis also seemed to be mediators of angiogenesis. This culture system may therefore prove to be a

  17. Isolation and cytokine analysis of lamina propria lymphocytes from mucosal biopsies of the human colon.

    PubMed

    Bowcutt, Rowann; Malter, Lisa B; Chen, Lea Ann; Wolff, Martin J; Robertson, Ian; Rifkin, Daniel B; Poles, Michael; Cho, Ilseug; Loke, P'ng

    2015-06-01

    Much of our understanding of gut-microbial interactions has come from mouse models. Intestinal immunity is complex and a combination of host genetics and environmental factors play a significant role in regulating intestinal immunity. Due to this complexity, no mouse model to date gives a complete and accurate representation of human intestinal diseases, such as inflammatory bowel diseases. However, intestinal tissue from patients undergoing bowel resection reflects a condition of severe disease that has failed treatment; hence a more dynamic perspective of varying inflammatory states in IBD could be obtained through the analyses of pinch biopsy material. Here we describe our protocol for analyzing mucosal pinch biopsies collected predominantly during colonoscopies. We have optimized flow cytometry panels to analyze up to 8 cytokines produced by CD4+ and CD8+ cells, as well as for characterizing nuclear proteins and transcription factors such as Ki67 and Foxp3. Furthermore, we have optimized approaches to analyze the production of cytokines, including TGF-beta from direct ex vivo cultures of pinch biopsies and LPMCs isolated from biopsies. These approaches are part of our workflow to try and understand the role of the gut microbiota in complex and dynamic human intestinal diseases. PMID:25769417

  18. 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.

  19. Variations in cytokine mRNA expression during normal human pregnancy

    PubMed Central

    Kruse, N; Greif, M; Moriabadi, N F; Marx, L; Toyka, K V; Rieckmann, P

    2000-01-01

    Epidemiological data provide evidence that disease activity of T cell-mediated, organ-specific autoimmune diseases is reduced during pregnancy. Although there are several experimental animal studies on the effect of pregnancy on the immune system, the situation in humans is less clear. We therefore performed a prospective analysis of cytokine mRNA expression in whole blood by a new on-line reverse transcriptase-polymerase chain reaction technique and of serum hormone levels during pregnancy in healthy women. The control group included age-matched non-pregnant healthy women. Quantitativecytokine mRNA expression revealed significantly reduced IL-18, interferon-gamma (IFN-γ), and IL-2 mRNA levels in the first and second trimester in pregnancy compared with non-pregnant women. No difference between groups was detected for tumour necrosis factor-alpha (TNF-α) mRNA. IL-4 and IL-10 mRNA were detected at low levels in only 20% of pregnant women and were reduced to a statistically significant extent in the second and third trimester compared with the control group. Changes in IL-18 mRNA expression correlated inversely with serum values for human choriogonadotropin (HCG) and IL-10 serum levels correlated with increases in serum 17β-oestradiol levels. These data indicate immunomodulatory effects of pregnancy at the cytokine level which may be related to the variations in the clinical course of organ-specific, T cell-mediated autoimmune diseases during pregnancy. PMID:10632669

  20. The cytokines effect on EBV-immortalized human B cells producing antisperm antibody.

    PubMed

    Fiszer, D; Niedbała, W; Fernandez, N; Kurpisz, M

    1992-01-01

    We have analyzed the antisperm antibody production of autoimmunized male subjects using Epstein-Barr virus (EBV) immortalization of B lymphocytes. We evaluated the influence of several in vitro culture variants applied prior to EBV infection on the frequence of antibody-producing cells and affinity of secreted antibodies. The following variants were applied: a) polyclonal antigenic stimulation of lymphocytes with PWM, b) PWM (pokeweed mitogen) + IL-2 + interferon gamma and c) PWM + IL-2 + interferon gamma + sperm antigenic extract. The variants where the cytokines were added did not increase the frequency of EBV-infected antibody-producing cells as comparing to EBV infection previously amplified by the use of polyclonal activator. Furthermore the cytokine activation either in combination with mitogen or in vitro secondary antigenic sensitization (prior to EBV transformation) did not seem to have beneficial effect on affinity of antibodies produced by EBV-infected cells in comparison to straight EBV infection. On the other hand, the attempt to promote an immunoglobulin secretion (IgM) by previously obtained human-human antisperm hybridomas by adding of IL-2 was quite successful. PMID:1338684

  1. Cytokine regulation of human lung fibroblast hyaluronan (hyaluronic acid) production. Evidence for cytokine-regulated hyaluronan (hyaluronic acid) degradation and human lung fibroblast-derived hyaluronidase.

    PubMed Central

    Sampson, P M; Rochester, C L; Freundlich, B; Elias, J A

    1992-01-01

    We characterized the mechanisms by which recombinant (r) tumor necrosis factor (TNF), IFN-gamma, and IL-1, alone and in combination, regulate human lung fibroblast hyaluronic acid (HA) production. Each cytokine stimulated fibroblast HA production. The combination of rTNF and rIFN-gamma resulted in a synergistic increase in the production of high molecular weight HA. This was due to a synergistic increase in hyaluronate synthetase activity and a simultaneous decrease in HA degradation. In contrast, when rTNF and rIL-1 were combined, an additive increase in low molecular weight HA was noted. This was due to a synergistic increase in hyaluronate synthetase activity and a simultaneous increase in HA degradation. Human lung fibroblasts contained a hyaluronidase that, at pH 3.7, depolymerized high molecular weight HA to 10-40 kD end products of digestion. However, hyaluronidase activity did not correlate with fibroblast HA degradation. Instead, HA degradation correlated with fibroblast-HA binding, which was increased by rIL-1 plus rTNF and decreased by rIFN-gamma plus rTNF. Recombinant IL-1 and rTNF weakly stimulated and rIL-1 and rTNF in combination further augmented the levels of CD44 mRNA in lung fibroblasts. In contrast, rIFN-gamma did not significantly alter the levels of CD44 mRNA in unstimulated or rTNF stimulated cells. These studies demonstrate that rIL-1, rTNF, and rIFN-gamma have complex effects on biosynthesis and degradation which alter the quantity and molecular weight of the HA produced by lung fibroblasts. They also show that fibroblast HA degradation is mediated by a previously unrecognized lysosomal-type hyaluronidase whose function may be regulated by altering fibroblast-HA binding. Lastly, they suggest that the CD44 HA receptor may be involved in this process. Images PMID:1401082

  2. Lactobacillus rhamnosus GG and Streptococcus thermophilus induce suppressor of cytokine signalling 3 (SOCS3) gene expression directly and indirectly via interleukin-10 in human primary macrophages

    PubMed Central

    Latvala, S; Miettinen, M; Kekkonen, R A; Korpela, R; Julkunen, I

    2011-01-01

    In the present study we have characterized T helper type 2 (Th2) [interleukin (IL)-10]/Th1 (IL-12) cytokine expression balance in human primary macrophages stimulated with multiple non-pathogenic Gram-positive bacteria used in the food industry and as probiotic substances. Bacteria representing Lactobacillus, Bifidobacterium, Lactococcus, Leuconostoc, Propionibacterium and Streptococcus species induced anti-inflammatory IL-10 production, although quantitative differences between the bacteria were observed. S. thermophilus was able to induce IL-12 production, while the production of IL-12 induced by other bacteria remained at a low level. The highest anti-inflammatory potential was seen with bifidobacteria, as evidenced by high IL-10/IL-12 induction ratios. All studied non-pathogenic bacteria were able to stimulate the expression of suppressor of cytokine signalling (SOCS) 3 that controls the expression of proinflammatory cytokine genes. Lactobacillus and Streptococcus species induced SOCS3 mRNA expression directly in the absence of protein synthesis and indirectly via bacteria-induced IL-10 production, as demonstrated by experiments with cycloheximide (CHX) and anti-IL-10 antibodies, respectively. The mitogen-activated protein kinase (MAPK) p38 signalling pathway played a key role in bacteria-induced SOCS3 gene expression. Enhanced IL-10 production and SOCS3 gene expression induced by live non-pathogenic Lactobacillus and Streptococcus is also likely to contribute to their immunoregulatory effects in vivo. PMID:21545585

  3. Role of the AMP kinase in cytokine-induced human EndoC-βH1 cell death.

    PubMed

    Fred, Rikard G; Kappe, Camilla; Ameur, Adam; Cen, Jing; Bergsten, Peter; Ravassard, Phillippe; Scharfmann, Raphael; Welsh, Nils

    2015-10-15

    The aim of the present investigation was to delineate cytokine-induced signaling and death using the EndoC-βH1 cells as a model for primary human beta-cells. The cytokines IL-1β and IFN-γ induced a rapid and transient activation of NF-κB, STAT-1, ERK, JNK and eIF-2α signaling. The EndoC-βH1 cells died rapidly when exposed to IL-1β + IFN-γ, and this occurred also in the presence of the actinomycin D. Inhibition of NF-κB and STAT-1 did not protect against cell death, nor did the cytokines activate iNOS expression. Instead, cytokines promoted a rapid decrease in EndoC-βH1 cell respiration and ATP levels, and we observed protection by the AMPK activator AICAR against cytokine-induced cell death. It is concluded that EndoC-βH1 cell death can be prevented by AMPK activation, which suggests a role for ATP depletion in cytokine-induced human beta-cell death. PMID:26213325

  4. Rotavirus Infects Human Biliary Epithelial Cells and Stimulates Secretion of Cytokines IL-6 and IL-8 via MAPK Pathway

    PubMed Central

    Clemente, Maria Grazia; Patton, John T.; Anders, Robert A.; Yolken, Robert H.; Schwarz, Kathleen B.

    2015-01-01

    Biliary atresia (BA) is an infantile inflammatory cholangiopathy of unknown etiology although epidemiologic studies and animal models utilizing rotavirus (RV) have suggested a role for viral infection. Proinflammatory and profibrotic cytokines have been detected in infants with BA. The purpose of our study was to investigate the susceptibility of human cholangiocytes (H69 cells) to infection with RRV and to determine if this infection resulted in cytokine secretion. Infection of H69 cells by RRV was noncytolytic and resulted in a time-dependent increase in the release of both infectious virions and cytokines IL-6 and IL-8 into the supernate. The greatest difference in cytokine supernatant levels between infected and mock-infected cells was noted at 24 hours postinfection (h p.i.) for IL-8, 556 ± 111 versus 77 ± 68 pg/mL (p < 0.0001), and at 48 h p.i. for IL-6, 459 ± 64 versus 67 ± 2 pg/mL (p < 0.0001). Production of both cytokines following RRV infection was significantly reduced by pretreating the H69 cells with inhibitors of mitogen-activated protein kinase (MAPK). Conclusion. RRV can infect human cholangiocytes resulting in the production of proinflammatory and profibrotic cytokines via the MAPK pathway. RRV-infected H69 cells could be a useful model system for investigating the viral hypothesis of BA. PMID:26247025

  5. TSLP is differentially regulated by vitamin D3 and cytokines in human skin

    PubMed Central

    Landheer, Janneke; Giovannone, Barbara; Sadekova, Svetlana; Tjabringa, Sandra; Hofstra, Claudia; Dechering, Koen; Bruijnzeel-Koomen, Carla; Chang, Charlie; Ying, Yu; de Waal Malefyt, Rene; Hijnen, DirkJan; Knol, Edward

    2015-01-01

    Thymic stromal lymphopoietin (TSLP) plays an important role in allergic diseases and is highly expressed in keratinocytes in human lesional atopic dermatitis (AD) skin. In nonlesional AD skin TSLP expression can be induced by applying house dust mite allergen onto the skin in the atopy patch test. Several studies have demonstrated that the induction of TSLP expression in mouse skin does not only lead to AD-like inflammation of the skin, but also predisposes to severe inflammation of the airways. In mice, TSLP expression can be induced by application of the 1,25-dihydroxyvitamin D3 (VD3) analogue calcipotriol and results in the development of eczema-like lesions. The objective is to investigate the effect of VD3 (calcitriol) or calcipotriol on TSLP expression in normal human skin and skin from AD patients. Using multiple ex vivo experimental setups, the effects of calci(po)triol on TSLP expression by normal human skin, and skin from AD patients were investigated and compared to effects of calcipotriol on mouse and non-human primates (NHP) skin. No induction of TSLP expression (mRNA or protein) was observed in human keratinocytes, normal human skin, nonlesional AD skin, or NHP skin samples after stimulation with calcipotriol or topical application of calcitriol. The biological activity of calci(po)triol in human skin samples was demonstrated by the increased expression of the VD3-responsive Cyp24a1 gene. TSLP expression was induced by cytokines (IL-4, IL-13, and TNF-α) in skin samples from all three species. In contrast to the findings in human and NHP, a consistent increase in TSLP expression was confirmed in mouse skin biopsies after stimulation with calcipotriol. VD3 failed to induce expression of TSLP in human or monkey skin in contrast to mouse, implicating careful extrapolation of this often-used mouse model to AD patients. PMID:25866638

  6. 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. PMID:24908297

  7. The effect of pro-inflammatory cytokines on immunophenotype, differentiation capacity and immunomodulatory functions of human mesenchymal stem cells.

    PubMed

    Pourgholaminejad, Arash; Aghdami, Nasser; Baharvand, Hossein; Moazzeni, Seyed Mohammad

    2016-09-01

    Mesenchymal stem cells (MSCs), as cells with potential clinical utilities, have demonstrated preferential incorporation into inflammation sites. Immunophenotype and immunomodulatory functions of MSCs could alter by inflamed-microenvironments due to the local pro-inflammatory cytokine milieu. A major cellular mediator with specific function in promoting inflammation and pathogenicity of autoimmunity are IL-17-producing T helper 17 (Th17) cells that polarize in inflamed sites in the presence of pro-inflammatory cytokines such as Interleukin-1β (IL-1β), IL-6 and IL-23. Since MSCs are promising candidate for cell-based therapeutic strategies in inflammatory and autoimmune diseases, Th17 cell polarizing factors may alter MSCs phenotype and function. In this study, human bone-marrow-derived MSCs (BM-MSC) and adipose tissue-derived MSCs (AD-MSC) were cultured with or without IL-1β, IL-6 and IL-23 as pro-inflammatory cytokines. The surface markers and their differentiation capacity were measured in cytokine-untreated and cytokine-treated MSCs. MSCs-mediated immunomodulation was analyzed by their regulatory effects on mixed lymphocyte reaction (MLR) and the level of IL-10, TGF-β, IL-4, IFN-γ and TNF-α production as immunomodulatory cytokines. Pro-inflammatory cytokines showed no effect on MSCs morphology, immunophenotype and co-stimulatory molecules except up-regulation of CD45. Adipogenic and osteogenic differentiation capacity increased in CD45+ MSCs. Moreover, cytokine-treated MSCs preserved the suppressive ability of allogeneic T cell proliferation and produced higher level of TGF-β and lower level of IL-4. We concluded pro-inflammatory cytokines up-regulate the efficacy of MSCs in cell-based therapy of degenerative, inflammatory and autoimmune disorders. PMID:27288632

  8. 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. PMID:27357508

  9. Human Cytomegalovirus-Induces Cytokine Changes in the Placenta with Implications for Adverse Pregnancy Outcomes

    PubMed Central

    Hamilton, Stuart T.; Scott, Gillian; Naing, Zin; Iwasenko, Jenna; Hall, Beverley; Graf, Nicole; Arbuckle, Susan; Craig, Maria E.; Rawlinson, William D.

    2012-01-01

    Human cytomegalovirus (CMV) infection of the developing fetus can result in adverse pregnancy outcomes including death in utero. Fetal injury results from direct viral cytopathic damage to the CMV-infected fetus, although evidence suggests CMV placental infection may indirectly cause injury to the fetus, possibly via immune dysregulation with placental dysfunction. This study investigated the effects of CMV infection on expression of the chemokine MCP-1 (CCL2) and cytokine TNF-α in placentae from naturally infected stillborn babies, and compared these changes with those found in placental villous explant histocultures acutely infected with CMV ex vivo. Tissue cytokine protein levels were assessed using quantitative immunohistochemistry. CMV-infected placentae from stillborn babies had significantly elevated MCP-1 and TNF-α levels compared with uninfected placentae (p = 0.001 and p = 0.007), which was not observed in placentae infected with other microorganisms (p = 0.62 and p = 0.71) (n = 7 per group). Modelling acute clinical infection using ex vivo placental explant histocultures showed infection with CMV laboratory strain AD169 (0.2 pfu/ml) caused significantly elevated expression of MCP-1 and TNF-α compared with uninfected explants (p = 0.0003 and p<0.0001) (n = 25 per group). Explant infection with wild-type Merlin at a tenfold lower multiplicity of infection (0.02 pfu/ml), caused a significant positive correlation between increased explant infection and upregulation of MCP-1 and TNF-α expression (p = 0.0001 and p = 0.017). Cytokine dysregulation has been associated with adverse outcomes of pregnancy, and can negatively affect placental development and function. These novel findings demonstrate CMV infection modulates the placental immune environment in vivo and in a multicellular ex vivo model, suggesting CMV-induced cytokine modulation as a potential initiator and/or exacerbator of placental and fetal injury. PMID

  10. Effects of ethanol on cytokine generation and NFkappaB activity in human lung epithelial cell.

    PubMed

    Johansson, Anne-Sofie M; Lidén, Johan; Okret, Sam; Palmblad, Jan E W

    2005-08-15

    Alcohol abuse is associated with enhanced risk for pulmonary infections, but the mechanisms remain obscure. We assessed whether ethanol reduced generation of cytokines from a human lung epithelial cell line (A549) in vitro and if effects on the NFkappaB transcription factor were involved. Exposure of A549 to ethanol (0.1-1%) dose-dependently inhibited (by 15-49%) the release of G-CSF and IL-8, but not of M-CSF, triggered by IL1beta or TNFalpha. Ethanol also inhibited by 49% the IL-1beta stimulated translocation of the p65 subunit of NFkappaB from the cytoplasm into the nucleus. Using a kappaB binding and luciferase coupled construct, transfected into A549 cells, we found that 1% ethanol specifically reduced IL-1beta and TNFalpha induced luciferase activity with 34 and 40%, respectively. Thus, in vitro exposure of lung epithelial cells to ethanol reduced the generation of cytokines, as well as translocation and gene activation by NFkappaB. PMID:15993849

  11. Stroke-induced migration of human umbilical cord blood cells: time course and cytokines.

    PubMed

    Newman, Mary B; Willing, Alison E; Manresa, John J; Davis-Sanberg, Cyndy; Sanberg, Paul R

    2005-10-01

    The therapeutic window for treatment of individuals after stroke is narrow, regardless of the treatment regime; extension of this window would provide a major therapeutic advance. In prior reports, we demonstrated significant improvements in the behavioral defects of rats that received human umbilical cord blood (HUCB) cells 24 h after a middle cerebral arterial occlusion. These effects paralleled the recruitment of these cells to the site of tissue damage. While the administration of HUCB cells 24 h after stroke was effective, the optimal time to administer these cells after stroke has not been established. Here, we investigated the migration of HUCB cells to ischemic tissue extracts. After ischemic assault, brain tissue was homogenized, and the supernatants were assayed for their ability to attract HUCB mononuclear cells as well as for levels of several cytokines. We demonstrate increased migratory activity of HUCB cells toward the extracts harvested at 24-72 h after stroke. The extracts possessed increased levels of certain cytokines and chemokines, suggesting their participation in HUCB cell migration. The results from this study are promising in that the current 3-h therapeutic window for the treatment of stroke victims, using approved anticoagulant treatment, may be extended with the use of HUCB cell therapy 24-72 h post stroke. Last, the chemokines present in the supernatant provide a sound starting point to start examining the mechanisms responsible for the in vivo migration of HUCB cells after the induction of stroke. PMID:16305342

  12. Complement regulatory protein expression by a human oligodendrocyte cell line: cytokine regulation and comparison with astrocytes.

    PubMed Central

    Gasque, P; Morgan, B P

    1996-01-01

    Rat oligodendrocytes spontaneously activate complement (C) and lack the C inhibitor CD59. As a consequence, rat oligodendrocytes are susceptible to lysis by autologous C in vitro. Expression of C inhibitors on human oligodendrocytes in vitro and other human glia has yet to be well characterized. We have previously shown expression at the mRNA level of the membrane inhibitors CD59, decay-accelerating factor (DAF; CD55) and membrane cofactor protein (MCP; CD46) in human astrocytes. We here examine the expression of membrane and secreted C inhibitors by the oligodendrocyte cell line, HOG. HOG cells abundantly expressed CD59, assessed at protein and mRNA level, and expressed DAF and MCP, albeit at a lower level. Expression of all three inhibitors was enhanced by incubation with interferon-gamma or with phorbol ester (PMA). Complement receptor type 1 (CR1; CD35) was neither expressed constitutively nor induced by cytokines. HOG also constitutively secreted C1-inhibitor, S-protein and clusterin. Factor H was secreted only after stimulation with cytokines. C4b binding protein was expressed at a very low level and was detected only at the mRNA level by reverse transcriptase-polymerase chain reaction (RT-PCR). For comparison, astrocyte expression of CD59, DAF, MCP and CR1 was confirmed at the mRNA and protein levels. HOG did not activate C spontaneously, as judged by the lack of deposition of C fragments, and were not lysed by C even after inhibition of CD59 and DAF using specific monoclonal antibodies. Images Figure 1 Figure 2 Figure 3 Figure 4 PMID:8958045

  13. 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

  14. Induction of cytokine synthesis and fever suppresses REM sleep and improves mood in patients with major depression.

    PubMed

    Bauer, J; Hohagen, F; Gimmel, E; Bruns, F; Lis, S; Krieger, S; Ambach, W; Guthmann, A; Grunze, H; Fritsch-Montero, R

    1995-11-01

    Beneficial effects of inflammatory events on certain psychiatric disorders, including depression, were reported sporadically by ancient Greek physicians, but have been described also in our times by a few psychiatrists during the past decades. During febrile inflammatory events, mediators of the immune system such as interleukin-1 can be detected in the brain and may act on their respective receptors which have also been demonstrated in the brain. Since cytokines such as interleukin-1 have been shown in animal studies to exert sedative behavioral effects, to be somnogenic, and to induce slow-wave sleep (SWS), we performed a pilot study to evaluate scientifically the anecdotically reported beneficial effects of inflammatory states on depressive disorders. Mood and sleep parameters were monitored in seven drug-free, severely depressed patients before, during, and after the administration of a single dose of endotoxin. All patients responded with a short pulse of increased synthesis of the cytokines tumor necrosis factor, interleukin-1, and interleukin-6 and elevated body temperature for several hours. During the night following endotoxin administration, rapid eye movement (REM) sleep was significantly suppressed, while changes in slow wave sleep were not significant. During the next day, all patients were in a significantly improved mood; however a rebound of REM sleep was observed in the second night after endotoxin administration and mood worsened again during the next days, indicating an only transient beneficial effect of the treatment. PMID:8573663

  15. Constitutive and cytokine-induced expression of human leukocyte antigens and cell adhesion molecules by human myotubes.

    PubMed Central

    Michaelis, D.; Goebels, N.; Hohlfeld, R.

    1993-01-01

    Understanding the immunobiology of muscle is relevant to muscular autoimmune diseases and to gene therapies based on myoblast transfer. We have investigated the constitutive and cytokine-induced intra- and extracellular expression of histocompatibility human leukocyte antigens (HLA) and cell adhesion molecules by multinucleated human myotubes using immunofluorescence microscopy. Myotubes constitutively expressed HLA class I but not HLA class II. Exposure to interferon-gamma, but not tumor necrosis factor-alpha, induced HLA-DR in the cytoplasm and on the surface membrane of approximately 40 to 95% of cultured myotubes. Surface expression was strongest in perinuclear membrane areas, and cytoplasmic expression was strongest at branching points and at the tips of myotubes. HLA-DP and HLA-DQ were not expressed in detectable amounts. Both interferon-gamma and tumor necrosis factor-alpha induced the intercellular adhesion molecule-1 (CD54) in the cytoplasm and on the surface of nearly all myotubes. The distribution of intercellular adhesion molecule-1 and HLA-DR was similar but not identical in double-positive myotubes. The leukocyte function-associated (LFA) adhesion molecules LFA-1 (CD11a/CD18), LFA-2 (CD2), and LFA-3 (CD58) could not be detected in the cytoplasm or on the surface. Our results indicate that cytokine-induced myotubes can participate in immune interactions with T lymphocytes. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 PMID:8214008

  16. The effect of the pro-inflammatory cytokine tumor necrosis factor-alpha on human joint capsule myofibroblasts

    PubMed Central

    2010-01-01

    Introduction Previous studies have shown that the number of myoblastically differentiated fibroblasts known as myofibroblasts (MFs) is significantly increased in stiff joint capsules, indicating their crucial role in the pathogenesis of post-traumatic joint stiffness. Although the mode of MFs' function has been well defined for different diseases associated with tissue fibrosis, the underlying mechanisms of their regulation in the pathogenesis of post-traumatic joint capsule contracture are largely unknown. Methods In this study, we examined the impact of the pro-inflammatory cytokine tumor necrosis factor-alpha (TNF-α) on cellular functions of human joint capsule MFs. MFs were challenged with different concentrations of TNF-α with or without both its specifically inactivating antibody infliximab (IFX) and cyclooxygenase-2 (COX2) inhibitor diclofenac. Cell proliferation, gene expression of both alpha-smooth muscle actin (α-SMA) and collagen type I, the synthesis of prostaglandin derivates E2, F1A, and F2A, as well as the ability to contract the extracellular matrix were assayed in monolayers and in a three-dimensional collagen gel contraction model. The α-SMA and COX2 protein expressions were evaluated by immunofluorescence staining and Western blot analysis. Results The results indicate that TNF-α promotes cell viability and proliferation of MFs, but significantly inhibits the contraction of the extracellular matrix in a dose-dependent manner. This effect was associated with downregulation of α-SMA and collagen type I by TNF-α application. Furthermore, we found a significant time-dependent upregulation of prostaglandin E2 synthesis upon TNF-α treatment. The effect of TNF-α on COX2-positive MFs could be specifically prevented by IFX and partially reduced by the COX2 inhibitor diclofenac. Conclusions Our results provide evidence that TNF-α specifically modulates the function of MFs through regulation of prostaglandin E2 synthesis and therefore may play a

  17. Cytokines and fever.

    PubMed

    Conti, Bruno; Tabarean, Iustin; Andrei, Cristina; Bartfai, Tamas

    2004-05-01

    Cytokines are highly inducible, secreted proteins mediating intercellular communication in the nervous and immune system. Fever is the multiphasic response of elevation and decline of the body core temperature regulated by central thermoregulatory mechanisms localized in the preoptic area of the hypothalamus. The discovery that several proinflammatory cytokines act as endogenous pyrogens and that other cytokines can act as antipyretic agents provided a link between the immune and the central nervous systems and stimulated the study of the central actions of cytokines. The proinflammatory cytokines interleukin 1 (IL-1), interleukin 6 (IL-6) and the tumor necrosis factor alpha (TNF) as well as the antiinflammatory cytokines interleukin 1 receptor antagonist (IL-1ra) and interleukin 10 (IL-10) have been most investigated for their pyrogenic or antipyretic action. The experimental evidence demonstrating the role of these secreted proteins in modulating the fever response is as follows: 1) association between cytokine levels in serum and CSF and fever; 2) finding of the presence of cytokine receptors on various cell types in the brain and demonstration of the effects of pharmacological application of cytokines and of their neutralizing antibodies on the fever response; 3) fever studies on cytokine- and cytokine receptor- transgenic models. Studies on the peripheral and the central action of cytokines demonstrated that peripheral cytokines can communicate with the brain in several ways including stimulation of afferent neuronal pathways and induction of the synthesis of a non cytokine pyrogen, i.e. PGE2, in endothelial cells in the periphery and in the brain. Cytokines synthesized in the periphery may act by crossing the blood brain barrier and acting directly via neuronal cytokine receptors. The mechanisms that ultimately mediate the central action of cytokines and of LPS on the temperature-sensitive neurons in the preoptic hypothalamic region involved in

  18. 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

    2016-08-01

    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. PMID:26441050

  19. IL-1β (interleukin-1β) stimulates the production and release of multiple cytokines and chemokines by human preadipocytes.

    PubMed

    Alomar, Suliman Y; Gentili, Alessandra; Zaibi, Mohamed S; Kępczyńska, Malgorzata A; Trayhurn, Paul

    2016-07-01

    The effect of IL-1β on cytokine and chemokine production by human preadipocytes has been examined. Preadipocytes were incubated with IL-1β, and cytokine and chemokine release was measured at 24 h by protein arrays, while the expression of cytokine/chemokine genes was assessed by qPCR at 4 and 24 h. IL-1β stimulated the secretion of multiple cytokines/chemokines, including IL-6, IL-8, IL-10, IL-13, MCP-4, TNFα and IP-10. IL-10 was not released by un-stimulated preadipocytes, while IL-6 exhibited the greatest response to IL-1β (453-fold increase). IL-16 and IL-12p40 did not respond to IL-1β. qPCR demonstrated that IL-1β markedly stimulated CCL3, CSF3 and CXCL10 expression at 4 h (>900-fold mRNA increase). A time-course indicated that while CCL13 (encoding MCP-4) exhibited minimal basal expression in preadipocytes, expression increased progressively following differentiation. Human preadipocytes are highly sensitive to IL-1β, the cytokine stimulating a major inflammatory response in these cells similar to that in mature adipocytes. PMID:26890442

  20. 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

  1. 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.

  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. Inhibition of zymosan-induced cytokine and chemokine expression in human corneal fibroblasts by triptolide

    PubMed Central

    Liu, Yang; Li, Jing; Liu, Ye; Wang, Ping; Jia, Hui

    2016-01-01

    AIM To investigate the effects of triptolide on proinflammatory cytokine and chemokine expression induced by the fungal component zymosan in cultured human corneal fibroblasts (HCFs). METHODS HCFs were cultured in the absence or presence of zymosan or triptolide. The release of interleukin (IL)-6, IL-8, and monocyte chemoattractant protein-1 (MCP-1) into culture supernatants was measured with enzyme-linked immunosorbent assays. The cellular abundance of the mRNAs for these proteins was determined by reverse transcription and real-time polymerase chain reaction analysis. The phosphorylation of mitogen-activated protein kinases (MAPKs) and the endogenous nuclear factor-κB (NF-κB) inhibitor IκB-α was examined by immunoblot analysis. The release of lactate dehydrogenase (LDH) activity from HCFs was measured with a colorimetric assay. RESULTS Triptolide inhibited the zymosan-induced release of IL-6, IL-8, and MCP-1 from HCFs in a concentration- and time-dependent manner. It also inhibited the zymosan-induced up-regulation of IL-6, IL-8, and MCP-1 mRNA abundance in these cells. Furthermore, triptolide attenuated zymosan-induced phosphorylation of the MAPKs extracellular signal-regulated kinase (ERK), c-Jun NH2-terminal kinase (JNK), and p38 as well as the phosphorylation and degradation of IκB-α. Triptolide did not exhibit cytotoxicity for HCFs. CONCLUSION Triptolide inhibited proinflammatory cytokine and chemokine production by HCFs exposed to zymosan, with this action likely being mediated by suppression of MAPK and NF-κB signaling pathways. This compound might thus be expected to limit the infiltration of inflammatory cells into the cornea associated with fungal infection. PMID:26949603

  4. Characteristics of human CD34+ cells exposed to ionizing radiation under cytokine-free conditions

    PubMed Central

    Ishikawa, Junya; Hayashi, Naoki; Yamaguchi, Masaru; Monzen, Satoru; Kashiwakura, Ikuo

    2015-01-01

    To clarify the mechanisms underlying radiation-induced hematopoietic stem cell death, we investigated the effects of excessive ionizing radiation on the clonogenic potential of CD34+ cells obtained from human umbilical cord blood under cytokine-free conditions. The CD34+ cells were X-ray–irradiated (up to 2 Gy) and were cultured for 0–48 h under cytokine-free conditions. At various time-points, the CD34+ cells were investigated for survival, clonogenic potential and the generation of mitochondrial superoxide. At 12 h after X-ray irradiation, the number of viable cells had decreased to ∼70–80% compared with the 0-h non-irradiated control, whereas the clonogenic potential in the X-ray–irradiated cells had decreased to ∼50%–60% compared with the 0-h non-irradiated control. Furthermore, significant generation of mitochondrial superoxide was observed at 6 h, and reached a maximum value between 12 and 24 h after X-ray irradiation. However, no significant differences were observed between non-irradiated and X-ray–irradiated cells in terms of the generation of reactive oxygen species or in the intracellular mitochondrial contents. In addition, a cDNA microarray analysis showed that the majority of the altered genes in the CD34+ cells at 6 h after X-ray irradiation were apoptosis-related genes. These results suggest the possibility that the elimination of the clonogenic potentials of CD34+ cells involves the generation of mitochondrial superoxide induced by ionizing radiation. PMID:25877692

  5. Smoke exposure of human macrophages reduces HDAC3 activity, resulting in enhanced inflammatory cytokine production.

    PubMed

    Winkler, Aaron R; Nocka, Karl N; Williams, Cara M M

    2012-08-01

    Chronic obstructive pulmonary disease (COPD) is a debilitating condition resulting from exposure to pollutants such as cigarette smoke. Pulmonary macrophages secrete a plethora of inflammatory mediators that are increased in the lungs of COPD patients, but whether this phenotype results directly from smoke exposure remains unknown. Using an in vitro model for alveolar macrophages (AM) derived from human peripheral blood monocytes with granulocyte-macrophage stimulating factor (GM-MØ), we analyzed the mechanistic connection between cigarette smoke exposure and histone deacetylase (HDAC) regulation, hypothesized to be a contributing factor in COPD pathophysiology. Here we show that acute smoke exposure inhibits HDAC enzymatic activity in GM-MØ. Analysis of mRNA and total cellular proteins for expression of class I (1, 2, 3 and 8), class II (4, 5, 6, 7, 9, 10), and class IV (11) HDAC revealed no effect of smoke exposure, whereas nuclear HDAC3 protein content was reduced. To better understand the physiological significance of reduced HDAC3 activity, we utilized siRNA to knockdown HDAC1, 2 and 3 individually. Interestingly, siRNA-mediated reduction of HDAC3 resulted in increased production of IL8 and IL1β in response to LPS stimulation, while HDAC2 knockdown had no effect on either cytokine. Lower nuclear content of HDAC3 in the context of equivalent total HDAC protein levels following smoke exposure may reflect increased nuclear export of HDAC3, allowing increased nuclear factor kappa b (NF-κB ) driven cytokine expression that can contribute to inflammation. PMID:22613758

  6. 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

  7. 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 (...

  8. A distinct array of proinflammatory cytokines is expressed in human colon epithelial cells in response to bacterial invasion.

    PubMed Central

    Jung, H C; Eckmann, L; Yang, S K; Panja, A; Fierer, J; Morzycka-Wroblewska, E; Kagnoff, M F

    1995-01-01

    Pathogenic bacteria that penetrate the intestinal epithelial barrier stimulate an inflammatory response in the adjacent intestinal mucosa. The present studies asked whether colon epithelial cells can provide signals that are important for the initiation and amplification of an acute mucosal inflammatory response. Infection of monolayers of human colon epithelial cell lines (T84, HT29, Caco-2) with invasive strains of bacteria (Salmonella dublin, Shigella dysenteriae, Yersinia enterocolitica, Listeria monocytogenes, enteroinvasive Escherichia coli) resulted in the coordinate expression and upregulation of a specific array of four proinflammatory cytokines, IL-8, monocyte chemotactic protein-1, GM-CSF, and TNF alpha, as assessed by mRNA levels and cytokine secretion. Expression of the same cytokines was upregulated after TNF alpha or IL-1 stimulation of these cells. In contrast, cytokine gene expression was not altered after infection of colon epithelial cells with noninvasive bacteria or the noninvasive protozoan parasite, G. lamblia. Notably, none of the cell lines expressed mRNA for IL-2, IL-4, IL-5, IL-6, IL-12p40, IFN-gamma, or significant levels of IL-1 or IL-10 in response to the identical stimuli. The coordinate expression of IL-8, MCP-1, GM-CSF and TNF alpha appears to be a general property of human colon epithelial cells since an identical array of cytokines, as well as IL-6, also was expressed by freshly isolated human colon epithelial cells. Since the cytokines expressed in response to bacterial invasion or other proinflammatory agonists have a well documented role in chemotaxis and activation of inflammatory cells, colon epithelial cells appear to be programmed to provide a set of signals for the activation of the mucosal inflammatory response in the earliest phases after microbial invasion. Images PMID:7814646

  9. In Vitro Effects of Propranolol on T Helper Type 1 Cytokine Profile in Human Leukemic T Cells

    PubMed Central

    Hajighasemi, Fatemeh; Mirshafiey, Abbas

    2016-01-01

    Introduction: Cytokines are a large group of proteins play a key role in inflammation. Down-regulation of pro-inflammatory cytokines has beneficial effect on heart function. Propranolol, as a non selective beta-adrenergic blocker, has been extensively used for treatment of many cardiovascular problems such as arrhythmias and heart malfunction. In addition anti-inflammatory effects of propranolol have been revealed. In this study the propranolol effect on T helper type 1 cytokine profile in human leukemic T cells has been assessed in vitro. Materials and methods: Human leukemic T cells (Molt-4 and Jurkat) were cultured in complete RPMI medium. The cells were then incubated with different concentrations of propranolol (0.03- 30 µM) in the presence or absence of PHA (10 µg/ml) for 48 hours. The supernatants of cell culture media were collected and used for cytokines assay. Results: Propranolol significantly decreased the T helper type 1 cytokine profile [Interleukin-2 (IL-2) and Interferon- γ (IFN-γ)] production in PHA stimulated Molt-4 and Jurkat cells, after 48 hour of incubation time, dose-dependently compared to untreated control cells. Conclusion: Our data showed a dose dependent inhibitory effect of propranolol on the IL-2 and IFN-γ production in human leukemic Molt-4 and Jurkat cells. The anti- inflammatory effect of propranolol reported by other investigators may be in part due to its suppressive effect on production of inflammatory cytokines such as IL-2 and IFN-γ. So, propranolol along with its chronic long-term usage in cardiovascular problems may have potential implication in treatment of inflammatory-based disorders. PMID:27252810

  10. 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

  11. 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.

  12. 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.

  13. Entamoeba lysyl-tRNA Synthetase Contains a Cytokine-Like Domain with Chemokine Activity towards Human Endothelial Cells

    PubMed Central

    Han, Jung Min; Kim, Sunghoon; Celada, Antonio; Ribas de Pouplana, Lluís

    2011-01-01

    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. PMID:22140588

  14. The Retinoic Acid Receptor-a Mediates Human T-Cell Activation and Th2 Cytokine Production

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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-g and TNF-a expression by activated human T cells and reducing the synthesis of IL-12p70 from accessory cells. Here, we have demonstrated ...

  15. 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.

  16. 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

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

    PubMed Central

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

    2015-01-01

    Objective: 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. Methods: 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. Results: 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. Conclusion: 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. PMID:26722438

  18. 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. PMID:26518119

  19. 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.

  20. 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.

  1. Pro-inflammatory cytokine levels in human apical periodontitis: Correlation with clinical and histological findings.

    PubMed

    Jakovljevic, Aleksandar; Knezevic, Aleksandra; Karalic, Danijela; Soldatovic, Ivan; Popovic, Branka; Milasin, Jelena; Andric, Miroslav

    2015-08-01

    This study aimed to compare the levels of tumour necrosis factor-alpha (TNF-α), interleukin-1 beta (IL-1β) and interleukin-6 (IL-6) between apical periodontitis lesions with different clinical and histological features. Based on clinical data and history of disease, 100 human apical periodontitis lesions were categorised as either asymptomatic or symptomatic lesions. According to histological examination, lesions were divided into periapical granulomas and radicular cysts. Pulp tissues of 25 impacted wisdom teeth were used as controls. Homogenised tissue samples were centrifuged and supernatants were used for the determination of cytokine levels by enzyme-linked immunosorbent assay. Significantly higher levels of IL-1β and IL-6 were found in symptomatic lesions compared with asymptomatic lesions and control tissues (P < 0.001, P < 0.001, respectively). The concentration of IL-1β was significantly higher in radicular cysts compared with periapical granulomas (P = 0.003). Symptomatic lesions, as judged by high local production of IL-1β and IL-6, represent an immunologically active stage of the disease. PMID:25163634

  2. 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

  3. Human Mesenchymal Stem Cells Modulate Inflammatory Cytokines after Spinal Cord Injury in Rat

    PubMed Central

    Machová Urdzíková, Lucia; Růžička, Jiří; LaBagnara, Michael; Kárová, Kristýna; Kubinová, Šárka; Jiráková, Klára; Murali, Raj; Syková, Eva; Jhanwar-Uniyal, Meena; Jendelová, Pavla

    2014-01-01

    Transplantation of mesenchymal stem cells (MSC) improves functional recovery in experimental models of spinal cord injury (SCI); however, the mechanisms underlying this effect are not completely understood. We investigated the effect of intrathecal implantation of human MSC on functional recovery, astrogliosis and levels of inflammatory cytokines in rats using balloon-induced spinal cord compression lesions. Transplanted cells did not survive at the lesion site of the spinal cord; however, functional recovery was enhanced in the MSC-treated group as was confirmed by the Basso, Beattie, and Bresnahan (BBB) and the flat beam test. Morphometric analysis showed a significantly higher amount of remaining white matter in the cranial part of the lesioned spinal cords. Immunohistochemical analysis of the lesions indicated the rearrangement of the glial scar in MSC-treated animals. Real-time PCR analysis revealed an increased expression of Irf5, Mrc1, Fgf2, Gap43 and Gfap. Transplantation of MSCs into a lesioned spinal cord reduced TNFα, IL-4, IL-1β, IL-2, IL-6 and IL-12 and increased the levels of MIP-1α and RANTES when compared to saline-treated controls. Intrathecal implantation of MSCs reduces the inflammatory reaction and apoptosis, improves functional recovery and modulates glial scar formation after SCI, regardless of cell survival. Therefore, repeated applications may prolong the beneficial effects induced by MSC application. PMID:24968269

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

    PubMed

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

    2014-04-22

    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

  5. 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. PMID:26090085

  6. Inborn errors of mucocutaneous immunity to Candida albicans in humans: a role for IL-17 cytokines?

    PubMed Central

    Puel, Anne; Picard, Capucine; Cypowyj, Sophie; Lilic, Desa; Abel, Laurent; Casanova, Jean-Laurent

    2013-01-01

    The various clinical manifestations of chronic mucocutaneous candidiasis (CMC) often result from acquired T-cell immunodeficiencies. More rarely, CMC results from inborn errors of immunity, the recent dissection of which has shed light on the molecular mechanisms of mucocutaneous immunity to Candida albicans. CMC may accompany various other infectious diseases in patients with almost any broad and profound T-cell primary immunodeficiency. By contrast, CMC is one of the few key infections in patients with autosomal dominant hyper IgE syndrome (mutations in STAT3), and in rare patients with autosomal recessive predisposition to mucocutaneous and invasive fungal infections (mutation in CARD9). In patients with mutations in STAT3 and CARD9 the development of IL-17-producing T cells is impaired. Moreover, CMC is the principal, if not only infection in patients with autosomal recessive autoimmune polyendocrinopathy syndrome-I (mutations in AIRE). Patients with this condition have high titers of neutralizing autoantibodies (auto-Abs) against the IL-17 cytokines IL-17A, IL-17F, and IL-22. Collectively, these data suggest that human IL-17A, IL-17F, and IL-22 are essential for mucocutaneous immunity to Candida albicans. They also suggest that the distinct syndrome of isolated CMC, without autoimmunity or other infections, may be caused by inborn errors of IL-17 immunity. PMID:20674321

  7. 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

  8. Effects of pro-inflammatory cytokines, lipopolysaccharide and COX-2 mediators on human colonic neuromuscular function and epithelial permeability.

    PubMed

    Safdari, B K; Sia, T C; Wattchow, D A; Smid, S D

    2016-07-01

    Chronic colitis is associated with decreased colonic muscle contraction and loss of mucosal barrier function. Pro-inflammatory cytokines and bacterial lipopolysaccharide (LPS) are important in the generation and maintenance of inflammation. While colitis is associated with upregulated COX-2 -derived prostanoids and nitric oxide (NO), the direct activity of pro-inflammatory cytokines on human colonic neuromuscular function is less clear. This study investigated the effects of IBD-associated pro-inflammatory cytokines IL-17, TNF-α, IL-1β and LPS on human colonic muscle strip contractility, alone and following inhibition of COX-2 or nitric oxide production. In addition, human colonic epithelial Caco-2 cell monolayers were treated with LPS or COX-2 mediators including prostaglandins (PGE2, PGF2α) or their corresponding ethanolamides (PGE2-EA or PGF2α-EA) over 48h and trans-epithelial electrical resistance used to record permeability changes. Longitudinal muscle strips were obtained from healthy colonic resection margins and mounted in organ baths following IL-17, TNF-α, IL-1β and bacterial LPS incubations in an explant setting over 20h. Contraction in response to acetylcholine (ACh) was then measured, before and after either COX-2 inhibition (nimesulide; 10(-5)M) or nitric oxide synthase (NOS) inhibition (l-NNA; 10(-4)M). None of the cytokine or LPS explant incubations affected the potency or maximum cholinergic contraction in vitro, and subsequent COX-2 blockade with nimesulide revealed a significant but similar decrease in potency of ACh-evoked contraction in control, LPS and cytokine-incubated muscle strips. Pre-treatment with l-NNA provided no functional differences in the potency or maximum contractile responses to ACh in cytokine or LPS-incubated colonic longitudinal smooth muscle. Only PGE2 transiently increased Caco-2 monolayer permeability at 24h, while LPS (10μg/ml) increased permeability over 24-48h. These findings indicate that cholinergic

  9. An ethyl acetate fraction of Moringa oleifera Lam. Inhibits human macrophage cytokine production induced by cigarette smoke.

    PubMed

    Kooltheat, Nateelak; Sranujit, Rungnapa Pankla; 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

  10. MUTZ-3, a human cell line model for the cytokine-induced differentiation of dendritic cells from CD34+ precursors.

    PubMed

    Masterson, Allan J; Sombroek, Claudia C; De Gruijl, Tanja D; Graus, Yvo M F; van der Vliet, Hans J J; Lougheed, Sinéad M; van den Eertwegh, Alfons J M; Pinedo, Herbert M; Scheper, Rik J

    2002-07-15

    Many human myeloid leukemia-derived cell lines possess the ability to acquire a dendritic cell (DC) phenotype. However, cytokine responsiveness is generally poor, requiring direct manipulation of intracellular signaling mechanisms for differentiation. In contrast, the CD34+ human acute myeloid leukemia cell line MUTZ-3 responds to granulocyte macrophage- colony-stimulating factor (GM-CSF), interleukin 4 (IL-4), and tumor necrosis factor alpha (TNFalpha), cytokines known to be pivotal both in vivo and in vitro for DC generation from monocytes and CD34+ stem cells. In all respects, MUTZ-3 cells behave as the immortalized equivalent of CD34+ DC precursors. Upon stimulation with specific cytokine cocktails, they acquire a phenotype consistent with either interstitial- or Langerhans-like DCs and upon maturation (mDC), express CD83. MUTZ-3 DC display the full range of functional antigen processing and presentation pathways. These findings demonstrate the unique suitability of MUTZ-3 cells as an unlimited source of CD34+ DC progenitors for the study of cytokine-induced DC differentiation. PMID:12091369

  11. 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

  12. The cytomegalovirus homolog of interleukin-10 requires phosphatidylinositol 3-kinase activity for inhibition of cytokine synthesis in monocytes.

    PubMed

    Spencer, Juliet V

    2007-02-01

    Human cytomegalovirus (CMV) has evolved numerous strategies for evading host immune defenses, including piracy of cellular cytokines. A viral homolog of interleukin-10, designated cmvIL-10, binds to the cellular IL-10 receptor and effects potent immune suppression. The signaling pathways employed by cmvIL-10 were investigated, and the classic IL-10R/JAK1/Stat3 pathway was found to be activated in monocytes. However, inhibition of JAK1 had little effect on cmvIL-10-mediated suppression of tumor necrosis factor alpha (TNF-alpha) production. Inhibition of the phosphatidylinositol 3-kinase/Akt pathway had a more significant impact on TNF-alpha levels but did not completely relieve the immune suppression, demonstrating that cmvIL-10 stimulates multiple signaling pathways to modulate cell function. PMID:17121792

  13. Unstimulated human CD4 lymphocytes express a cytoplasmic immature form of the common cytokine receptor gamma-chain.

    PubMed

    Bani, L; Pasquier, V; Kryworuchko, M; Salamero, J; Thèze, J

    2001-07-01

    As a component of various cytokine receptors, common cytokine receptor gamma-chain (gamma(c)) is essential in the development of the immune system and plays an important role in different stages of inflammatory and immune responses. Here we establish that resting CD4 T cells and the Jurkat CD4 T cell line do not express the mature form of gamma(c) (64 kDa) recognized by mAb Tugh4. However, these cells constitutively transcribe the corresponding gamma(c) gene. This apparent paradox was solved by the demonstration that polyclonal anti-gamma(c) Abs detected endoglycosidase-H-sensitive immature forms of gamma(c) (54-58 kDa) expressed by quiescent CD4 T lymphocytes and Jurkat cells. Immature gamma(c) is characterized as an intracellular component localized in the endoplasmic reticulum. Pulse-chase analysis shows that the immature gamma(c) is rapidly degraded after synthesis. After activation of CD4 T lymphocytes, and as seen in the CD4 T cell line Kit 225, the endoglycosidase-H-resistant mature form of gamma(c) is detectable at the cell surface and in the endosomal compartment. For the first time, our results demonstrate that a cytokine receptor chain may be constitutively produced as an immature form. Furthermore, this supports the notion that expression of the functional form of gamma(c) may require intracellular interactions with lineage- or subset-specific molecular partners. PMID:11418669

  14. 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

  15. 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. PMID:19665552

  16. 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

  17. Multiplex Analysis of Serum Cytokines in Humans with Hantavirus Pulmonary Syndrome

    PubMed Central

    Morzunov, Sergey P.; Khaiboullina, Svetlana F.; St. Jeor, Stephen; Rizvanov, Albert A.; Lombardi, Vincent C.

    2015-01-01

    Hantavirus pulmonary syndrome (HPS) is an acute zoonotic disease transmitted primarily through inhalation of virus-contaminated aerosols. Hantavirus infection of endothelial cells leads to increased vascular permeability without a visible cytopathic effect. For this reason, it has been suggested that the pathogenesis of HPS is indirect with immune responses, such as cytokine production, playing a dominant role. In order to investigate their potential contribution to HPS pathogenesis, we analyzed the serum of hantavirus-infected subjects and healthy controls for 68 different cytokines, chemokines, angiogenic, and growth factors. Our analysis identified differential expression of cytokines that promote tissue migration of mononuclear cells including T lymphocytes, natural killer cells, and dendritic cells. Additionally, we observed a significant upregulation of cytokines known to regulate leukocyte migration and subsequent repair of lung tissue, as well as cytokines known to increase endothelial monolayer permeability and facilitate leukocyte transendothelial migration. Conversely, we observed a downregulation of cytokines associated with platelet numbers and function, consistent with the thrombocytopenia observed in subjects with HPS. This study corroborates clinical findings and extends our current knowledge regarding immunological and laboratory findings in subjects with HPS. PMID:26379668

  18. Multiplex Analysis of Serum Cytokines in Humans with Hantavirus Pulmonary Syndrome.

    PubMed

    Morzunov, Sergey P; Khaiboullina, Svetlana F; St Jeor, Stephen; Rizvanov, Albert A; Lombardi, Vincent C

    2015-01-01

    Hantavirus pulmonary syndrome (HPS) is an acute zoonotic disease transmitted primarily through inhalation of virus-contaminated aerosols. Hantavirus infection of endothelial cells leads to increased vascular permeability without a visible cytopathic effect. For this reason, it has been suggested that the pathogenesis of HPS is indirect with immune responses, such as cytokine production, playing a dominant role. In order to investigate their potential contribution to HPS pathogenesis, we analyzed the serum of hantavirus-infected subjects and healthy controls for 68 different cytokines, chemokines, angiogenic, and growth factors. Our analysis identified differential expression of cytokines that promote tissue migration of mononuclear cells including T lymphocytes, natural killer cells, and dendritic cells. Additionally, we observed a significant upregulation of cytokines known to regulate leukocyte migration and subsequent repair of lung tissue, as well as cytokines known to increase endothelial monolayer permeability and facilitate leukocyte transendothelial migration. Conversely, we observed a downregulation of cytokines associated with platelet numbers and function, consistent with the thrombocytopenia observed in subjects with HPS. This study corroborates clinical findings and extends our current knowledge regarding immunological and laboratory findings in subjects with HPS. PMID:26379668

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

    PubMed Central

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

    2016-01-01

    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 concomitant

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

    PubMed

    Khalfaoui, Soumaya; Eichhorn, Vivien; 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

  1. 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

  2. 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

  3. Modulation of the plasminogen activation system by inflammatory cytokines in human colon carcinoma cells.

    PubMed Central

    Trân-Thang, C.; Kruithof, E.; Lahm, H.; Schuster, W. A.; Tada, M.; Sordat, B.

    1996-01-01

    Inflammation may promote malignant invasion by enhancing cancer cell-associated proteolysis. Here we present the effect of inflammatory cytokines on the plasminogen activation system of eight human colon carcinoma cell lines. Tumour necrosis factor alpha (TNF-alpha) and interleukin-1 beta (IL-1 beta) increased in several, but not all, cell lines the production of urokinase-type plasminogen activator (uPA), tissue-type PA (tPA) and plasminogen activator inhibitor type 1 (PAI-1) as analysed by zymography, enzyme immunoassays and Northern analysis. Interleukin 6 (IL-6) had no effect. uPA receptor (uPAR) mRNA levels were also upregulated. However, each individual cell line responded differently following exposure to TNF-alpha or IL-1 beta. For example, there was a dose-dependent up-regulation of uPA and PAI-1 in SW 620 cells, whereas increased uPA production in SW 1116 cells was not accompanied by an increase in PAI-1. The TNF-alpha stimulatory effect was blocked by anti-TNF-alpha Fab fragments. All cell lines expressed both types of TNF receptor mRNAs, whereas no transcript for TNF-alpha, IL-1 beta, IL-6, IL-6 receptor or the IL-1 receptors was found. Our results demonstrate that TNF-alpha and IL-1 beta stimulate the plasminogen activation system in tumour cell but the responses differed even in cells derived from the same tissue origin. Images Figure 1 Figure 2 Figure 3 Figure 5 PMID:8826848

  4. Localized expression of mRNA for phagocyte-specific chemotactic cytokines in human periodontal infections.

    PubMed Central

    Tonetti, M S; Imboden, M A; Gerber, L; Lang, N P; Laissue, J; Mueller, C

    1994-01-01

    In bacterial infections, mononuclear and polymorphonuclear phagocytes are key components of host defenses. Recent investigations have indicated that chemokines are able to recruit and activate phagocytes. In particular, interleukin-8 (IL-8) attracts polymorphonuclear leukocytes (PMNs), while monocyte chemoattractant protein-1 (MCP-1) is selective for cells of the monocyte/macrophage lineage. In this investigation, we analyzed the in situ expression of IL-8 and MCP-1 mRNAs in human periodontal infections. Specific mRNA was detected by in situ hybridization using 35S-labeled riboprobes in frozen tissue sections. Phagocytes (PMNs and macrophages) were specifically detected as elastase-positive or CD68+ cells by a three-stage immunoperoxidase technique. Results indicated that expression of phagocyte-specific cytokines was confined to selected tissue locations and, in general, paralleled phagocyte infiltration. In particular, IL-8 expression was maximal in the junctional epithelium adjacent to the infecting microorganisms; PMN infiltration was more prominent in the same area. MCP-1 was expressed in the chronic inflammatory infiltrate and along the basal layer of the oral epithelium. Cells of the monocyte/macrophage lineage were demonstrated to be present in the same areas. The observed expression pattern may be the most economic way to establish a cell-type-selective chemotactic gradient within the tissue that is able to effectively direct polymorphonuclear phagocyte migration toward the infecting microorganisms and modulate mononuclear phagocyte infiltration in the surrounding tissues. This process may optimize host defenses and contribute to containing leukocyte infiltration to the infected and inflamed area, thus limiting tissue damage. Images PMID:8063420

  5. 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

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

    PubMed

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

    2016-05-01

    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

  7. Signal Strength and Metabolic Requirements Control Cytokine-Induced Th17 Differentiation of Uncommitted Human T Cells.

    PubMed

    Kastirr, Ilko; Crosti, Mariacristina; Maglie, Stefano; Paroni, Moira; Steckel, Bodo; Moro, Monica; Pagani, Massimilliano; Abrignani, Sergio; Geginat, Jens

    2015-10-15

    IL-17 production defines Th17 cells, which orchestrate immune responses and autoimmune diseases. Human Th17 cells can be efficiently generated with appropriate cytokines from precommitted precursors, but the requirements of uncommitted T cells are still ill defined. In standard human Th17 cultures, IL-17 production was restricted to CCR6(+)CD45RA(+) T cells, which expressed CD95 and produced IL-17 ex vivo, identifying them as Th17 memory stem cells. Uncommitted naive CD4(+) T cells upregulated CCR6, RORC2, and IL-23R expression with Th17-promoting cytokines but in addition required sustained TCR stimulation, late mammalian target of rapamycin (mTOR) activity, and HIF-1α to produce IL-17. However, in standard high-density cultures, nutrients like glucose and amino acids became progressively limiting, and mTOR activity was consequently not sustained, despite ongoing TCR stimulation and T cell proliferation. Sustained, nutrient-dependent mTOR activity also induced spontaneous IL-22 and IFN-γ production, but these cytokines had also unique metabolic requirements. Thus, glucose promoted IL-12-independent Th1 differentiation, whereas aromatic amino acid-derived AHR ligands were selectively required for IL-22 production. The identification of Th17 memory stem cells and the stimulation requirements for induced human Th17/22 differentiation have important implications for T cell biology and for therapies targeting the mTOR pathway. PMID:26378072

  8. [CYTOKINES DURING THE HUMAN IMMUNODEFICIENCY VIRUS INFECTION TYPE 1(HIV-1)].

    PubMed

    Selimova, L M; Kalnina, L B; Serebrovskaya, L V; Ivanova, L A; Gulyaeva, A N; Nosik, D N

    2016-01-01

    In this work the proinflammatory (IL-1β, IFN-γ, TNF-α, IL-2) and anti-inflammatory (IL-4, IL-10) plasma cytokine levels were evaluated in HIV-infected patients with or without antiretroviral treatment (ART). IFN-γ was detected in 94% samples with and without ART, TNF-α in 88% and IL-2 in 38% samples without ART, as well as in 12% and 30% samples with ART, respectively. Positive correlation was detected between viral RNA and IFN-γ levels (rs = 0.13) and negative correlation (rs = -0.242) in the patients without or with ART. Cosecretion of three cytokines (IFN-γ, TNF-α, IL-2) was detected in 31% samples and two cytokines (IFN-γ, TNF-α) in 35% samples of persons without ART. Cosecretion of three cytokines (IFN-γ, TNF-α, IL-2) was detected in 20% samples with ART; cosecretion of IFN-γ and IL-2 was detected in 10% samples. The higher percentage of the proinflammatory cytokines with cosecretion was detected in plasma HIV-infected patients without ART in the course of 6 and more years, which suggests that their immune system is able to provide disease control. PMID:27145600

  9. 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

  10. Potential regulatory molecules in the human trabecular meshwork of patients with glaucoma: immunohistochemical profile of a number of inflammatory cytokines.

    PubMed

    Taurone, Samanta; Ripandelli, Guido; Pacella, Elena; Bianchi, Enrica; Plateroti, Andrea Maria; De Vito, Stefania; Plateroti, Pasquale; Grippaudo, Francesca Romana; Cavallotti, Carlo; Artico, Marco

    2015-02-01

    Glaucoma occurs when there are imbalances between the production and the drainage of the eye liquid. The vast majority of the aqueous humor leaves the eye through the trabecular meshwork (TM). The cause of hypertonicity may be due to an alteration in the thickness of the TM. In the majority of cases the molecular changes that determine primary open‑angle glaucoma (POAG) are unclear. However, it has been hypothesized that the significant increase in the extracellular matrix (ECM) of the fibrillary bands in the TM is associated with possible inflammatory conditions. In this study the tissue distribution of interleukin (IL)‑6, IL‑1β, transforming growth factor-β1 (TGF‑β1), vascular endothelial growth factor (VEGF) and tumor necrosis factor α (TNF‑α) was analyzed in TM samples from patients with POAG by immunohistochemistry. Seven specimens from patients with POAG and three control tissues were analyzed by immunohistochemistry using specific antibodies against these cytokines. Morphological changes in the TM, such as increased cell content, macrophages, fibrosis and accumulation of neutrophils, were observed by transmission electron microscopy. In human TM tissues, an evident immunoreactivity for IL‑6, IL‑1β and TNF‑α was observed in patients with POAG when compared with the control subjects, indicating that these cytokines may be correlated with disease activity. TM endothelial cells secrete a number of factors and cytokines that modulate the functions of the cells and the ECM of the conventional outflow pathway. In the TM in glaucoma, macrophages produce cytokines, including IL‑6, IL‑1β and TNF‑α, leading to an acute inflammatory response and recruitment of other immune cells, including T lymphocytes. In addition, TGF‑β1 regulates and induces the expression of IL‑6 in TM that indirectly induces angiogenesis by stimulating VEGF expression. The present results support previous evidence that suggests that growth factors and cytokines

  11. T Helper 1 and T Helper 2 Cytokines Differentially Modulate Expression of Filaggrin and its Processing Proteases in Human Keratinocytes

    PubMed Central

    Di, Zheng-Hong; Ma, Lei; Qi, Rui-Qun; Sun, Xiao-Dong; Huo, Wei; Zhang, Li; Lyu, Ya-Ni; Hong, Yu-Xiao; Chen, Hong-Duo; Gao, Xing-Hua

    2016-01-01

    Background: Atopic dermatitis (AD) is characterized by defective skin barrier and imbalance in T helper 1/T helper 2 (Th1/Th2) cytokine expression. Filaggrin (FLG) is the key protein to maintaining skin barrier function. Recent studies indicated that Th1/Th2 cytokines influence FLG expression in keratinocytes. However, the role of Th1/Th2 cytokines on FLG processing is not substantially documented. Our aim was to investigate the impact of Th1/Th2 cytokines on FLG processing. Methods: HaCaT cells and normal human keratinocytes were cultured in low and high calcium media and stimulated by either interleukin (IL)-4, 13 or interferon-γ (IFN-γ). FLG, its major processing proteases and key protease inhibitor lymphoepithelial Kazal-type-related inhibitor (LEKTI) were measured by both real-time quantitative polymerase chain reaction and Western blotting. Their expression was also evaluated in acute and chronic AD lesions by immunohistochemistry. Results: IL-4/13 significantly reduced, while IFN-γ significantly up-regulated FLG expression. IL-4/13 significantly increased, whereas IFN-γ significantly decreased the expression of kallikreins 5 and 7, matriptase and channel-activating serine protease 1. On the contrary, IL-4/13 significantly decreased, while IFN-γ increased the expression of LEKTI and caspase-14. Similar trends were observed in AD lesions. Conclusions: Our results suggested that Th1/Th2 cytokines differentially regulated the expression of major FLG processing enzymes. The imbalance between Th1 and Th2 polarized immune response seems to extend to FLG homeostasis, through the network of FLG processing enzymes. PMID:26831231

  12. Reconfigurable microfluidic device with integrated antibody arrays for capture, multiplexed stimulation, and cytokine profiling of human monocytes.

    PubMed

    Vu, Tam; Rahimian, Ali; Stybayeva, Gulnaz; Gao, Yandong; Kwa, Timothy; Van de Water, Judy; Revzin, Alexander

    2015-07-01

    Monocytes represent a class of immune cells that play a key role in the innate and adaptive immune response against infections. One mechanism employed by monocytes for sensing foreign antigens is via toll-like receptors (TLRs)-transmembrane proteins that distinguish classes of foreign pathogens, for example, bacteria (TLR4, 5, and 9) vs. fungi (TLR2) vs. viruses (TLR3, 7, and 8). Binding of antigens activates a signaling cascade through TLR receptors that culminate in secretion of inflammatory cytokines. Detection of these cytokines can provide valuable clinical data for drug developers and disease investigations, but this usually requires a large sample volume and can be technically inefficient with traditional techniques such as flow cytometry, enzyme-linked immunosorbent assay, or luminex. This paper describes an approach whereby antibody arrays for capturing cells and secreted cytokines are encapsulated within a microfluidic device that can be reconfigured to operate in serial or parallel mode. In serial mode, the device represents one long channel that may be perfused with a small volume of minimally processed blood. Once monocytes are captured onto antibody spots imprinted into the floor of the device, the straight channel is reconfigured to form nine individually perfusable chambers. To prove this concept, the microfluidic platform was used to capture monocytes from minimally processed human blood in serial mode and then to stimulate monocytes with different TLR agonists in parallel mode. Three cytokines, tumor necrosis factor-α, interleukin (IL)-6, and IL-10, were detected using anti-cytokine antibody arrays integrated into each of the six chambers. We foresee further use of this device in applications such as pediatric immunology or drug/vaccine testing where it is important to balance small sample volume with the need for high information content. PMID:26339315

  13. A Cytokine-Independent Approach To Identify Antigen-Specific Human Germinal Center T Follicular Helper Cells and Rare Antigen-Specific CD4+ T Cells in Blood.

    PubMed

    Dan, Jennifer M; Lindestam Arlehamn, Cecilia S; Weiskopf, Daniela; da Silva Antunes, Ricardo; Havenar-Daughton, Colin; Reiss, Samantha M; Brigger, Matthew; Bothwell, Marcella; Sette, Alessandro; Crotty, Shane

    2016-08-01

    Detection of Ag-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 Ag-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 from Th1 or Th17 cells in the quantities of cytokines produced. Conventional identification of Ag-specific cells by intracellular cytokine staining 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 Ag-specific GC Tfh cells in human lymphoid tissue. Whereas Group A Streptococcus-specific GC Tfh cells produced minimal detectable cytokines by intracellular cytokine staining, the AIM method identified 85-fold more Ag-specific GC Tfh cells. Intriguingly, these GC Tfh cells consistently expressed programmed death ligand 1 upon activation. AIM also detected non-Tfh cells in lymphoid tissue. As such, we applied AIM for identification of rare Ag-specific CD4(+) T cells in human peripheral blood. Dengue, tuberculosis, and pertussis vaccine-specific CD4(+) T cells were readily detectable by AIM. In summary, cytokine assays missed 98% of Ag-specific human GC Tfh cells, reflecting the biology of these cells, which could instead be sensitively identified by coexpression of TCR-dependent activation markers. PMID:27342848

  14. 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. PMID:26679997

  15. 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

  16. Proinflammatory Cytokine Gene Induction by Human T-Cell Leukemia Virus Type 1 (HTLV-1) and HTLV-2 Tax in Primary Human Glial Cells▿

    PubMed Central

    Banerjee, Prabal; Rochford, Rosemary; Antel, J.; Canute, G.; Wrzesinski, Stephen; Sieburg, Michelle; Feuer, Gerold

    2007-01-01

    Infection with human T-cell leukemia virus type 1 (HTLV-1) can result in the development of HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP), a chronic inflammatory disease of the central nervous system (CNS). HTLV-2 is highly related to HTLV-1 at the genetic level and shares a high degree of sequence homology, but infection with HTLV-2 is relatively nonpathogenic compared to HTLV-1. Although the pathogenesis of HAM/TSP remains to be fully elucidated, previous evidence suggests that elevated levels of the proinflammatory cytokines in the CNS are associated with neuropathogenesis. We demonstrate that HTLV-1 infection in astrogliomas results in a robust induction of interleukin-1β (IL-1β), IL-1α, tumor necrosis factor alpha (TNF-α), TNF-β, and IL-6 expression. HTLV encodes for a viral transcriptional transactivator protein named Tax that also induces the transcription of cellular genes. To investigate and compare the effects of Tax1 and Tax2 expression on the dysregulation of proinflammatory cytokines, lentivirus vectors were used to transduce primary human astrocytomas and oligodendrogliomas. The expression of Tax1 in primary human astrocytomas and oligodendrogliomas resulted in significantly higher levels of proinflammatory cytokine gene expression compared to Tax2. Notably, Tax1 expression uniquely sensitized primary human astrocytomas to apoptosis. A Tax2/Tax1 chimera encoding the C-terminal 53 amino acids of the Tax1 fused to the Tax2 gene (Tax221) demonstrated a phenotype that resembled Tax1, with respect to proinflammatory cytokine gene expression and sensitization to apoptosis. The patterns of differential cytokine induction and sensitization to apoptosis displayed by Tax1 and Tax2 may reflect differences relating to the heightened neuropathogenicity associated with HTLV-1 infection and the development of HAM/TSP. PMID:17121800

  17. 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.

  18. Time dependent production of cytokines and eicosanoids by human monocytic leukaemia U937 cells; effects of glucocorticosteroids.

    PubMed Central

    Garrelds, I M; van Hal, P T; Haakmat, R C; Hoogsteden, H C; Saxena, P R; Zijlstra, F J

    1999-01-01

    In the present study the human monoblast cell line U937 has been used as a model to study the function of human mononuclear phagocytes in asthma. The kinetics of the production of eicosanoids and cytokines, which are thought to play a role in the pathogenesis of asthma, were studied. In addition, the effects of glucocorticosteroids were investigated, as these drugs are of great importance for the treatment of asthmatic patients. After stimulation with phorbol-12 myristate acetate (PMA) for 24 h, U937 cells were cultured in the absence or presence of lipopolysaccharide (LPS: 1 and 5 microg ml(-1)) and glucocorticosteroids (budesonide, fluticasone propionate and prednisolone: 10(-11), 10(-9) and 10(-7) M) for 96 h. The production of interleukin-1beta (IL-1beta), interleukin-6 (IL-6), prostaglandin E2 (PGE2) and thromboxane B2 (TxB2) gradually increased in time after stimulation with LPS, whereas the transient production of tumor necrosis factor alpha (TNF-alpha) reached its maximum between 6 and 12 h. Interferon-gamma (IFN-gamma), interleukin-10 (IL-10) and leukotriene B4 (LTB4) were not detectable. All three glucocorticosteroids (budesonide, fluticasone propionate and prednisolone) completely inhibited the production of both eicosanoids and cytokines. The production of eicosanoids was more sensitive to these glucocorticoids than the production of cytokines. The observed differences in the kinetics of the production of eicosanoids and cytokines stress the importance of time course experiments in studies on the effect of drugs on mononuclear cells. PMID:10704077

  19. 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

  20. 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. PMID:26644088

  1. Proteomic and cytokine plasma biomarkers for predicting progression from colorectal adenoma to carcinoma in human patients.

    PubMed

    Choi, Jung-Won; Liu, Hao; Shin, Dong Hoon; Yu, Gyeong Im; Hwang, Jae Seok; Kim, Eun Soo; Yun, Jong Won

    2013-08-01

    In the present study, we screened proteomic and cytokine biomarkers between patients with adenomatous polyps and colorectal cancer (CRC) in order to improve our understanding of the molecular mechanisms behind turmorigenesis and tumor progression in CRC. To this end, we performed comparative proteomic analysis of plasma proteins using a combination of 2DE and MS as well as profiled differentially regulated cytokines and chemokines by multiplex bead analysis. Proteomic analysis identified 11 upregulated and 13 downregulated plasma proteins showing significantly different regulation patterns with diagnostic potential for predicting progression from adenoma to carcinoma. Some of these proteins have not previously been implicated in CRC, including upregulated leucine-rich α-2-glycoprotein, hemoglobin subunit β, Ig α-2 chain C region, and complement factor B as well as downregulated afamin, zinc-α-2-glycoprotein, vitronectin, and α-1-antichymotrypsin. In addition, plasma levels of three cytokines/chemokines, including interleukin-8, interferon gamma-induced protein 10, and tumor necrosis factor α, were remarkably elevated in patients with CRC compared to those with adenomatous polyps. Although further clinical validation is required, these proteins and cytokines can be established as novel biomarkers for CRC and/or its progression from colon adenoma. PMID:23606366

  2. Cross-reactivity of anti-human cytokine monoclonal antibodies used as a tool to identify novel immunological biomarkers in domestic ruminants.

    PubMed

    Dorneles, E M S; Araújo, M S S; Teixeira-Carvalho, A; Martins-Filho, O A; Lage, A P

    2015-01-01

    Eleven commercially available PE-labeled anti-human (IL-1-α, IL-6, IL-8, TNF-α, IL-17A, IL-5, IL-10, IL-12 and IL-13) and anti-mouse (IL-10, TNF-α) cytokine monoclonal antibodies (mAbs) were tested for cross-reactivity with cattle, goat, and sheep cytokines. Cross-reactivity was assessed by comparative analysis with the standard reactivity of the target species. Our data demonstrated that anti-human IL-1-α, IL-6, IL-8, IL-17A and IL-10 mAbs cross-react with all ruminant species tested. Anti-human IL-5 mAb showed a strong cross-reactivity with cattle and goat IL-5, while anti-human TNF-α mAb showed a selective cross-reactivity with goat TNF-α. No cross-reactivity with the ruminant cytokines was observed for anti-human IL-12 and IL-13 mAbs or for the two anti-mouse cytokine mAbs tested. The present study demonstrated the cross-reactivity of various anti-human cytokine mAbs with cattle, sheep, and goat cytokines, increasing the range of immunological biomarkers for studies in veterinary medicine. PMID:25730032

  3. 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

  4. Sterol synthesis in the human arterial intima

    PubMed Central

    Chobanian, Aram V.

    1968-01-01

    Intimal sterol synthesis was examined in isolated human arterial segments obtained at surgery or at postmortem examination. The tissues were incubated with acetate-1-14C and mevalonate-2-14C and the incorporation of these precursors into sterols was determined. Intimal sterols were isolated by multiple chromatographic techniques and purified by bromination and oxidation procedures. The results indicate that the arterial intima can incorporate acetate and mevalonate into cholesterol, cholestanol, and squalene. Cholestanol was the major sterol synthesized by the arterial wall, but cholesterol production was also consistently observed. The findings suggest that local synthesis is a potential source of sterol accumulation within the arterial wall. The conversion of cholesterol to other sterols was also studied in terminally ill patients receiving labeled cholesterol before death. Tissue analyses revealed the presence of labeled cholestanol as well as cholesterol in the tissue 5-104 days after labeled cholesterol administration. The results demonstrate the conversion of cholesterol to cholestanol in man and suggest that the exchange of cholestanol between the blood and tissues is similar to that of cholesterol. PMID:5637146

  5. 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.

  6. Gene deleted live attenuated Leishmania vaccine candidates against visceral leishmaniasis elicit pro-inflammatory cytokines response in human PBMCs.

    PubMed

    Avishek, Kumar; Kaushal, Himanshu; Gannavaram, Sreenivas; Dey, Ranadhir; Selvapandiyan, Angamuthu; Ramesh, V; Negi, Narender Singh; Dubey, Uma S; Nakhasi, Hira L; Salotra, Poonam

    2016-01-01

    Currently no effective vaccine is available for human visceral leishmaniasis(VL) caused by Leishmania donovani. Previously, we showed that centrin1 and p27gene deleted live attenuated Leishmania parasites (LdCen1(-/-) and Ldp27(-/-)) are safe, immunogenic and protective in animal models. Here, to assess the correlates of protection, we evaluated immune responses induced by LdCen1(-/-) and Ldp27(-/-) in human blood samples obtained from healthy, healed VL (HVL), post kala-azar dermal leishmaniasis(PKDL) and VL subjects. Both parasites infected human macrophages, as effectively as the wild type parasites. Further, LdCen1(-/-) and Ldp27(-/-) strongly stimulated production of pro-inflammatory cytokines including, IL-12, IFN-γ, TNF-α, IL-2, IL-6 and IL-17 in the PBMCs obtained from individuals with a prior exposure to Leishmania (HVL and PKDL). There was no significant stimulation of anti-inflammatory cytokines (IL-4 and IL-10). Induction of Th1 biased immune responses was supported by a remarkable increase in IFN-γ secreting CD4(+) and CD8(+) T cells and IL-17 secreting CD4(+) cells in PBMCs from HVL cases with no increase in IL-10 secreting T cells. Hence, LdCen1(-/-) and Ldp27(-/-) are promising as live vaccine candidates against VL since they elicit strong protective immune response in human PBMCs from HVL, similar to the wild type parasite infection, mimicking a naturally acquired protection following cure. PMID:27624408

  7. Synthesis of a novel cytokine and its gene (LD78) expressions in hematopoietic fresh tumor cells and cell lines.

    PubMed Central

    Yamamura, Y; Hattori, T; Obaru, K; Sakai, K; Asou, N; Takatsuki, K; Ohmoto, Y; Nomiyama, H; Shimada, K

    1989-01-01

    The gene coding for the protein LD78 was isolated from stimulated human tonsillar lymphocytes by differential hybridization. The gene product consisted of 92 amino acids with characteristics of cytokines. LD78 gene transcripts were detected in eight of eight fresh samples of cells from patients with acute nonlymphocytic leukemia (ANLL) by Northern blot analysis. ANLL cells with monocytic features gave the strongest bands. RNA transcripts were found in two of three samples of cells from patients with adult T cell leukemia (ATL), eight of nine samples from patients with acute lymphocytic leukemia (ALL) of B cell lineage, and one of the three samples from patients with T cell ALL. KG-1, HL-60, HUT 102, MT-2, and MJ cell lines expressed the LD78 gene constitutively. The LD78 protein was detected in culture supernatants and cell lysates of HUT 102, MT-2, MJ, and fresh ATL cells by Western blot analysis. This protein was not found in culture supernatants or cell lysates of monocytic leukemia cells and HL-60 cells, although LD78 transcripts were found in those cells. The discrepancy between gene and protein expression might be explained by the stability of the mRNA. Thus, the protein may be involved in the neoplastic transformation of hematopoietic cells. Images PMID:2687328

  8. Distinct TLR-mediated cytokine production and immunoglobulin secretion in human newborn naïve B cells.

    PubMed

    Pettengill, Matthew A; van Haren, Simon D; Li, Ning; Dowling, David J; Bergelson, Ilana; Jans, Jop; Ferwerda, Gerben; Levy, Ofer

    2016-08-01

    Neonatal innate immunity is distinct from that of adults, which may contribute to increased susceptibility to infection and limit vaccine responses. B cells play critical roles in protection from infection and detect PAMPs via TLRs, that, when co-activated with CD40, can drive B-cell proliferation and Ab production. We characterized the expression of TLRs in circulating B cells from newborns and adults, and evaluated TLR- and CD40-mediated naïve B-cell class-switch recombination (CSR) and cytokine production. Gene expression levels of most TLRs was similar between newborn and adult B cells, except that newborn naïve B cells expressed more TLR9 than adult naïve B cells. Neonatal naïve B cells demonstrated impaired TLR2- and TLR7- but enhanced TLR9-mediated cytokine production. Significantly fewer newborn naïve B cells underwent CSR to produce IgG, an impairment also noted with IL-21 stimulation. Additionally, co-stimulation via CD40 and TLRs induced greater cytokine production in adult B cells. Thus, while newborn naïve B cells demonstrate adult-level expression of TLRs and CD40, the responses to stimulation of these receptors are distinct. Relatively high expression of TLR9 and impaired CD40-mediated Ig secretion contributes to distinct innate and adaptive immunity of human newborns and may inform novel approaches to early-life immunization. PMID:27252169

  9. Antioxidants inhibit SAA formation and pro-inflammatory cytokine release in a human cell model of alkaptonuria

    PubMed Central

    Spreafico, Adriano; Millucci, Lia; Ghezzi, Lorenzo; Geminiani, Michela; Braconi, Daniela; Amato, Loredana; Chellini, Federico; Frediani, Bruno; Moretti, Elena; Collodel, Giulia; Bernardini, Giulia

    2013-01-01

    Objective. 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. Methods. 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. Results. 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. Conclusion. 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. PMID:23704321

  10. TAM receptor-dependent regulation of SOCS3 and MAPKs contributes to pro-inflammatory cytokine downregulation following chronic NOD2 stimulation of human macrophages1

    PubMed Central

    Zheng, Shasha; Hedl, Matija; Abraham, Clara

    2014-01-01

    Microbial-induced cytokine regulation is critical to intestinal immune homeostasis. Acute stimulation of NOD2, the Crohn’s disease-associated sensor of bacterial peptidoglycan, induces cytokines. However, cytokines are attenuated after chronic NOD2 and pattern recognition receptor (PRR) stimulation of macrophages; similar attenuation is observed in intestinal macrophages. The role of Tyro3, Axl and Mer (TAM) receptors in regulating chronic PRR stimulation and NOD2-induced outcomes has not been examined. Moreover, TAM receptors have been relatively less investigated in human macrophages. Whereas TAM receptors did not downregulate acute NOD2-induced cytokines in primary human macrophages, they were essential for downregulating signaling and pro-inflammatory cytokine secretion after chronic NOD2 and TLR4 stimulation. Axl and Mer were similarly required in mice for cytokine downregulation after chronic NOD2 stimulation in vivo and in intestinal tissues. Consistently, TAM expression was increased in human intestinal myeloid-derived cells. Chronic NOD2 stimulation led to IL-10- and TGFβ-dependent TAM upregulation in human macrophages, which in turn, upregulated SOCS3 expression. Restoring SOCS3 expression under TAM knockdown conditions restored chronic NOD2-mediated pro-inflammatory cytokine downregulation. In contrast to the upregulated pro-inflammatory cytokines, attenuated IL-10 secretion was maintained in TAM-deficient macrophages upon chronic NOD2 stimulation. The level of MAPK activation in TAM-deficient macrophages after chronic NOD2 stimulation was insufficient to upregulate IL-10 secretion; however, full restoration of MAPK activation under these conditions restored c-Fos, c-Jun, MAFK and PU.1 binding to the IL-10 promoter and IL-10 secretion. Therefore, TAM receptors are critical for downregulating pro-inflammatory cytokines under the chronic NOD2 stimulation conditions observed in the intestinal environment. PMID:25567680

  11. Dose-dependent modulation of the in vitro cytokine production of human immune competent cells by lead salts.

    PubMed

    Hemdan, Nasr Y A; Emmrich, Frank; Adham, Khadiga; Wichmann, Gunnar; Lehmann, Irina; El-Massry, Azza; Ghoneim, Hossam; Lehmann, Jörg; Sack, Ulrich

    2005-07-01

    Lead pollution constitutes a major health problem that has been intensively debated. To reveal its effects on the immune response, the influence of lead on the in vitro cytokine production of human peripheral mononuclear blood cells was investigated. Isolated cells were exposed to lead acetate or lead chloride for 24 h in the presence of either heat-killed Salmonella enteritidis (hk-SE) or monoclonal antibodies (anti-CD3, anti-CD28, anti-CD40) as cell activators. Our results showed that while higher lead doses are toxic, lower ones evoke immunomodulatory effects. All tested lead doses significantly reduced cell vitality and/or proliferation and affected secretion of proinflammatory, T helper cell type (T(H))1 and T(H)2 cytokines. Expression of interferon (IFN)-gamma, interleukin (IL)-1beta, and tumor necrosis factor (TNF)-alpha was reduced at lower lead doses in both models of cell stimulation. Although hk-SE failed to induce detectable IL-4 levels, monoclonal antibody-induced IL-4, IL-6, and IL-10 secretion increased in the presence of lower lead doses. Also, levels of hk-SE-induced IL-10 and IL-6 secretion were increased at lower lead doses. Thus, exposure to lower doses leads to suppression of the T(H)1 cytokine IFN-gamma and the proinflammatory cytokines TNF-alpha and IL-1beta. The elevated production of IL-4 and/or IL-10 can induce and maintain a T(H)2 immune response and might contribute to increased susceptibility to pathologic agents as well as the incidence of allergic hypersensitivity and/or T(H)2-dominated autoimmune diseases. PMID:15843504

  12. Human decidual macrophages and NK cells differentially express Toll-like receptors and display distinct cytokine profiles upon TLR stimulation.

    PubMed

    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

  13. A human cytokine/single-chain antibody fusion protein for simultaneous delivery of GM-CSF and IL-2 to Ep-CAM overexpressing tumor cells.

    PubMed

    Schanzer, Juergen M; Baeuerle, Patrick A; Dreier, Torsten; Kufer, Peter

    2006-01-01

    Pro-inflammatory cytokines regulate the growth, differentiation, and activation of immune cells and can play a role in antitumor responses. GM-CSF and IL-2 induce tumor rejection in animal models when expressed by tumor cells, and IL-2 is used for the treatment of melanoma and renal cell cancer. However, high doses of GM-CSF and IL-2 are associated with severe side effects in cancer patients. We generated a dual cytokine fusion protein for simultaneous targeted delivery of human GM-CSF and IL-2 to human tumors. The fusion protein is based on a heterodimeric core structure formed by human CH1 and C kappa domains (heterominibody) with C-terminally fused human cytokines and N-terminally fused human single-chain Ab fragments (scFv) specific for the tumor-associated surface antigen epithelial cell adhesion molecule (Ep-CAM). The dual cytokine heterominibody (DCH) was well expressed and secreted by CHO cells, preserved the specific proliferative activities of the two cytokines, and showed Ep-CAM-specific binding to tumor cells. DCH induced potent tumor cell lysis in vitro by two distinct mechanisms. One was activating PBMCs to lyse tumor cells, which was superior to cytotoxicity induced by equimolar ratios of free recombinant human IL-2 and GM-CSF. The other mechanism was redirected lysis, as seen with isolated human T cells, which was solely dependent on the IL-2 fusion part. The therapeutic principle of dual cytokine targeting may warrant in vivo testing of murine-specific analogues in appropriate mouse models and further preclinical development of the less immunogenic, human cytokine- and human Ep-CAM-specific DCH molecule described here. PMID:16483188

  14. Cytokine Gene Polymorphisms and Human Autoimmune Disease in the Era of Genome-Wide Association Studies

    PubMed Central

    2012-01-01

    Cytokine (receptor) genes have traditionally attracted great interest as plausible genetic risk factors for autoimmune disease. Since 2007, the implementation of genome-wide association studies has facilitated the robust identification of allelic variants in more than 35 cytokine loci as susceptibility factors for a wide variety of over 15 autoimmune disorders. In this review, we catalog the gene loci of interleukin, chemokine, and tumor necrosis factor receptor superfamily and ligands that have emerged as autoimmune risk factors. We examine recent progress made in the clarification of the functional mechanisms by which polymorphisms in the genes coding for interleukin-2 receptor alpha (IL2RA), IL7R, and IL23R may alter risk for autoimmune disease, and discuss opposite autoimmune risk alleles found, among others, at the IL10 locus. PMID:22191464

  15. Effect of surgical menopause and estrogen replacement on cytokine release from human blood mononuclear cells.

    PubMed Central

    Pacifici, R; Brown, C; Puscheck, E; Friedrich, E; Slatopolsky, E; Maggio, D; McCracken, R; Avioli, L V

    1991-01-01

    To determine whether mononuclear cell secretory products contribute to the changes in bone turnover that characterize the development of postmenopausal osteoporosis, we evaluated the effects of oophorectomy and subsequent estrogen replacement on the spontaneous secretion of interleukin 1 (IL-1) and tumor necrosis factor alpha (TNF-alpha) and on the phytohemagglutinin A-induced secretion of granulocyte-macrophage colony-stimulating factor (GM-CSF) from peripheral blood mononuclear cells. In 15 healthy premenopausal women who underwent oophorectomy, increases in GM-CSF activity were observed as early as 1 week after surgery, whereas elevations in IL-1 and TNF-alpha and in hydroxyproline/creatinine and calcium/creatinine ratios, two urinary indices of bone resorption, were detectable 2 weeks after the surgical procedure. Six of the oophorectomized women received no estrogen therapy after surgery and in these subjects hydroxyproline/creatinine and calcium/creatinine ratios plateaued 6 weeks postoperatively, and all three cytokines reached the highest levels 8 weeks after oophorectomy, when the study ended. In the remaining 9 women, who were started on estrogen replacement therapy 4 weeks after oophorectomy, decreases in the indices of bone resorption paralleled decreases in the secretion of the cytokines, with lower levels detected after 2 weeks of therapy. In the women who did not receive estrogen therapy, circulating osteocalcin, a marker of bone formation, increased beyond preoperative levels 8 weeks after oophorectomy, whereas in the estrogen-treated subjects osteocalcin remained unchanged in the entire study period. In 9 female controls who underwent simple hysterectomy, cytokine release and biochemical indices of bone turnover did not change after surgery. These data indicate that changes in estrogen status in vivo are associated with the secretion of mononuclear cell immune factors in vitro and suggest that alterations in the local production of bone

  16. p38 MAPK alpha mediates cytokine-induced IL-6 and MMP-3 expression in human cardiac fibroblasts.

    PubMed

    Sinfield, John K; Das, Anupam; O'Regan, David J; Ball, Stephen G; Porter, Karen E; Turner, Neil A

    2013-01-01

    Pre-clinical studies suggest that the p38 MAPK signaling pathway plays a detrimental role in cardiac remodeling, but its role in cardiac fibroblast (CF) function is not well defined. We aimed to identify the p38 MAPK subtypes expressed by human CF, study their activation in response to proinflammatory cytokines, and determine which subtypes were important for expression of specific cytokines and matrix metalloproteinases (MMPs). Quantitative real-time RT-PCR analysis of mRNA levels in human CF cultured from multiple patients revealed a consistent pattern of expression with p38α being most abundant, followed by p38γ, then p38δ and only low expression of p38β (3% of p38α mRNA levels). Immunoblotting confirmed marked protein expression of p38α, γ and δ, with little or no expression of p38β. Phospho-ELISA and combined immunoprecipitation/immunoblotting techniques demonstrated that the proinflammatory cytokines IL-1α and TNFα selectively activated p38α and p38γ, but not p38δ. Selective p38α siRNA gene silencing reduced IL-1α-induced IL-6 and MMP-3 mRNA expression and protein secretion, without affecting IL-1α-induced IL-1β and MMP-9 mRNA expression. In conclusion, human CF express the α, γ and δ subtypes of p38 MAPK, and the α subtype is important for IL-1α-induced IL-6 and MMP-3 expression in this cell type. PMID:23206705

  17. p38 MAPK alpha mediates cytokine-induced IL-6 and MMP-3 expression in human cardiac fibroblasts

    PubMed Central

    Sinfield, John K.; Das, Anupam; O’Regan, David J.; Ball, Stephen G.; Porter, Karen E.; Turner, Neil A.

    2013-01-01

    Pre-clinical studies suggest that the p38 MAPK signaling pathway plays a detrimental role in cardiac remodeling, but its role in cardiac fibroblast (CF) function is not well defined. We aimed to identify the p38 MAPK subtypes expressed by human CF, study their activation in response to proinflammatory cytokines, and determine which subtypes were important for expression of specific cytokines and matrix metalloproteinases (MMPs). Quantitative real-time RT-PCR analysis of mRNA levels in human CF cultured from multiple patients revealed a consistent pattern of expression with p38α being most abundant, followed by p38γ, then p38δ and only low expression of p38β (3% of p38α mRNA levels). Immunoblotting confirmed marked protein expression of p38α, γ and δ, with little or no expression of p38β. Phospho-ELISA and combined immunoprecipitation/immunoblotting techniques demonstrated that the proinflammatory cytokines IL-1α and TNFα selectively activated p38α and p38γ, but not p38δ. Selective p38α siRNA gene silencing reduced IL-1α-induced IL-6 and MMP-3 mRNA expression and protein secretion, without affecting IL-1α-induced IL-1β and MMP-9 mRNA expression. In conclusion, human CF express the α, γ and δ subtypes of p38 MAPK, and the α subtype is important for IL-1α-induced IL-6 and MMP-3 expression in this cell type. PMID:23206705

  18. Human intestinal dendritic cells decrease cytokine release against Salmonella infection in the presence of Lactobacillus paracasei upon TLR activation.

    PubMed

    Bermudez-Brito, Miriam; Muñoz-Quezada, Sergio; Gomez-Llorente, Carolina; Matencio, Esther; Bernal, María J; Romero, Fernando; Gil, Angel

    2012-01-01

    Probiotic bacteria have been shown to modulate immune responses and could have therapeutic effects in allergic and inflammatory disorders. However, little is known about the signalling pathways that are engaged by probiotics. Dendritic cells (DCs) are antigen-presenting cells that are involved in immunity and tolerance. Monocyte-derived dendritic cells (MoDCs) and murine DCs are different from human gut DCs; therefore, in this study, we used human DCs generated from CD34+ progenitor cells (hematopoietic stem cells) harvested from umbilical cord blood; those DCs exhibited surface antigens of dendritic Langerhans cells, similar to the lamina propria DCs in the gut. We report that both a novel probiotic strain isolated from faeces of exclusively breast-fed newborn infants, Lactobacillus paracasei CNCM I-4034, and its cell-free culture supernatant (CFS) decreased pro-inflammatory cytokines and chemokines in human intestinal DCs challenged with Salmonella. Interestingly, the supernatant was as effective as the bacteria in reducing pro-inflammatory cytokine expression. In contrast, the bacterium was a potent inducer of TGF-β2 secretion, whereas the supernatant increased the secretion of TGF-β1 in response to Salmonella. We also showed that both the bacteria and its supernatant enhanced innate immunity through the activation of Toll-like receptor (TLR) signalling. These treatments strongly induced the transcription of the TLR9 gene. In addition, upregulation of the CASP8 and TOLLIP genes was observed. This work demonstrates that L. paracasei CNCM I-4034 enhanced innate immune responses, as evidenced by the activation of TLR signalling and the downregulation of a broad array of pro-inflammatory cytokines. The use of supernatants like the one described in this paper could be an effective and safe alternative to using live bacteria in functional foods. PMID:22905233

  19. The cytokine profile of human NKT cells and PBMCs is dependent on donor sex and stimulus.

    PubMed

    Bernin, Hannah; Fehling, Helena; Marggraff, Claudia; Tannich, Egbert; Lotter, Hannelore

    2016-08-01

    Sex-related variations in natural killer T (NKT) cells may influence immunoregulation and outcome of infectious and autoimmune diseases. We analyzed sex-specific differences in peripheral blood NKTs and peripheral blood mononuclear cells (PBMCs) from men and women and determined the frequencies of NKT cells and their subpopulations [CD4(+); CD8(+); double negative (DN)] and the levels of cytokine production following stimulation with the NKT cell ligands α-Galactosylceramide (αGalCer) and Entamoeba histolytica lipopeptidephosphoglycan (Lotter et al. in PLoS Pathog 5(5):e1000434, 2009). Total and DN NKT cells were more abundant in women than in men. In women, αGalCer induced higher production of intracellular IFNγ, IL-4, IL-17 and TNF by CD4(+) and DN(+)NKT cells. Both ligands induced expression of multiple cytokines in PBMCs and influenced the ratio of NKT cell subpopulations during long-term culture. Although the sex-specific differences in frequencies of NKT cells and their subpopulations were marginal, the significant sex-specific differences in cytokine production might influence disease outcomes. PMID:26895635

  20. Medium and Long Chain Fatty Acids Differentially Modulate Apoptosis and Release of Inflammatory Cytokines in Human Liver Cells.

    PubMed

    Li, Lumin; Wang, Baogui; Yu, Ping; Wen, Xuefang; Gong, Deming; Zeng, Zheling

    2016-06-01

    Medium chain fatty acids (MCFA) can be more easily absorbed and supply energy more rapidly than long chain fatty acids (LCFA). However, little is known about the inflammatory response by the treatment of MCFA in human liver cells. Thus this study used human liver cells (LO2) to evaluate the effects of MCFA on apoptosis and inflammatory response. Tetrazolim-based colorimetric assay and lactate dehydrogenase assay were used to measure the viability of LO2 cells, isolated spleens and liver cells from BALB/C mice. Inverted fluorescence microscopy and flow cytometry were used to assess the cell apoptosis. Activity of superoxide dismutase and malondialdehyde level were measured to determine the oxidative damage. mRNA or protein levels of classical pro-inflammatory cytokines were analyzed by quantitative real-time polymerase chain reaction (qPCR), enzyme-linked immunosorbent assay and western blotting. The results showed that the liver cells treated with the fatty acids at 200 μM for 24 h exhibited good viability. Fatty acids induced inflammatory cytokines at transcriptional and translational levels to a lesser extent than lipopolysaccharide. LCFA (oleic acid) up-regulated tumor necrosis fator-α, monocyte chemoattractant-1 and interleukin-1β while down-regulated IL-6 and IL-8 secretion to a higher extent than MCFA in mRNA and protein levels. These findings suggested that MCFA may induce apoptosis to a less extent and exert more gentle inflammation than LCFA in human liver cells. PMID:27145239

  1. Oligonol, a lychee fruit-derived low-molecular form of polyphenol mixture, suppresses inflammatory cytokine production from human monocytes.

    PubMed

    Lee, Naeun; Shin, Min Sun; Kang, Youna; Park, Kieyoung; Maeda, Takahiro; Nishioka, Hiroshi; Fujii, Hajime; Kang, Insoo

    2016-06-01

    Monocytes produce high levels of inflammatory cytokines including IL-6 and TNF-α that are involved in autoimmunity, inflammatory diseases, cardiovascular disease and obesity. Therapies targeting IL-6 and TNF-α have been utilized in treating chronic inflammatory diseases. Oligonol is a lychee fruit-derived low-molecular form of polyphenol mixture, typically catechin-type monomers and oligomers of proanthocyanidins, which are produced by an oligomerization process. Although previous studies reported anti-inflammatory properties of Oligonol, it is unknown whether and how Oligonol suppresses IL-6 and TNF-α production in human monocytes. The results of our study demonstrate that Oligonol (25μg/ml) decreases the production of IL-6 and TNF-α from human primary monocytes as measured by flow cytometry and ELISA. Such an anti-cytokine effect was likely mediated by the suppression of NF-κB activation without inducing cell death. Our findings raise the possibility of exploring the benefits of Oligonol in controlling inflammatory conditions, especially those associated with monocytes, in humans. PMID:27079270

  2. CD14-dependent and -independent cytokine and chemokine production by human THP-1 monocytes stimulated by Streptococcus suis capsular type 2

    PubMed Central

    SEGURA, M; VADEBONCOEUR, N; GOTTSCHALK, M

    2002-01-01

    Streptococcus suis capsular type 2 is an important aetiologic agent of swine meningitis, and it has been highlighted as a cause of occupational disease leading to meningitis and fulminant sepsis in humans. The objective of the present work was to study the ability of S. suis type 2 to induce the release of tumour necrosis factor alpha (TNF-α), interleukin-1 (IL-1), IL-6, IL-8 and monocyte chemotactic protein one (MCP-1) by human monocytic THP-1 cells. The induction of these five cytokines was dose- and incubation time-dependent, and it was significantly enhanced by pre-treatment of cells with interferon gamma. IL-8 levels were markedly higher compared with those obtained with the other cytokines. However, elevated levels of MCP-1 and IL-6 were also observed. Levels of cytokine induced by heat-killed or live bacteria were similar. Pre-treatment of cells with anti-CD14 monoclonal antibodies suggested that this important host receptor is partially implicated in TNF, IL-1, IL-6 andMCP-1 production, while CD14-independent pathways seem to be responsible for IL-8 production after S. suis stimulation. In addition, blocking studies with anti-TNF and anti-IL-1 antibodies revealed that these cytokines are involved in amplification of the S. suis-induced cytokine cascade. When several different S. suis strains of human or porcine origin were compared, a very heterogeneous pattern of cytokine production was observed. Human strains did not exhibit a clear tendency to induce higher cytokine release by human THP-1 monocytes. The synergistic effect of the up-regulation of cytokines during S. suis meningitis may mediate many of the inflammatory reactions, including the sequestration of leucocytes at the site of infection. PMID:11876746

  3. 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

  4. Butyrate regulates the expression of inflammatory and chemotactic cytokines in human acute leukemic cells during apoptosis.

    PubMed

    Pulliam, Stephanie R; Pellom, Samuel T; Shanker, Anil; Adunyah, Samuel E

    2016-08-01

    Butyrate is a histone deacetylase inhibitor implicated in many studies as a potential therapy for various forms of cancer. High concentrations of butyrate (>1.5mM) 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 24h, 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. 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.

  7. 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

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

    PubMed

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

    2016-01-01

    Interleukin (IL)-38 is the latest member of the IL-1 cytokine family. However, as a result of lacking efficient method to generate relatively large quantity of IL-38, its precise functions are poorly understood. In the present study, the cloning, expression, purification, and activity analysis of recombinant human IL-38 was described. Human IL-38 cDNA was cloned into the prokaryotic expression vector pET-44. The recombinant IL-38 containing a C-hexahistidine tag was expressed in Escherichia coli BL21 (DE3) which induced by isopropyl-β-D-thiogalactoside. The expressed fusion protein was purified by Ni-NTA affinity chromatography. IL-38 protein was largely found in the soluble fraction. The purified IL-38 appeared a single band on SDS-PAGE, the yield of IL-38 was 4 mg from 1 L of bacterial culture, and the purity was more than 98% with low endotoxin level (<0.1 EU/μg). Western blotting confirmed the identity of the purified protein. Activity analysis showed that IL-38 can inhibit effectively the expression of proinflammatory cytokines, such as tumor necrosis factor-α, IL-1β, IL-17, and monocyte chemoattractant protein-1 in lipopolysaccharide-activated THP-1 cells. The production and characterization of biologically active IL-38 will be beneficial for its potential role in clinical applications. PMID:27414784

  9. Dengue virus NS1 enhances viral replication and pro-inflammatory cytokine production in human dendritic cells.

    PubMed

    Alayli, Farah; Scholle, Frank

    2016-09-01

    Dengue virus (DV) has become the most prevalent arthropod borne virus due to globalization and climate change. It targets dendritic cells during infection and leads to production of pro-inflammatory cytokines and chemokines. Several DV non-structural proteins (NS) modulate activation of human dendritic cells. We investigated the effect of DV NS1 on human monocyte-derived dendritic cells (mo-DCs) during dengue infection. NS1 is secreted into the serum of infected individuals where it interacts with various immune mediators and cell types. We purified secreted DV1 NS1 from supernatants of 293T cells that over-express the protein. Upon incubation with mo-DCs, we observed NS1 uptake and enhancement of early DV1 replication. As a consequence, mo-DCs that were pre-exposed to NS1 produced more pro-inflammatory cytokines in response to subsequent DV infection compared to DCs exposed to heat-inactivated NS1 (HNS1). Therefore the presence of exogenous NS1 is able to modulate dengue infection in mo-DCs. PMID:27348054

  10. 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

  11. Regulation and function of the IL-1 family cytokine IL-1F9 in human bronchial epithelial cells.

    PubMed

    Chustz, Regina T; Nagarkar, Deepti R; Poposki, Julie A; Favoreto, Silvio; Avila, Pedro C; Schleimer, Robert P; Kato, Atsushi

    2011-07-01

    The IL-1 family of cytokines, which now includes 11 members, is well known to participate in inflammation. Although the most recently recognized IL-1 family cytokines (IL-1F5-11) have been shown to be expressed in airway epithelial cells, the regulation of their expression and function in the epithelium has not been extensively studied. We investigated the regulation of IL-1F5-11 in primary normal human bronchial epithelial cells. Messenger (m)RNAs for IL-1F6 and IL-1F9, but not IL-1F5, IL-1F8 or IL-1F10, were significantly up-regulated by TNF, IL-1β, IL-17 and the Toll-like receptor (TLR)3 ligand double-stranded (ds)RNA. mRNAs for IL-1F7 and IL-1F11 (IL-33) were weakly up-regulated by some of the cytokines tested. Notably, mRNAs for IL-1F6 and IL-1F9 were synergistically enhanced by the combination of TNF/IL-17 or dsRNA/IL-17. IL-1F9 protein was detected in the supernatant following stimulation with dsRNA or a combination of dsRNA and IL-17. IL-1F6 protein was detected in the cell lysate but was not detected in the supernatant. We screened for the receptor for IL-1F9 and found that lung fibroblasts expressed this receptor. We found that IL-1F9 activated mitogen-activated protein kinases and the transcription factor NF-κB in primary normal human lung fibroblasts. IL-1F9 also stimulated the expression of the neutrophil chemokines IL-8 and CXCL3 and the Th17 chemokine CCL20 in lung fibroblasts. These results suggest that epithelial activation by TLR3 (e.g., by respiratory viral infection) and exposure to cytokines from Th17 cells (IL-17) and inflammatory cells (TNF) may amplify neutrophilic inflammation in the airway via induction of IL-1F9 and activation of fibroblasts. PMID:20870894

  12. 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

  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. Computational and biological analysis of 680 kb of DNA sequence from the human 5q31 cytokine gene cluster region.

    PubMed

    Frazer, K A; Ueda, Y; Zhu, Y; Gifford, V R; Garofalo, M R; Mohandas, N; Martin, C H; Palazzolo, M J; Cheng, J F; Rubin, E M

    1997-05-01

    With the human genome project advancing into what will be a 7- to 10-year DNA sequencing phase, we are presented with the challenge of developing strategies to convert genomic sequence data, as they become available, into biologically meaningful information. We have analyzed 680 kb of noncontiguous DNA sequence from a 1-Mb region of human chromosome 5q31, coupling computational analysis with gene expression studies of tissues isolated from humans as well as from mice containing human YAC transgenes. This genomic interval has been noted previously for containing the cytokine gene cluster and a quantitative trait locus associated with inflammatory diseases. Our analysis identified and verified expression of 16 new genes, as well as 7 previously known genes. Of the total of 23 genes in this region, 78% had similarity matches to sequences in protein databases and 83% had exact expressed sequence tag (EST) database matches. Comparative mapping studies of eight of the new human genes discovered in the 5q31 region revealed that all are located in the syntenic region of mouse chromosome 11q. Our analysis demonstrates an approach for examining human sequence as it is made available from large sequencing programs and has resulted in the discovery of several biomedically important genes, including a cyclin, a transcription factor that is homologous to an oncogene, a protein involved in DNA repair, and several new members of a family of transporter proteins. PMID:9149945

  15. TNF-alpha-independent IL-8 expression: alterations in bacterial challenge dose cause differential human monocytic cytokine response.

    PubMed

    Patrone, Julia B; Bish, Samuel E; Stein, Daniel C

    2006-07-15

    We examined the effects of different bacterial doses of Neisseria gonorrhoeae on the cytokine response of primary human monocytes. The data indicate that a low multiplicity of infection (MOI) challenge (MOI = 0.1) results in substantial production of IL-8 and other chemokines/cytokines, in the absence of significant TNF-alpha production. Positive control challenges (MOI = 10) induced levels of IL-8 that were comparable to the low MOI challenges, but now induced significant levels of TNF-alpha. Induction of IL-8 expression in low MOI challenges was not mediated by an autocrine response as pretreatment of monocytes with neutralizing Abs against TNF-alpha or IL-1beta had no effect on IL-8 expression. IL-8 induction resulting from gonococcal challenge was shown to require NF-kappaB activation, though this activation was limited by the inoculating dose. These data indicate that IL-8 induction results from direct contact between bacteria and monocytes. Analysis of the overall cytokine profile revealed patterns of expression for growth-regulated oncogene, MCP-1, and IL-6 that were similar to IL-8. Analysis of various MAPKs indicated that low MOI challenges were able to efficiently activate both the ERK and p38 pathways, but in contrast to positive control samples, failed to activate the JNK pathway. A lack of phosphorylated JNK leads to decreased production of AP-1 dimers, transcription factors that are critical for efficient transcription of TNF-alpha. Therefore, we propose a mechanism where a low MOI gonococcal challenge results in diminished AP-1 activity and TNF-alpha production while IL-8 levels remain constant. PMID:16818792

  16. 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

  17. 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

    2016-08-01

    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. PMID:27284006

  18. 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

  19. The influence of physical activity on the profile of immune response cells and cytokine synthesis in mice with experimental breast tumors induced by 7,12-dimethylbenzanthracene.

    PubMed

    Abdalla, Douglas R; Murta, Eddie F C; Michelin, Márcia A

    2013-05-01

    This study aims to investigate cytokine synthesis by lymphocytes in the presence of mammary tumors and the interaction with physical activity. For this study, we used 56 female Balb/c, 8-week-old, virgin mice with a body mass between 20 and 30 g. The mice were divided into four groups: a no tumor/nontrained control group; a no tumor/trained group subjected to physical training of swimming in water (30 ± 4°C) for 45 min, five times per week for 8 weeks; a tumor/nontrained (sedentary) group in which the animals received 7,12-dimethylbenzanthracene [(DMBA) 1 mg/ml weekly for 6 weeks)]; and a tumor/trained group in which animals were subjected to the aforementioned DMBA tumor induction and swim training protocols. After the experimental period, immune cells were collected from spleen cell specimens, placed in culture, and stimulated with lipopolysaccharide. The presence of cluster of differentiation (CD)3, CD4, and CD8 markers and the expression of interferon-γ, interleukin (IL)-2, IL-4, IL-10, IL-12, transforming growth factor β, and tumor necrosis factor α cytokines were assessed by flow cytometry and enzyme-linked immunosorbent assay. Physical activity increased the quantities of lymphocytes producing interferon γ, IL-2, IL-12, and tumor necrosis factor α and decreased the quantities of lymphocytes and macrophages expressing IL-4, IL-10, and transforming growth factor β. In contrast, tumor induction, in the absence of swim training, reduced Th1 cytokine levels while increasing the presence of Th2 cytokines and Treg cells. Physical activity promoted reductions in the incidence of tumor development and promoted immune system polarization toward an antitumor Th1 response pattern profile. PMID:22976388

  20. GPU-based skin texture synthesis for digital human model.

    PubMed

    Shen, Zhe; Wang, Lili; Zhao, Yaqian; Zhao, Qinping; Zhao, Meng

    2014-01-01

    Skin synthesis is important for the actual appearance of digital human models. However, it is difficult to design a general algorithm to efficiently produce high quality results. This paper proposes a parallel texture synthesis method for large scale skin of digital human models. Two major procedures are included in this method, a parallel matching procedure and a multi-pass optimizing procedure. Compared with other methods, this algorithm is easy to use, requires only a small size of skin image as input, and generates an arbitrary size of skin texture with high quality. As demonstrated by experiments, the effectiveness of this skin texture synthesis method is confirmed. PMID:25226921

  1. The effects of Saccharum officinarium (sugar cane) molasses on cytokine secretion by human blood cultures.

    PubMed

    Rahiman, Farzana; Pool, Edmund John

    2010-01-01

    This study investigated the effects of sugar cane molasses on the immune system, using cytokines as biomarkers. Whole blood cultures, stimulated in vitro with endotoxin or PHA, were incubated with various concentrations of molasses. No cell death occurred in whole blood cultures incubated with molasses samples. The addition of molasses (800 microg/mL) to unstimulated whole blood cultures resulted in increased levels of the biomarker of inflammation, Interleukin-6 (P < 0.001) and also the biomarker of humoral immunity, Interleukin-10 (P < 0.001). Molasses addition (800 microg/mL) to unstimulated whole blood cultures has no effect on the cell mediated immunity biomarker, Interferon gamma secretion. Molasses has no effect on Interleukin-6, Interleukin-10 and Interferon gamma secretion in stimulated whole blood cultures. Immunostimulation by molasses requires further investigation as it may have potential health impacts. PMID:20391026

  2. Cytokine Production and Antigen Recognition by Human Mucosal Homing Conjunctival Effector Memory CD8+ T Cells

    PubMed Central

    Williams, Geraint P.; Pachnio, Annette; Long, Heather M.; Rauz, Saaeha; Curnow, S. John

    2014-01-01

    Purpose. Conjunctival epithelial T cells are dominated by CD3+CD56-TCRαβ+CD8αβ+ lymphocytes. In this study we explored the antigen experience status, mucosal homing phenotype, cytokine expression, and viral antigen recognition of conjunctival epithelial CD8+ T cells from healthy individuals. Methods. Following ocular surface impression cytology, conjunctival cells were recovered by gentle agitation and analyzed by flow cytometry for cell surface markers, cytokine production (stimulated by phorbol 12-myristate 13-acetate [PMA]/ionomycin), and Epstein-Barr virus (EBV)/cytomegalovirus (CMV) immunodominant epitope recognition using major histocompatibility complex (MHC) class I peptide tetramers. Results. In contrast to peripheral blood, conjunctival epithelial CD8+ T cells were dominantly CD45RA−CCR7− effector memory cells, and the vast majority expressed the mucosal homing integrin αEβ7. Conjunctival memory CD8+ T cells maintained effector functions with the ability to secrete IFN-γ and expression of Granzyme B, although they expressed significantly reduced amounts per cell compared to peripheral blood T cells. Interestingly, herpetic virus-specific CD8+ T cells recognizing epitopes derived from EBV and CMV could be detected in the conjunctival cells of healthy virus carriers, although they were generally at lower frequencies than in the peripheral blood of the same donor. Virus-specific conjunctival CD8+ T cells were dominated by CD45RA−CCR7− effector memory cells that expressed αEβ7. Conclusions. These data demonstrate that the majority of conjunctival epithelial CD8+ T cells are mucosal homing αEβ7+ effector memory T cells, which can recognize viral epitopes and are capable of secreting Granzyme B and IFN-γ. PMID:25395484

  3. The Human Pancreatic Islet Transcriptome: Expression of Candidate Genes for Type 1 Diabetes and the Impact of Pro-Inflammatory Cytokines

    PubMed Central

    Eizirik, Décio L.; Sammeth, Michael; Bouckenooghe, Thomas; Bottu, Guy; Sisino, Giorgia; Igoillo-Esteve, Mariana; Ortis, Fernanda; Santin, Izortze; Colli, Maikel L.; Barthson, Jenny; Bouwens, Luc; Hughes, Linda; Gregory, Lorna; Lunter, Gerton; Marselli, Lorella; Marchetti, Piero; McCarthy, Mark I.; Cnop, Miriam

    2012-01-01

    Type 1 diabetes (T1D) is an autoimmune disease in which pancreatic beta cells are killed by infiltrating immune cells and by cytokines released by these cells. Signaling events occurring in the pancreatic beta cells are decisive for their survival or death in diabetes. We have used RNA sequencing (RNA–seq) to identify transcripts, including splice variants, expressed in human islets of Langerhans under control conditions or following exposure to the pro-inflammatory cytokines interleukin-1β (IL-1β) and interferon-γ (IFN-γ). Based on this unique dataset, we examined whether putative candidate genes for T1D, previously identified by GWAS, are expressed in human islets. A total of 29,776 transcripts were identified as expressed in human islets. Expression of around 20% of these transcripts was modified by pro-inflammatory cytokines, including apoptosis- and inflammation-related genes. Chemokines were among the transcripts most modified by cytokines, a finding confirmed at the protein level by ELISA. Interestingly, 35% of the genes expressed in human islets undergo alternative splicing as annotated in RefSeq, and cytokines caused substantial changes in spliced transcripts. Nova1, previously considered a brain-specific regulator of mRNA splicing, is expressed in islets and its knockdown modified splicing. 25/41 of the candidate genes for T1D are expressed in islets, and cytokines modified expression of several of these transcripts. The present study doubles the number of known genes expressed in human islets and shows that cytokines modify alternative splicing in human islet cells. Importantly, it indicates that more than half of the known T1D candidate genes are expressed in human islets. This, and the production of a large number of chemokines and cytokines by cytokine-exposed islets, reinforces the concept of a dialog between pancreatic islets and the immune system in T1D. This dialog is modulated by candidate genes for the disease at both the immune system and

  4. Actinobacillus actinomycetemcomitans serotype b-specific polysaccharide antigen stimulates production of chemotactic factors and inflammatory cytokines by human monocytes.

    PubMed Central

    Yamaguchi, N; Yamashita, Y; Ikeda, D; Koga, T

    1996-01-01

    Serotype b-specific polysaccharide antigen (SPA) was extracted from whole cells of Actinobacillus actinomycetemcomitans Y4 by autoclaving and purified by chromatography on DEAE-Sephadex A-25 and Sephacryl S-300. SPA induced the release of monocyte and leukocyte chemotactic factors by human monocytes. Polymyxin B had almost no effect on the release of monocyte chemotactic factor, but a monoclonal antibody against SPA markedly inhibited it. Human monocytes stimulated with SPA exhibited the increased mRNA expression of monocyte chemoattractant protein 1 (MCP-1) and a neutrophil chemotactic factor, interleukin-8 (IL-8). On the other hand, SPA induced the release of IL-1, IL-6, and tumor necrosis factor (TNF) and enhanced the expression of IL-1alpha, IL-1beta, IL-6, and TNF alpha (TNF-alpha) mRNAs. Human monocytes expressed MCP-1 and IL-8 mRNAs when stimulated by human recombinant IL-1alpha, I1-1beta, IL-6, and TNF-alpha, suggesting that these inflammatory cytokines induced by SPA might participate in the production of chemotactic factors in human monocytes. PMID:8698480

  5. 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

    2016-05-01

    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. PMID:26924657

  6. Serum Cytokine Responses over the Entire Clinical-Immunological Spectrum of Human Leishmania (L.) infantum chagasi Infection

    PubMed Central

    Ramos, Patrícia Karla; Carvalho, Karina Inácio; Rosa, Daniela Santoro; Rodrigues, Ana Paula; Lima, Luciana Vieira; Campos, Marliane Batista; Gomes, Claudia Maria C.; Laurenti, Márcia Dalastra; Corbett, Carlos Eduardo

    2016-01-01

    The clinical-immunological spectrum of human Leishmania (L.) infantum chagasi infection in Amazonian Brazil was recently reviewed based on clinical, DTH, and IFAT (IgG) evaluations that identified five profiles: three asymptomatic (asymptomatic infection, AI; subclinical resistant infection, SRI; and indeterminate initial infection, III) and two symptomatic (symptomatic infection, SI; American visceral leishmaniasis, AVL; and subclinical oligosymptomatic infection, SOI). TNF-α, IL-4, IL-6, and IL-10 serum cytokines were analyzed using multiplexed Cytometric Bead Array in 161 samples from endemic areas in the Brazilian Amazon: SI [AVL] (21 cases), III (49), SRI (19), SOI (12), AI (36), and a control group [CG] (24). The highest IL-6 serum levels were observed in the SI profile (AVL); higher IL-10 serum levels were observed in SI than in SOI or CG and in AI and III than in SOI; higher TNF-α serum levels were seen in SI than in CG. Positive correlations were found between IL-6 and IL-10 serum levels in the SI and III profiles and between IL-6 and TNF-α and between IL-4 and TNF-α in the III profile. These results provide strong evidence for associating IL-6 and IL-10 with the immunopathogenesis of AVL and help clarify the role of these cytokines in the infection spectrum. PMID:27051668

  7. 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

  8. 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. PMID:27452928

  9. Phloridzin derivatives inhibiting pro-inflammatory cytokine expression in human cystic fibrosis IB3-1 cells.

    PubMed

    Milani, R; Marcellini, A; Montagner, G; Baldisserotto, A; Manfredini, S; Gambari, R; Lampronti, I

    2015-10-12

    Cystic Fibrosis (CF) is the most diffuse autosomal recessive genetic disease affecting Caucasians. A persistent recruitment of neutrophils in the bronchi of CF patients contributes to exacerbate the airway tissue damage, suggesting that modulation of chemokine expression may be an important target for the patient's well being thus the identification of innovative anti-inflammatory drugs is considered a longterm goal to prevent progressive tissue deterioration. Phloridzin, isolated from Malus domestica by a selective molecular imprinting extraction, and its structural analogues, Phloridzin heptapropionate (F1) and Phloridzin tetrapropionate (F2), were initially investigated because of their ability to reduce IL-6 and IL-8 expression in human CF bronchial epithelial cells (IB3-1) stimulated with TNF-α. Release of these cytokines by CF cells was shown to be controlled by the Transcription Factor (TF) NF-kB. The results of the present investigation show that of all the derivatives tested, Phloridzin tetrapropionate (F2) is the most interesting and has greatest potential as it demonstrates inhibitory effects on the expression and production of different cytokines involved in CF inflammation processes, including RANTES, VEGF, GM-CSF, IL-12, G-CSF, MIP-1b, IL-17, IL-10 and IP-10, without any correlated anti-proliferative and pro-apoptotic effects. PMID:26209880

  10. House dust mite extracts activate cultured human dermal endothelial cells to express adhesion molecules and secrete cytokines.

    PubMed

    Arlian, Larry G; Elder, B Laurel; Morgan, Marjorie S

    2009-05-01

    The human skin contacts molecules from house dust mites that are ubiquitous in many environments. These mite-derived molecules may penetrate the skin epidermis and dermis and contact microvascular endothelial cells and influence their function. The purpose of this study was to determine the response of normal human dermal microvascular endothelial cells to extracts of the dust mites, Dermatophagoides farinae, D. pteronyssinus, and Euroglyphus maynei with and without endotoxin (lipopolysaccharide). Endothelial cells were stimulated with mite extracts and the expression of surface molecules and the secretion of cytokines were measured in the absence and presence of polymyxin B to bind endotoxin. All three mite extracts stimulated endothelial cells to express intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), and E-selectin and to secrete interleukin (IL)-6, IL-8, monocyte chemoattractant protein (MCP-1), and granulocyte/macrophage colony stimulating factor (GM-CSF). Euroglyphus maynei-induced expression of all the cell surface molecules was not inhibited when the endotoxin activity in the mite extract was inhibited. In contrast, endothelial cells challenged with D. farinae or D. pteronyssinus extract depleted of endotoxin activity expressed only constitutive levels of ICAM-1, VCAM-1, and E-selectin. D. farinae and E. maynei extracts depleted of endotoxin activity still induced secretion of IL-8 and MCP-1 but at reduced levels. Only constitutive amounts of IL-6, G-CSF, and GM-CSF were secreted in response to any of the endotoxin-depleted mite extracts. Extracts of D. farinae, D. pteronyssinus, and E. maynei contain both endotoxins and other molecules that can stimulate expression of cell adhesion molecules and chemokine receptors and the secretion of cytokines by normal human microvascular endothelial cells. PMID:19496432

  11. 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

  12. 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...

  13. A Human Anti-M2 Antibody Mediates Antibody-Dependent Cell-Mediated Cytotoxicity (ADCC) and Cytokine Secretion by Resting and Cytokine-Preactivated Natural Killer (NK) Cells

    PubMed Central

    Simhadri, Venkateswara R.; Dimitrova, Milena; Mariano, John L.; Zenarruzabeitia, Olatz; Zhong, Weimin; Ozawa, Tatsuhiko; Muraguchi, Atsushi; Kishi, Hiroyuki; Eichelberger, Maryna C.; Borrego, Francisco

    2015-01-01

    The highly conserved matrix protein 2 (M2) is a good candidate for the development of a broadly protective influenza vaccine that induces long-lasting immunity. In animal models, natural killer (NK) cells have been proposed to play an important role in the protection provided by M2-based vaccines through a mechanism of antibody-dependent cell-mediated cytotoxicity (ADCC). We investigated the ability of the human anti-M2 Ab1-10 monoclonal antibody (mAb) to activate human NK cells. They mediated ADCC against M2-expressing cells in the presence of Ab1-10 mAb. Furthermore, NK cell pro-inflammatory cytokine and chemokine secretion is also enhanced when Ab1-10 mAb is present. We also generated cytokine-preactivated NK cells and showed that they still displayed increased effector functions in the presence of Ab1-10 mAb. Thus, our study has demonstrated that human resting and cytokine-preactivated NK cells may have a very important role in the protection provided by anti-M2 Abs. PMID:25915748

  14. 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

  15. Blocking of cell proliferation, cytokines production and genes expression following administration of Chinese herbs in the human mesangial cells.

    PubMed

    Kuo, Y C; Sun, C M; Tsai, W J; Ou, J C; Chen, W P; Lin, C Y

    1999-01-01

    In the hope of identifying agents of therapeutic value in immuoglobulin A nephropathy (IgA-N), we tested crude methanol extracts of 15 Chinese herbs for their effect on human mesangial cell proliferation. The results indicated that 4 out of the 15 crude extracts inhibited human cells proliferation activated by IL-1beta and IL-6. The extracts and their median inhibitory concentrations were as follows (in microg/ml): Ludwiga octovalvis (MLS-052), 49.9 +/- 1.8; Rhus semialata (MLS-053), 31.2 +/- 1.6; Tabernaemontana divaricata (MLS-054), 50.0 +/- 2.1; Amepelopsis brevipedunculata (MLS-059), 42.9 +/- 1.1. These findings indicate that human mesangial cells were most sensitive to MLS-053 treatment. These herbs also decreased interleukin-1beta (IL-1beta) and tumor necrosis factor-alpha (TNF-alpha) production. Moreover, IL- 1beta mRNA expression was inhibited by Rhus semialata (R. semialata; MLS-053). It is unlikely that cytotoxicity was involved, because no cell deaths were observable. We hypothesize that the inhibitory mechanisms of these Chinese herbs may be related to the impairments of gene expression and production of cytokines in human mesangial cells. Plans are underway for the isolation of pure compounds from these Chinese herbs and the elucidation of their mechanisms of action. PMID:10372651

  16. Inflammatory cytokines regulate secretion of VEGF and chemokines by human conjunctival fibroblasts: Role in dysfunctional tear syndrome.

    PubMed

    Nagineni, Chandrasekharam N; William, Abitha; Cherukuri, Aswini; Samuel, William; Hooks, John J; Detrick, Barbara

    2016-02-01

    Ocular surface inflammation is one of the primary mechanisms associated with dysfunctional tear syndrome (DTS), also known as dry eye disease. DTS, more prevalent in older populations, causes ocular discomfort and visual disturbance due to dryness on the surface layer in the eye. We used human conjunctival fibroblast cultures (HCJVF) to investigate the effects of inflammatory cytokines IFN-γ, TNF-α and IL-1β (ITI) on the secretions of VEGF and chemokines. Our results demonstrate the elevated secretion of angiogenic VEGF molecules by ITI without affecting anti-angiogenic molecules, PEDF, endostatin, thrombospondin and sVEGF-R1. The secretion of interferon-γ inducible chemokines, CXCL9, -10, -11 by HCJVF were significantly enhanced by ITI. Our in vitro study supports previously reported observations of elevated VEGF and chemokines in tear fluids of DTS patients, reiterating the role of inflammatory reactions in DTS. PMID:26615568

  17. Human esophageal myofibroblasts secrete proinflammatory cytokines in response to acid and Toll-like receptor 4 ligands.

    PubMed

    Gargus, Matthew; Niu, Chao; Vallone, John G; Binkley, Jana; Rubin, Deborah C; Shaker, Anisa

    2015-06-01

    The pathophysiology of esophageal injury, repair, and inflammation in gastroesophageal reflux-disease (GERD) is complex. Whereas most studies have focused on the epithelial response to GERD injury, we are interested in the stromal response. We hypothesized that subepithelial esophageal myofibroblasts in GERD secrete proinflammatory cytokines in response to injurious agents encountered via epithelial barrier breaches or through dilated epithelial intercellular spaces. We determined the percentage of myofibroblasts [-smooth muscle actin (-SMA)+vimentin+CD31-] in the subepithelial GERD and normal esophageal stroma by immunomorphologic analysis. We performed -SMA coimmunostaining with IL-6 and p65. We established and characterized primary cultures of -SMA+vimentin+CD31-CD45- human esophageal myofibroblasts (HuEso MFs). We modeled GERD by treatment with pH 4.5-acidified media and Toll-like receptor 4 (TLR4) ligands, LPS and high-mobility group box 1 protein (HMGB1), and determined myofibroblast cytokine secretion in response to GERD injury. We demonstrate that spindle-shaped cell myofibroblasts are located near the basement membrane of stratified squamous epithelium in normal esophagus. We identify an increase in subepithelial myofibroblasts and activation of proinflammatory pathways in patients with GERD. Primary cultures of stromal cells obtained from normal esophagus retain myofibroblast morphology and express the acid receptor transient receptor potential channel vanilloid subfamily 1 (TRPV1) and TLR4. HuEso MFs stimulated with acid and TLR4 agonists LPS and HMGB1 increase IL-6 and IL-8 secretion via TRPV1 and NF-B activation. Our work implicates a role for human subepithelial stromal cells in the pathogenesis of GERD-related esophageal injury. Findings of this study can be extended to the investigation of epithelial-stromal interactions in inflammatory esophageal mucosal disorders. PMID:25882613

  18. Human Rhinovirus-induced Proinflammatory Cytokine and Interferon-β Responses in Nasal Epithelial Cells From Chronic Rhinosinusitis Patients

    PubMed Central

    Kim, Ji Heui; Kim, You-Sun; Cho, Gye Song; Kim, Nam Hee; Gong, Chang-Hoon; Lee, Bong-Jae

    2015-01-01

    Purpose Asthma exacerbation from human rhinovirus (HRV) infection is associated with deficient antiviral interferon (IFN) secretion. Although chronic rhinosinusitis (CRS), an inflammatory upper airway disease, is closely linked to asthma, IFN-β responses to HRV infections in human nasal epithelial cells (HNECs) from CRS patients remain to be studied. We evaluated inflammatory and antiviral responses to HRV infection in HNECs from CRS patients. Methods HNECs, isolated from turbinate tissue of 13 patients with CRS and 14 non-CRS controls, were infected with HRV16 for 4 hours. The HRV titer, LDH activity, production of proinflammatory cytokines and IFN-β proteins, and expression levels of RIG-I and MDA5 mRNA were assessed at 8, 24, and 48 hours after HRV16 infection. Results The reduction in viral titer was slightly delayed in the CRS group compared to the non-CRS control group. IL-6 and IL-8 were significantly increased to a similar extent in both groups after HRV infection. In the control group, IFN-β production and MDA5 mRNA expression were significantly increased at 8 and 24 hours after HRV16 infection, respectively. By contrast, in the CRS group, IFN-β was not induced by HRV infection; however, HRV-induced MDA5 mRNA expression was increased, but the increase was slightly delayed compared to the non-CRS control group. The RIG-I mRNA level was not significantly increased by HRV16 infection in either group. Conclusions HRV-induced secretion of proinflammatory cytokines in CRS patients was not different from that in the non-CRS controls. However, reductions in viral titer, IFN-β secretion, and MDA5 mRNA expression in response to HRV infection in CRS patients were slightly impaired compared to those in the controls, suggesting that HRV clearance in CRS patients might be slightly deficient. PMID:26122508

  19. Human esophageal myofibroblasts secrete proinflammatory cytokines in response to acid and Toll-like receptor 4 ligands

    PubMed Central

    Gargus, Matthew; Niu, Chao; Vallone, John G.; Binkley, Jana; Rubin, Deborah C.

    2015-01-01

    The pathophysiology of esophageal injury, repair, and inflammation in gastroesophageal reflux-disease (GERD) is complex. Whereas most studies have focused on the epithelial response to GERD injury, we are interested in the stromal response. We hypothesized that subepithelial esophageal myofibroblasts in GERD secrete proinflammatory cytokines in response to injurious agents encountered via epithelial barrier breaches or through dilated epithelial intercellular spaces. We determined the percentage of myofibroblasts [α-smooth muscle actin (α-SMA)+vimentin+CD31−] in the subepithelial GERD and normal esophageal stroma by immunomorphologic analysis. We performed α-SMA coimmunostaining with IL-6 and p65. We established and characterized primary cultures of α-SMA+vimentin+CD31−CD45− human esophageal myofibroblasts (HuEso MFs). We modeled GERD by treatment with pH 4.5-acidified media and Toll-like receptor 4 (TLR4) ligands, LPS and high-mobility group box 1 protein (HMGB1), and determined myofibroblast cytokine secretion in response to GERD injury. We demonstrate that spindle-shaped cell myofibroblasts are located near the basement membrane of stratified squamous epithelium in normal esophagus. We identify an increase in subepithelial myofibroblasts and activation of proinflammatory pathways in patients with GERD. Primary cultures of stromal cells obtained from normal esophagus retain myofibroblast morphology and express the acid receptor transient receptor potential channel vanilloid subfamily 1 (TRPV1) and TLR4. HuEso MFs stimulated with acid and TLR4 agonists LPS and HMGB1 increase IL-6 and IL-8 secretion via TRPV1 and NF-κB activation. Our work implicates a role for human subepithelial stromal cells in the pathogenesis of GERD-related esophageal injury. Findings of this study can be extended to the investigation of epithelial-stromal interactions in inflammatory esophageal mucosal disorders. PMID:25882613

  20. Lactodifucotetraose, a human milk oligosaccharide, attenuates platelet function and inflammatory cytokine release.

    PubMed

    Newburg, David S; Tanritanir, Ayse C; Chakrabarti, Subrata

    2016-07-01

    Human milk strongly quenches inflammatory processes in vitro, and breastfed infants have lower incidence of inflammatory diseases than those fed artificially. Platelets from neonates, in contrast to those from adults, are less responsive to platelet agonists such as collagen, thrombin, ADP, and epinephrine. Breastfed infants absorb oligosaccharides intact from the human milk in their gut to the circulation. This study was to determine whether these oligosaccharides can attenuate platelet function and platelet secretion of pro-inflammatory proteins, and to identify the active component. The natural mixture of oligosaccharides from human milk and pure individual human milk oligosaccharides were tested for their ability to modulate responses of platelets isolated from human blood following exposure to thrombin, ADP, and collagen. Human milk and the natural mixture of human milk oligosaccharides inhibited platelet release of inflammatory proteins. Of the purified human milk oligosaccharides tested, only lactodifucotetraose (LDFT) significantly inhibited thrombin induced release of the pro-inflammatory proteins RANTES and sCD40L. LDFT also inhibited platelet adhesion to a collagen-coated surface, as well as platelet aggregation induced by ADP or collagen. These data indicate that LDFT may help modulate hemostasis by suppressing platelet-induced inflammatory processes in breastfed infants. This activity suggests further study of LDFT for its potential as a therapeutic agent in infants and adults. PMID:26743063

  1. Effects of cytokines on CYP3A4 expression and reversal of the effects by anti-cytokine agents in the three-dimensionally cultured human hepatoma cell line FLC-4.

    PubMed

    Mimura, Hanaka; Kobayashi, Kaoru; Xu, Linxiaoqing; Hashimoto, Mari; Ejiri, Yoko; Hosoda, Masaya; Chiba, Kan

    2015-02-01

    The expression of hepatic cytochrome P450 (CYP) enzymes is altered under pathological conditions with increased levels of cytokines. In this study, we analyzed the effects of cytokines (interleukin [IL]-1β, IL-6 and tumor necrosis factor α) on the expression of CYP3A4 using newly introduced three-dimensionally cultured human hepatocarcinoma FLC-4 cells. The mRNA level of CYP3A4 was significantly decreased by IL-1β, IL-6 and tumor necrosis factor-α. Formation of α-hydroxytriazolam catalyzed by CYP3A was decreased by IL-1β and IL-6. Pre-treatment with IL-6 enhanced the cytotoxic effects of gefitinib and paclitaxel. In addition, tocilizumab and IL-1 receptor antagonist restored the decreased expression of CYP3A4 mRNA by IL-6 and IL-1β, respectively. These results obtained by using three-dimensionally cultured FLC-4 cells are consistent with results obtained by using primary human hepatocytes and results of clinical studies. Therefore, three-dimensionally cultured FLC-4 cell system may be a promising cellular tool to assess the effects of cytokines on CYP3A4 expression. PMID:25760537

  2. Cytokine Therapies in Neurological Disease.

    PubMed

    Azodi, Shila; Jacobson, Steven

    2016-07-01

    Cytokines are a heterogeneous group of glycoproteins that coordinate physiological functions. Cytokine deregulation is observed in many neurological diseases. This article reviews current research focused on human clinical trials of cytokine and anticytokine therapies in the treatment of several neurological disease including stroke, neuromuscular diseases, neuroinfectious diseases, demyelinating diseases, and neurobehavioral diseases. This research suggests that cytokine therapy applications may play an important role in offering new strategies for disease modulation and treatment. Further, this research provides insights into the causal link between cytokine deregulation and neurological diseases. PMID:27388288

  3. Low Level Pro-inflammatory Cytokines Decrease Connexin36 Gap Junction Coupling in Mouse and Human Islets through Nitric Oxide-mediated Protein Kinase Cδ.

    PubMed

    Farnsworth, Nikki L; Walter, Rachelle L; Hemmati, Alireza; Westacott, Matthew J; Benninger, Richard K P

    2016-02-12

    Pro-inflammatory cytokines contribute to the decline in islet function during the development of diabetes. Cytokines can disrupt insulin secretion and calcium dynamics; however, the mechanisms underlying this are poorly understood. Connexin36 gap junctions coordinate glucose-induced calcium oscillations and pulsatile insulin secretion across the islet. Loss of gap junction coupling disrupts these dynamics, similar to that observed during the development of diabetes. This study investigates the mechanisms by which pro-inflammatory cytokines mediate gap junction coupling. Specifically, as cytokine-induced NO can activate PKCδ, we aimed to understand the role of PKCδ in modulating cytokine-induced changes in gap junction coupling. Isolated mouse and human islets were treated with varying levels of a cytokine mixture containing TNF-α, IL-1β, and IFN-γ. Islet dysfunction was measured by insulin secretion, calcium dynamics, and gap junction coupling. Modulators of PKCδ and NO were applied to determine their respective roles in modulating gap junction coupling. High levels of cytokines caused cell death and decreased insulin secretion. Low levels of cytokine treatment disrupted calcium dynamics and decreased gap junction coupling, in the absence of disruptions to insulin secretion. Decreases in gap junction coupling were dependent on NO-regulated PKCδ, and altered membrane organization of connexin36. This study defines several mechanisms underlying the disruption to gap junction coupling under conditions associated with the development of diabetes. These mechanisms will allow for greater understanding of islet dysfunction and suggest ways to ameliorate this dysfunction during the development of diabetes. PMID:26668311

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

    NASA Technical Reports Server (NTRS)

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

    1999-01-01

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

  5. Immune checkpoint inhibitors enhance cytotoxicity of cytokine-induced killer cells against human myeloid leukaemic blasts.

    PubMed

    Poh, Su Li; Linn, Yeh Ching

    2016-05-01

    We studied whether blockade of inhibitory receptors on cytokine-induced killer (CIK) cells by immune checkpoint inhibitors could increase its anti-tumour potency against haematological malignancies. CIK cultures were generated from seven normal donors and nine patients with acute myeloid leukaemia (AML), acute lymphoblastic leukaemia (ALL) or multiple myeloma (MM). The inhibitory receptors B and T lymphocyte attenuator, CD200 receptor, lymphocyte activation gene-3 (LAG-3) and T cell immunoglobulin and mucin-domain-containing-3 (TIM-3) were present at variable percentages in most CIK cultures, while cytotoxic T lymphocyte-associated protein 4 (CTLA-4), programmed death-1 (PD-1) and killer cell immunoglobulin-like receptors (KIR2DL1/2/3) were expressed at low level in most cultures. Without blockade, myeloid leukaemia cells were susceptible to autologous and allogeneic CIK-mediated cytotoxicity. Blockade of KIR, LAG-3, PD-1 and TIM-3 but not CTLA-4 resulted in remarkable increase in killing against these targets, even in those with poor baseline cytotoxicity. ALL and MM targets were resistant to CIK-mediated cytotoxicity, and blockade of receptors did not increase cytotoxicity to a meaningful extent. Combination of inhibitors against two receptors did not further increase cytotoxicity. Interestingly, potentiation of CIK killing by blocking antibodies was not predicted by expression of receptors on CIK and their respective ligands on the targets. Compared to un-activated T and NK cells, blockade potentiated the cytotoxicity of CIK cells to a greater degree and at a lower E:T ratio, but without significant increase in cytotoxicity against normal white cell. Our findings provide the basis for clinical trial combining autologous CIK cells with checkpoint inhibitors for patients with AML. PMID:26961084

  6. Combined effects of the signal sequence and the major chaperone proteins on the export of human cytokines in Escherichia coli.

    PubMed Central

    Bergès, H; Joseph-Liauzun, E; Fayet, O

    1996-01-01

    We have studied the export of two human proteins in the course of their production in Escherichia coli. The coding sequences of the granulocyte-macrophage colony-stimulating factor and of interleukin 13 were fused to those of two synthetic signal sequences to direct the human proteins to the bacterial periplasm. We found that the total amount of protein varies with the signal peptide-cytokine combination, as does the fraction of it that is soluble in a periplasmic extract. The possibility that the major chaperone proteins such as SecB and the GroEL-GroES and DnaK-DnaJ pairs are limiting factors for the export was tested by overexpressing one or the other of these chaperones concomitantly with the heterologous protein. The GroEL-GroES chaperone pair had no effect on protein production. Overproduction of SecB or DnaK plus DnaJ resulted in a marked increase of the quantity of human proteins in the periplasmic fraction, but this increase depends on the signal peptide-heterologous protein-chaperone association involved. PMID:8572712

  7. Alteration of somatotropic function by proinflammatory cytokines.

    PubMed

    Frost, R A; Lang, C H

    2004-01-01

    Infections direct amino acids away from growth and skeletal muscle accretion toward the hepatic synthesis of acute-phase proteins. The loss of skeletal muscle protein stores results in both a decrease in muscle function and an increase in mortality. In general, muscle protein synthesis is decreased in rodent models of sepsis, as well as after the injection of components of the bacterial cell wall, such as lipopolysaccharide. Although the overexpression of proinflammatory cytokines is known to hasten the loss of skeletal muscle protein, it is not known whether this represents a direct effect of cytokines or results from secondary changes in the IGF system. Plasma concentrations of IGF-I are dramatically lowered by infection in rats, mice, pigs, and steers. The drop in IGF-I often occurs despite an increase in the plasma concentration of somatotropin. Animals are therefore considered to be GH resistant. The IGF bioactivity is determined not only by the plasma concentration of the ligand, but also by IGFBP; IGFBP-3 is the most abundant of these binding proteins and undergoes proteolysis during some catabolic states. In contrast to IGFBP-3, the plasma concentration of inhibitory IGFBP, such as IGFBP-1, is increased during infection. Insulin-like growth factor-binding protein-1 accumulates in skeletal muscle, where it can potentially inhibit IGF-dependent protein synthesis. Insulin-like growth factor-I and IGFBP-1 are regulated at the level of gene transcription by proinflammatory cytokines. Recent studies demonstrate that bacterial components that activate immune cells also activate the innate immune response in skeletal muscle. Lipopolysaccharide increases proinflammatory cytokine messenger RNA expression in muscle from control mice, but not from mice with a mutation in the lipopolysaccharide receptor. Lipopolysaccharide also increases cytokine expression in human and mouse myoblasts. Local expression of cytokines in skeletal muscle may negatively regulate the

  8. Pathogenesis of Streptococcus urinary tract infection depends on bacterial strain and β-hemolysin/cytolysin that mediates cytotoxicity, cytokine synthesis, inflammation and virulence.

    PubMed

    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. 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

  10. Susceptibility of human tonsillar epithelial cells to enterovirus 71 with normal cytokine response.

    PubMed

    Xie, Guang-Cheng; Guo, Ni-Jun; Grénman, Reidar; Wang, Hong; Wang, Ying; Vuorenmma, Minna; Zhang, Qing; Zhang, Shuang; Li, Hui-Ying; Pang, Li-Li; Li, Dan-Di; Jin, Miao; Sun, Xiao-Man; Kong, Xiang-Yu; Duan, Zhao-Jun

    2016-07-01

    A recent histopathologic study implicated human tonsillar crypt epithelium as an important site for EV71 replication in EV71-caused fatal cases. This study aimed to confirm the susceptibility of human tonsillar epithelium to EV71. Two human tonsillar epithelial cell lines (UT-SCC-60A and UT-SCC-60B) were susceptive to EV71, and PI3K/AKT, p38, ERK1/2, and JNK1/2 signal pathways were activated. Interferon-α, IL-8, IL-1β, IL-6 and IL-12p40 were induced and regulated by PI3K/AKT, p38, ERK1/2, and JNK1/2 signal pathways. PI3K/AKT pathway activation appeared to suppress the induction of TNF-α, which induced cell survival by inhibiting GSK-3β. The activation of NF-κB was observed but inhibited by these pathways in EV71 infection. Furthermore, ERK1/2 and JNK1/2 were essential for efficient EV71 replication. Human tonsillar epithelial cells support EV71 replication and display innate antiviral immunity in vitro, indicating that human tonsillar epithelial cells may be novel targets for EV71 infection and replication in vivo. PMID:27107253