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Sample records for activated t-cells nfat

  1. Selective NFAT targeting in T cells ameliorates GvHD while maintaining antitumor activity.

    Vaeth, Martin; Bäuerlein, Carina A; Pusch, Tobias; Findeis, Janina; Chopra, Martin; Mottok, Anja; Rosenwald, Andreas; Beilhack, Andreas; Berberich-Siebelt, Friederike

    2015-01-27

    Graft-versus-host disease (GvHD) is a life-threatening immunological complication after allogenic hematopoietic stem cell transplantation (allo-HCT). The intrinsic graft-versus-leukemia (GvL) effect, however, is the desirable curative benefit. Patients with acute GvHD are treated with cyclosporine A (CsA) or tacrolimus (FK506), which not only often causes severe adverse effects, but also interferes with the anticipated GvL. Both drugs inhibit calcineurin, thus at first suppressing activation of the nuclear factor of activated T cells (NFAT). Therefore, we explored the specific contribution of individual NFAT factors in donor T cells in animal models of GvHD and GvL. Ablation of NFAT1, NFAT2, or a combination of both resulted in ameliorated GvHD, due to reduced proliferation, target tissue homing, and impaired effector function of allogenic donor T cells. In contrast, the frequency of Foxp3(+) regulatory T (Treg) cells was increased and NFAT-deficient Tregs were fully protective in GvHD. CD8(+) T-cell recall response and, importantly, the beneficial antitumor activity were largely preserved in NFAT-deficient effector T cells. Thus, specific inhibition of NFAT opens an avenue for an advanced therapy of GvHD maintaining protective GvL. PMID:25583478

  2. Nickel differentially regulates NFAT and NF-κB activation in T cell signaling

    Nickel is a potent hapten that induces contact hypersensitivity in human skin. While nickel induces the maturation of dendritic cells via NF-κB and p38 MAPK activation, it also exerts immunosuppressive effects on T cells through an unknown mechanism. To elucidate the molecular mechanisms of its effects on T cells, we examined the effects of NiCl2 on mRNA expression in human CD3+ T cells stimulated with CD3 and CD28 antibodies. Using a DNA microarray and Gene Ontology, we identified 70 up-regulated (including IL-1β, IL-6 and IL-8) and 61 down-regulated (including IL-2, IL-4, IL-10 and IFN-γ) immune responsive genes in NiCl2-treated T cells. The DNA microarray results were verified using real-time PCR and a Bio-PlexTM suspension protein array. Suppression of IL-2 and IFN-γ gene transcription by NiCl2 was also confirmed using Jurkat T cells transfected with IL-2 or IFN-γ luciferase reporter genes. To explore the NiCl2-regulated signaling pathway, we examined the binding activity of nuclear proteins to NFAT, AP-1, and NF-κB consensus sequences. NiCl2 significantly and dose-dependently suppressed NFAT- and AP-1-binding activity, but augmented NF-κB-binding activity. Moreover, NiCl2 decreased nuclear NFAT expression in stimulated T cells. Using Jurkat T cells stimulated with PMA/ionomycin, we demonstrated that NiCl2 significantly suppressed stimulation-evoked cytosolic Ca2+ increases, suggesting that NiCl2 regulates NFAT signals by acting as a blocker of Ca2+ release-activated Ca2+ (CRAC) channels. These data showed that NiCl2 decreases NFAT and increases NF-κB signaling in T cells. These results shed light on the effects of nickel on the molecular regulation of T cell signaling. - Graphical Abstract: Nickel suppresses stimulation-evoked cytosolic Ca2+ increase, which results in the suppression of NFAT signals. On the other hand, Ni rather stimulates NF-κB signaling. The effects of Nickel on these transcription factors modulate the expression of various immune

  3. Imaging TCR-Dependent NFAT-Mediated T-Cell Activation with Positron Emission Tomography In Vivo

    Vladimir Ponomarev

    2001-01-01

    Full Text Available A noninvasive method for molecular imaging of T-cell activity in vivo would be of considerable value. It would aid in understanding the role of specific genes and signal transduction pathways in the course of normal and pathologic immune responses, could elucidate temporal dynamics and immune regulation at different stages of disease and following therapy. We developed and assessed a novel method for monitoring the T-cell receptor (TCR -dependent nuclear factor of activated T cells (NFAT -mediated activation of T cells by optical fluorescence imaging (OFI and positron emission tomography (PET. The herpes simplex virus type 1 thymidine kinase/green fluorescent protein [HSV1-tk/GFP (TKGFP ] dual reporter gene was used to monitor NFAT-mediated transcriptional activation in human Jurkat cells. A recombinant retrovirus bearing the NFAT-TKGFP reporter system was constructed in which the TKGFP reporter gene was placed under control of an artificial cis-acting NFAT-specific enhancer. Transduced Jurkat cells were used to establish subcutaneous infiltrates in nude rats. We demonstrated that noninvasive OR and nuclear imaging of T-cell activation is feasible using the NFAT-TKGFP reporter system. PET imaging with [124]FIAU using the NFAT-TKGFP reporter system is sufficiently sensitive to detect T-cell activation in vivo. PET images were confirmed by independent measurements of T-cell activation (e.g., CD69 and induction of GFP fluorescence. PET imaging of TCR-induced NFAT-dependent transcriptional activity may be useful in the assessment of T cell responses, T-cell-based adoptive therapies, vaccination strategies and immunosuppressive drugs.

  4. Interleukin-7 induces HIV replication in primary naive T cells through a nuclear factor of activated T cell (NFAT)-dependent pathway

    Interleukin (IL)-7 plays several roles critical to T cell maturation, survival, and homeostasis. Because of these functions, IL-7 is under investigation as an immune-modulator for therapeutic use in lymphopenic clinical conditions, including HIV. We reported that naive T cells, typically not permissive to HIV, can be productively infected when pre-treated with IL-7. We evaluated the mechanism by which IL-7-mediates this effect. IL-7 potently up-regulated the transcriptional factor NFAT, but had no effect on NFκB. Blocking NFAT activity using a number of reagents, such as Cyclosporin A, FK-506, or the NFAT-specific inhibitor known as VIVIT peptide, all markedly reduced IL-7-mediated induction of HIV replication in naive T cells. Additional neutralization of cytokines present in IL-7-treated cultures and/or those that have NFAT-binding sequences within their promotors indicated that IL-10, IL-4, and most significantly IFNγ, all contribute to IL-7-induction of HIV productive replication in naive T cells. These data clarify the mechanism by which IL-7 can overcome the block to HIV productive infection in naive T cells, despite their quiescent cell status. These findings are relevant to the treatment of HIV disease and understanding HIV pathogenesis in the naive CD4+ T cell compartment, especially in light of the vigorous pursuit of IL-7 as an in vivo immune modulator

  5. Inhibition of nuclear factor of activated T-cells (NFAT suppresses accelerated atherosclerosis in diabetic mice.

    Anna V Zetterqvist

    Full Text Available OBJECTIVE OF THE STUDY: Diabetic patients have a much more widespread and aggressive form of atherosclerosis and therefore, higher risk for myocardial infarction, peripheral vascular disease and stroke, but the molecular mechanisms leading to accelerated damage are still unclear. Recently, we showed that hyperglycemia activates the transcription factor NFAT in the arterial wall, inducing the expression of the pro-atherosclerotic protein osteopontin. Here we investigate whether NFAT activation may be a link between diabetes and atherogenesis. METHODOLOGY AND PRINCIPAL FINDINGS: Streptozotocin (STZ-induced diabetes in apolipoprotein E(-/- mice resulted in 2.2 fold increased aortic atherosclerosis and enhanced pro-inflammatory burden, as evidenced by elevated blood monocytes, endothelial activation- and inflammatory markers in aorta, and pro-inflammatory cytokines in plasma. In vivo treatment with the NFAT blocker A-285222 for 4 weeks completely inhibited the diabetes-induced aggravation of atherosclerosis, having no effect in non-diabetic mice. STZ-treated mice exhibited hyperglycemia and higher plasma cholesterol and triglycerides, but these were unaffected by A-285222. NFAT-dependent transcriptional activity was examined in aorta, spleen, thymus, brain, heart, liver and kidney, but only augmented in the aorta of diabetic mice. A-285222 completely blocked this diabetes-driven NFAT activation, but had no impact on the other organs or on splenocyte proliferation or cytokine secretion, ruling out systemic immunosuppression as the mechanism behind reduced atherosclerosis. Instead, NFAT inhibition effectively reduced IL-6, osteopontin, monocyte chemotactic protein 1, intercellular adhesion molecule 1, CD68 and tissue factor expression in the arterial wall and lowered plasma IL-6 in diabetic mice. CONCLUSIONS: Targeting NFAT signaling may be a novel and attractive approach for the treatment of diabetic macrovascular complications.

  6. FOXP3 Inhibits Activation-Induced NFAT2 Expression in T Cells Thereby Limiting Effector Cytokine Expression

    Torgerson, Troy R; Genin, Anna; Chen, Chunxia; Zhang, Mingce; Zhou, Bin; Añover-Sombke, Stephanie; Frank, M Barton; Dozmorov, Igor; Ocheltree, Elizabeth; Kulmala, Petri; Centola, Michael; Ochs, Hans D.; Wells, Andrew D.; Cron, Randy Q

    2009-01-01

    The forkhead DNA-binding protein FOXP3 is critical for the development and suppressive function of CD4+CD25+ regulatory T cells (TREG), which play a key role in maintaining self-tolerance. Functionally, FOXP3 is capable of repressing transcription of cytokine genes regulated by the Nuclear Factor of Activated T cells (NFAT). Various mechanisms have been proposed by which FOXP3 mediates these effects. Using novel cell lines that inducibly express either wild-type (WT) or mutant FOXP3, we have ...

  7. An altered gp100 peptide ligand with decreased binding by TCR and CD8alpha dissects T cell cytotoxicity from production of cytokines and activation of NFAT

    Niels eSchaft

    2013-09-01

    Full Text Available Altered peptide ligands (APLs provide useful tools to study T cell activation and potentially direct immune responses to improve treatment of cancer patients. To better understand and exploit APLs, we studied the relationship between APLs and T cell function in more detail. Here, we tested a broad panel of gp100(280-288 APLs with respect to T cell cytotoxicity, production of cytokines and activation of Nuclear Factor of Activated T cells (NFAT by human T cells gene-engineered with a gp100-HLA-A2-specific TCRalpha/beta. We demonstrated that gp100-specific cytotoxicity, production of cytokines, and activation of NFAT were not affected by APLs with single amino acid substitutions, except for an APL with an amino acid substitution at position 3 (APL A3, which did not elicit any T cell response. A gp100 peptide with a double amino acid mutation (APL S4S6 elicited T cell cytotoxicity and production of IFNgamma, and to a lesser extent TNFalpha, IL-4, and IL-5, but not production of IL-2 and IL-10, or activation of NFAT. Notably, TCR-mediated functions showed decreases in sensitivities for S4S6 versus gp100 wt peptide, which were minor for cytotoxicity but at least a 1000-fold more prominent for the production of cytokines. TCR-engineered T cells did not bind A3-HLA-A2, but did bind S4S6-HLA-A2 although to a lowered extent compared to wt peptide-HLA-A2. Moreover, S4S6-induced T cell function demonstrated an enhanced dependency on CD8alpha. Taken together, most gp100 APLs functioned as agonists, but A3 and S4S6 peptides acted as a null ligand and partial agonist, respectively. Our results further suggest that TCR-mediated cytotoxicity can be dissected from production of cytokines and activation of NFAT, and that the agonist potential of peptide mutants relates to the extent of binding by TCR and CD8alpha. These findings may facilitate the design of APLs to advance the study of T cell activation and their use for therapeutic applications.

  8. Impaired NFAT and NFκB activation are involved in suppression of CD40 ligand expression by Δ9-tetrahydrocannabinol in human CD4+ T cells

    We have previously reported that Δ9-tetrahydrocannabinol (Δ9-THC), the main psychoactive cannabinoid in marijuana, suppresses CD40 ligand (CD40L) expression by activated mouse CD4+ T cells. CD40L is involved in pathogenesis of many autoimmune and inflammatory diseases. In the present study, we investigated the molecular mechanism of Δ9-THC-mediated suppression of CD40L expression using peripheral blood human T cells. Pretreatment with Δ9-THC attenuated CD40L expression in human CD4+ T cells activated by anti-CD3/CD28 at both the protein and mRNA level, as determined by flow cytometry and quantitative real-time PCR, respectively. Electrophoretic mobility shift assays revealed that Δ9-THC suppressed the DNA-binding activity of both NFAT and NFκB to their respective response elements within the CD40L promoter. An assessment of the effect of Δ9-THC on proximal T cell-receptor (TCR) signaling induced by anti-CD3/CD28 showed significant impairment in the rise of intracellular calcium, but no significant effect on the phosphorylation of ZAP70, PLCγ1/2, Akt, and GSK3β. Collectively, these findings identify perturbation of the calcium-NFAT and NFκB signaling cascade as a key mechanistic event by which Δ9-THC suppresses human T cell function. - Highlights: • Δ9-THC attenuated CD40L expression in activated human CD4+ T cells. • Δ9-THC suppressed DNA-binding activity of NFAT and NFκB. • Δ9-THC impaired elevation of intracellular Ca2+. • Δ9-THC did not affect phosphorylation of ZAP70, PLCγ1/2, Akt, and GSK3β

  9. Expression of NFAT-family proteins in normal human T cells.

    Lyakh, L; P. Ghosh; Rice, N R

    1997-01-01

    NFAT proteins constitute a family of transcription factors involved in mediating signal transduction. Using a panel of specific antisera in immunoprecipitation assays, we found that NFATp (135 kDa) is constitutively expressed in normal human T cells, while synthesis of NFATc (predominant form of 86 kDa) is induced by ionomycin treatment. NFAT4/x was very weakly expressed in unstimulated cells, and its level did not increase upon treatment with activating agents. NFAT3 protein was not observed...

  10. Nuclear factor of activated T cell (NFAT) transcription proteins regulate genes involved in adipocyte metabolism and lipolysis

    NFAT involvement in adipocyte physiological processes was examined by treatment with CsA and/or GSK3β inhibitors (Li+ or TZDZ-8), which prevent or increase NFAT nuclear translocation, respectively. CsA treatment reduced basal and TNFα-induced rates of lipolysis by 50%. Adipocytes preincubated with Li+ or TZDZ-8 prior to CsA and/or TNFα, exhibited enhanced basal rates of lipolysis and complete inhibition of CsA-mediated decreased rates of lipolysis. CsA treatment dramatically reduced the mRNA levels of adipocyte-specific genes (aP2, HSL, PPARγ, ACS and Adn), compared with control or TNFα-treatment, whereas Li+ pretreatment blocked the inhibitory effects of CsA, and mRNA levels of aP2, HSL, PPARγ, and ACS were found at or above control levels. NFAT nuclear localization, assessed by EMSA, confirmed that CsA or Li+ treatments inhibited or increased NFAT nuclear translocation, respectively. These results show that NFAT proteins in mature adipocytes participate in the transcriptional control of genes involved in adipocyte metabolism and lipolysis

  11. HIV-1 gp120 induces NFAT nuclear translocation in resting CD4+ T-cells

    The replication of human immunodeficiency virus (HIV) in CD4+ T-cells is strongly dependent upon the state of activation of infected cells. Infection of sub-optimally activated cells is believed to play a critical role in both the transmission of virus and the persistence of CD4+ T-cell reservoirs. There is accumulating evidence that HIV can modulate signal-transduction pathways in a manner that may facilitate replication in such cells. We previously demonstrated that HIV gp120 induces virus replication in resting CD4+ T cells isolated from HIV-infected individuals. Here, we show that in resting CD4+ T-cells, gp120 activates NFATs and induces their translocation into the nucleus. The HIV LTR encodes NFAT recognition sites, and NFATs may play a critical role in promoting viral replication in sub-optimally activated cells. These observations provide insight into a potential mechanism by which HIV is able to establish infection in resting cells, which may have implications for both transmission of HIV and the persistence of viral reservoirs

  12. Impaired NFAT and NFκB activation are involved in suppression of CD40 ligand expression by Δ{sup 9}-tetrahydrocannabinol in human CD4{sup +} T cells

    Ngaotepprutaram, Thitirat [Department of Pharmacology and Toxicology, Michigan State University (United States); Center for Integrative Toxicology, Michigan State University (United States); Kaplan, Barbara L.F. [Department of Pharmacology and Toxicology, Michigan State University (United States); Center for Integrative Toxicology, Michigan State University (United States); Neuroscience Program, Michigan State University (United States); Kaminski, Norbert E., E-mail: kamins11@msu.edu [Department of Pharmacology and Toxicology, Michigan State University (United States); Center for Integrative Toxicology, Michigan State University (United States)

    2013-11-15

    We have previously reported that Δ{sup 9}-tetrahydrocannabinol (Δ{sup 9}-THC), the main psychoactive cannabinoid in marijuana, suppresses CD40 ligand (CD40L) expression by activated mouse CD4{sup +} T cells. CD40L is involved in pathogenesis of many autoimmune and inflammatory diseases. In the present study, we investigated the molecular mechanism of Δ{sup 9}-THC-mediated suppression of CD40L expression using peripheral blood human T cells. Pretreatment with Δ{sup 9}-THC attenuated CD40L expression in human CD4{sup +} T cells activated by anti-CD3/CD28 at both the protein and mRNA level, as determined by flow cytometry and quantitative real-time PCR, respectively. Electrophoretic mobility shift assays revealed that Δ{sup 9}-THC suppressed the DNA-binding activity of both NFAT and NFκB to their respective response elements within the CD40L promoter. An assessment of the effect of Δ{sup 9}-THC on proximal T cell-receptor (TCR) signaling induced by anti-CD3/CD28 showed significant impairment in the rise of intracellular calcium, but no significant effect on the phosphorylation of ZAP70, PLCγ1/2, Akt, and GSK3β. Collectively, these findings identify perturbation of the calcium-NFAT and NFκB signaling cascade as a key mechanistic event by which Δ{sup 9}-THC suppresses human T cell function. - Highlights: • Δ{sup 9}-THC attenuated CD40L expression in activated human CD4+ T cells. • Δ{sup 9}-THC suppressed DNA-binding activity of NFAT and NFκB. • Δ{sup 9}-THC impaired elevation of intracellular Ca2+. • Δ{sup 9}-THC did not affect phosphorylation of ZAP70, PLCγ1/2, Akt, and GSK3β.

  13. Regulation of T cell activation by HIV-1 accessory proteins: Vpr acts via distinct mechanisms to cooperate with Nef in NFAT-directed gene expression and to promote transactivation by CREB

    Nef and Vpr are lentiviral accessory proteins that have been implicated in regulation of cellular gene expression. We noticed that Vpr can potentiate Nef-induced activation of nuclear factor of activated T cells (NFAT)-dependent transcription. Unlike Nef, which stimulated calcium signaling to activate NFAT, Vpr functioned farther downstream. Similar to the positive effects of Vpr on most of the transcriptional test systems that we used, potentiation of NFAT-directed gene expression was relatively modest in magnitude (two- to threefold) and depended on the cell cycle-arresting capacity of Vpr. By contrast, we found that Vpr could cause more than fivefold upregulation of cyclic AMP response element (CRE)-directed transcription via a mechanism that did not require Vpr-induced G2/M arrest. This effect, however, was only evident under suboptimal conditions known to lead to serine phosphorylation of the CRE binding factor (CREB) but not to CREB-dependent gene expression. This suggested that Vpr may act by stabilizing interactions with CREB and its transcriptional cofactor CREB binding protein (CBP). Indeed, this effect could be blocked by cotransfection of the adenoviral CBP inhibitor E1A. These results provide additional evidence for cell cycle-independent regulation of gene expression by Vpr and implicate CREB as a potentially important target for Vpr action in HIV-infected host cells

  14. Recombinant NFAT1 (NFATp) is regulated by calcineurin in T cells and mediates transcription of several cytokine genes.

    Luo, C.; Burgeon, E; Carew, J A; McCaffrey, P G; Badalian, T M; Lane, W S; Hogan, P G; Rao, A

    1996-01-01

    Transcription factors of the NFAT family play a key role in the transcription of cytokine genes and other genes during the immune response. We have identified two new isoforms of the transcription factor NFAT1 (previously termed NFATp) that are the predominant isoforms expressed in murine and human T cells. When expressed in Jurkat T cells, recombinant NFAT1 is regulated, as expected, by the calmodulin-dependent phosphatase calcineurin, and its function is inhibited by the immunosuppressive a...

  15. Nuclear factor of activated T cells negatively regulates expression of the tumor necrosis factor receptor-related 2 gene in T cells

    Kim, Woon-Ki; Sul, Ok-Ju; Kwak, Jung-Sook; Hur, Hye-Young; Latour, Anne M.; Koller, Beverly H.; Kwon, Byoung S.; Jeong, Choon-Soo

    2010-01-01

    Tumor necrosis factor receptor-related 2 (TR2, HVEM or TNFRSF-14) plays an important role in immune responses, however, the mechanisms regulating its expression are unclear. To understand the control of TR2 gene expression, we studied the upstream region of the gene. Gel supershift assays revealed inducible binding of nuclear factor of activated T cells (NFAT) to a putative NFAT site within the TR2 promoter. Furthermore, cotransfection of a dominant negative NFAT construct, or siRNA for NFAT,...

  16. T-cell activation triggers death receptor-6 expression in a NF-kappa B and NF-AT dependent manner

    Klíma, Martin; Broučková, Adéla; Koc, Michal; Anděra, Ladislav

    2011-01-01

    Roč. 48, 12-13 (2011), s. 1439-1447. ISSN 0161-5890 R&D Projects: GA MŠk 1M0506 Institutional research plan: CEZ:AV0Z50520514 Keywords : TNFRSF21 * T cell s * Jurkat Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.897, year: 2011

  17. Disparate effects of serum on basal and evoked NFAT activity in primary astrocyte cultures

    Furman, Jennifer L.; Artiushin, Irina A.; Norris, Christopher M.

    2009-01-01

    In astrocytes, the Ca2+-dependent protein phosphatase calcineurin (CN) strongly regulates neuro-immune/inflammatory cascades through activation of the transcription factor, nuclear factor of activated T cells (NFAT). While primary cell cultures provide a useful model system for investigating astrocytic CN/NFAT signaling, variable results may arise both within and across labs because of differences in culture conditions. Here, we determined the extent to which serum and cell confluency affect ...

  18. Dual effect of lithium on NFAT5 activity in kidney cells

    Christoph eKüper

    2015-09-01

    Full Text Available Lithium salts are used widely for treatment of bipolar and other mental disorders. Lithium therapy is accompanied frequently by renal side effects, such as nephrogenic diabetes insipidus or chronic kidney disease, but the molecular mechanisms underlying these effects are still poorly understood. In the present study we examined the effect of lithium on the activity of the osmosensitive transcriptional activator nuclear factor of activated T cells 5 (NFAT5, also known as TonEBP, which plays a key role in renal cellular osmoprotection and urinary concentrating ability. Interestingly, we found different effects of lithium on NFAT5 activity, depending on medium osmolality and incubation time. When cells were exposed to lithium for a relative short period (24 h, NFAT5 activity was significantly increased, especially under isosmotic conditions, resulting in an enhanced expression of the NFAT5 target gene heat shock protein 70 (HSP70. Further analysis revealed that the increase of NFAT5 activity depended primarily on an enhanced activity of the c-terminal transactivation domain (TAD, while NFAT5 protein abundance was largely unaffected. Enhanced activity of the TAD is probably mediated by lithium-induced inhibitory phosphorylation of glycogen synthase kinase 3b (GSK-3b, which is in accordance with previous studies. When cells were exposed to lithium for a longer period (96 h, cellular NFAT5 activity and subsequently expression of HSP70 significantly decreased under hyperosmotic conditions, due to diminished NFAT5 protein abundance, also resulting from GSK-3b inhibition. Taken together, our results provide evidence that lithium has opposing effects on NFAT5 activity, depending on environmental osmolality and exposure duration. The potential impacts of these observations on the diverse effects of lithium on kidney function are discussed.

  19. The Involvement of NFAT Transcriptional Activity Suppression in SIRT1-Mediated Inhibition of COX-2 Expression Induced by PMA/Ionomycin

    Yu-Yan Jia; Jie Lu; Yue Huang; Guang Liu; Peng Gao; Yan-Zhen Wan; Ran Zhang; Zhu-Qin Zhang; Rui-Feng Yang; Xiaoqiang Tang; Jing Xu; Xu Wang; Hou-Zao Chen; De-Pei Liu

    2014-01-01

    SIRT1, a class III histone deacetylase, acts as a negative regulator for many transcription factors, and plays protective roles in inflammation and atherosclerosis. Transcription factor nuclear factor of activated T cells (NFAT) has been previously shown to play pro-inflammatory roles in endothelial cells. Inhibition of NFAT signaling may be an attractive target to regulate inflammation in atherosclerosis. However, whether NFAT transcriptional activity is suppressed by SIRT1 remains unknown. ...

  20. Immune-suppressive activity of punicalagin via inhibition of NFAT activation

    Since T cell activation is central to the development of autoimmune diseases, we screened a natural product library comprising 1400 samples of medicinal herbal extracts, to identify compounds that suppress T cell activity. Punicalagin (PCG) isolated from the fruit of Punica granatum was identified as a potent immune suppressant, based on its inhibitory action on the activation of the nuclear factor of activated T cells (NFAT). PCG downregulated the mRNA and soluble protein expression of interleukin-2 from anti-CD3/anti-CD28-stimulated murine splenic CD4+ T cells and suppressed mixed leukocytes reaction (MLR) without exhibiting cytotoxicity to the cells. In vivo, the PCG treatment inhibited phorbol 12-myristate 13-acetate (PMA)-induced chronic ear edema in mice and decreased CD3+ T cell infiltration of the inflamed tissue. These results suggest that PCG could be a potential candidate for the therapeutics of various immune pathologies

  1. FOXP3 Inhibits HIV-1 Infection of CD4 T-cells via Inhibition of LTR Transcriptional Activity

    Selliah, Nithianandan; Zhang, Mingce; White, Sara; Zoltick, Philip; Sawaya, Bassel E.; Finkel, Terri H.; Cron, Randy Q

    2008-01-01

    FOXP3 is a necessary transcription factor for the development and function of CD4+ regulatory T-cells (Tregs). The role of Tregs in HIV-1 infection remains unclear. Here, we show expression of FOXP3 in primary human CD4 T-cells significantly inhibits HIV-1 infection. Since FOXP3 inhibits NFAT activity, and NFAT proteins contribute to HIV-1 transcription, we explore a transcriptional repressive function of HIV-1 LTR by FOXP3. Over-expression of FOXP3 in primary CD4 T-cells inhibits wild-type H...

  2. Characterization of Adapter Protein NRBP as a Negative Regulator of T Cell Activation

    WANG Hui; LIN Zhi-xin; WU Jun

    2008-01-01

    Adapter proteins can regulate the gene transcriptions in disparate signaling pathway by interacting with multiple signaling molecules, including T cell activation signaling. Nuclear receptor binding protein (NRBP), a novel adapter protein, represents a small family of evolutionarily conserved proteins with homologs in Caenorhabditis elegans (C. elegans), Drosophila melanogaster (D.melanogaster), mouse and human. Here, we demonstrated that overexpression of NRBP in Jurkat TAg cells specifically impairs T cell receptor (TCR) or phorbol myristate acetate (PMA)/ionomycin-mediated signaling leading to nuclear factor of activated T cells (NFAT) promoter activation. Furthermore, the N-terminal of NRBP is necessary for its regulation of NFAT activation. Finally, we showed that NRBP has minimal effect on both TCR- and PMA-induced CD69 up-regulation in Jurkat TAg cells, which suggests that NRBP may function downstream of protein kinase C (PKC)/Ras pathway.

  3. Band-pass processing in a GPCR signaling pathway selects for NFAT transcription factor activation.

    Sumit, M; Neubig, R R; Takayama, S; Linderman, J J

    2015-11-01

    Many biological processes are rhythmic and proper timing is increasingly appreciated as being critical for development and maintenance of physiological functions. To understand how temporal modulation of an input signal influences downstream responses, we employ microfluidic pulsatile stimulation of a G-protein coupled receptor, the muscarinic M3 receptor, in single cells with simultaneous real-time imaging of both intracellular calcium and NFAT nuclear localization. Interestingly, we find that reduced stimulation with pulses of ligand can give more efficient transcription factor activation, if stimuli are timed appropriately. Our experiments and computational analyses show that M3 receptor-induced calcium oscillations form a low pass filter while calcium-induced NFAT translocation forms a high pass filter. The combination acts as a band-pass filter optimized for intermediate frequencies of stimulation. We demonstrate that receptor desensitization and NFAT translocation rates determine critical features of the band-pass filter and that the band-pass may be shifted for different receptors or NFAT dynamics. As an example, we show that the two NFAT isoforms (NFAT4 and NFAT1) have shifted band-pass windows for the same receptor. While we focus specifically on the M3 muscarinic receptor and NFAT translocation, band-pass processing is expected to be a general theme that applies to multiple signaling pathways. PMID:26374065

  4. The Toxicity of 3-Monochloro-1,2-Propanediol (+ on Activated T Cell in Mice

    Shuang Guan

    2015-06-01

    Full Text Available This study aimed to clarify the toxicity effects of 3-monochloro-1,2- propanediol(+ (3-MCPD(+ on activated T cell in mice. The toxicity effects of 3-MCPD (+ on murine T lymphocyte responses were evaluated in vitro and in vivo. The data showed that 3-MCPD(+ markedly inhibited ConA-induced T lymphocytes proliferation, Th1/Th2 cytokines production, activated Ca2+/ CaM/I-κB-NF-κB and Ca2+/CaM/CaN/NFAT signal transduction pathways in vitro. Furthermore, administration of 3-MCPD significantly inhibited T cell-mediated DTH response in vivo. These observations indicated that 3-MCPD (+ exhibited suppressive effects on activated mouse T cell in vitro and in vivo.

  5. Micro-RNA Feedback Loops Modulating the Calcineurin/NFAT Signaling Pathway

    Shichina Kannambath

    2016-05-01

    Full Text Available Nuclear factor of activated T cells (NFAT is a family of transcription factors important for innate and adaptive immune responses. NFAT activation is tightly regulated through the calcineurin/NFAT signaling pathway. There is increasing evidence on non-coding RNAs such as miRNAs playing a crucial role in regulating transcription factors and signaling pathways. However, not much is known about microRNAs (miRNAs targeting the calcineurin/NFAT signaling pathway involved in immune response in human. In this study, a comprehensive pathway level analysis has been carried out to identify miRNAs regulating the calcineurin/NFAT signaling pathway. Firstly, by incorporating experimental data and computational predictions, 191 unique miRNAs were identified to be targeting the calcineurin/NFAT signaling pathway in humans. Secondly, combining miRNA expression data from activated T cells and computational predictions, 32 miRNAs were observed to be induced by NFAT transcription factors. Finally, 11 miRNAs were identified to be involved in a feedback loop to modulate the calcineurin/NFAT signaling pathway activity. This data demonstrate the potential role of miRNAs as regulators of the calcineurin/NFAT signaling pathway. The present study thus emphasizes the importance of pathway level analysis to identify miRNAs and understands their role in modulating signaling pathways and transcription factor activity.

  6. NFAT5 promotes proliferation and migration of lung adenocarcinoma cells in part through regulating AQP5 expression

    Guo, Kai, E-mail: gk161@163.com [Department of Respiration, Tangdu Hospital, Fourth Military Medical University, Xi' an 710038 (China); Department of Respiration, 161th Hospital, PLA, Wuhan 430015 (China); Jin, Faguang, E-mail: jinfag@fmmu.edu.cn [Department of Respiration, Tangdu Hospital, Fourth Military Medical University, Xi' an 710038 (China)

    2015-09-25

    The osmoregulated transcription factor nuclear factor of activated T-cells 5(NFAT5), has been found to play important roles in the development of many kinds of human cancers, including breast cancer, colon carcinoma, renal cell carcinoma and melanoma. The aim of the present study was to determine whether NFAT5 is involved in the proliferation and migration of lung adenocarcinoma cells. We found that NFAT5 was upregulated in lung adenocarcinoma cells and knockdown of NFAT5 decreased proliferation and migration of the cells, accompanied by a significant reduction in the expression of AQP5. AQP5 was upregulated in lung adenocarcinoma cells and knockdown of AQP5 also inhibited proliferation and migration of the cells as knockdown of NFAT5 did. Moreover, overexpression of NFAT5 promoted proliferation and migration of lung adenocarcinoma cells, accompanied by a significant increase in the expression of AQP5. These results indicate that NFAT5 plays important roles in proliferation and migration of human lung adenocarcinoma cells through regulating AQP5 expression, providing a new therapeutic option for lung adenocarcinoma therapy. - Highlights: • NFAT5 expression is higher in lung adenocarcinoma cells compared with normal cells. • NFAT5 knockdown decreases proliferation and migration of lung adenocarcinoma cells. • Knockdown of NFAT5 reduces AQP5 expression in human lung adenocarcinoma cells. • Overexpression of NFAT5 promotes proliferation and migration of lung adenocarcinoma cells. • Overexpression of NFAT5 increases AQP5 expression in human lung adenocarcinoma cells.

  7. NFAT5 promotes proliferation and migration of lung adenocarcinoma cells in part through regulating AQP5 expression

    The osmoregulated transcription factor nuclear factor of activated T-cells 5(NFAT5), has been found to play important roles in the development of many kinds of human cancers, including breast cancer, colon carcinoma, renal cell carcinoma and melanoma. The aim of the present study was to determine whether NFAT5 is involved in the proliferation and migration of lung adenocarcinoma cells. We found that NFAT5 was upregulated in lung adenocarcinoma cells and knockdown of NFAT5 decreased proliferation and migration of the cells, accompanied by a significant reduction in the expression of AQP5. AQP5 was upregulated in lung adenocarcinoma cells and knockdown of AQP5 also inhibited proliferation and migration of the cells as knockdown of NFAT5 did. Moreover, overexpression of NFAT5 promoted proliferation and migration of lung adenocarcinoma cells, accompanied by a significant increase in the expression of AQP5. These results indicate that NFAT5 plays important roles in proliferation and migration of human lung adenocarcinoma cells through regulating AQP5 expression, providing a new therapeutic option for lung adenocarcinoma therapy. - Highlights: • NFAT5 expression is higher in lung adenocarcinoma cells compared with normal cells. • NFAT5 knockdown decreases proliferation and migration of lung adenocarcinoma cells. • Knockdown of NFAT5 reduces AQP5 expression in human lung adenocarcinoma cells. • Overexpression of NFAT5 promotes proliferation and migration of lung adenocarcinoma cells. • Overexpression of NFAT5 increases AQP5 expression in human lung adenocarcinoma cells

  8. Mathematics of the NFAT signalling pathway

    Rendall, A.

    2012-01-01

    This paper is a mathematical study of some aspects of the signalling pathway leading to the activation of the transcription factor NFAT (nuclear factor of activated T cells). Activation takes place by dephosphorylation at multiple sites. This has been modelled by Salazar and H\\"ofer using a large system of ordinary differential equations depending on many parameters. With the help of chemical reaction network theory we show that for any choice of the parameters this system has a unique statio...

  9. Vitamin D controls T cell antigen receptor signaling and activation of human T cells

    von Essen, Marina Rode; Kongsbak, Martin; Schjerling, Peter;

    2010-01-01

    Phospholipase C (PLC) isozymes are key signaling proteins downstream of many extracellular stimuli. Here we show that naive human T cells had very low expression of PLC-gamma1 and that this correlated with low T cell antigen receptor (TCR) responsiveness in naive T cells. However, TCR triggering...... led to an upregulation of approximately 75-fold in PLC-gamma1 expression, which correlated with greater TCR responsiveness. Induction of PLC-gamma1 was dependent on vitamin D and expression of the vitamin D receptor (VDR). Naive T cells did not express VDR, but VDR expression was induced by TCR...... signaling via the alternative mitogen-activated protein kinase p38 pathway. Thus, initial TCR signaling via p38 leads to successive induction of VDR and PLC-gamma1, which are required for subsequent classical TCR signaling and T cell activation....

  10. T-Cell Tumor Elimination as a Result of T-Cell Receptor-Mediated Activation

    Ashwell, Jonathan D.; Longo, Dan L.; Bridges, Sandra H.

    1987-07-01

    It has recently been shown that activation of murine T-cell hybridomas with antigen inhibits their growth in vitro. The ``suicide'' of these neoplastic T cells upon stimulation with antigen suggested the possibility that activation via the antigen-specific receptor could also inhibit the growth of neoplastic T cells in vivo. To test this, mice were subcutaneously inoculated with antigen-specific T-cell hybridomas and then treated intraperitoneally with antigen. Administration of the appropriate antigen immediately after inoculation with the T-cell hybridoma abrogated tumor formation; antigen administered after tumors had become established decreased the tumor burden and, in a substantial fraction of animals, led to long-term survival. The efficacy of antigen therapy was due to both a direct inhibitory effect on tumor growth and the induction of host immunity. These studies demonstrate the utility of cellular activation as a means of inhibiting neoplastic T-cell growth in vivo and provide a rationale for studying the use of less selective reagents that can mimic the activating properties of antigen, such as monoclonal antibodies, in the treatment of T-cell neoplasms of unknown antigen specificity.

  11. Role of nuclear factor of activated T-cells 5 in regulating hypertonic-mediated secretin receptor expression in kidney collecting duct cells.

    Chua, Oscar W H; Wong, Kenneth K L; Ko, Ben C; Chung, Sookja K; Chow, Billy K C; Lee, Leo T O

    2016-07-01

    A growing body of evidence suggests that secretin (SCT) is an important element in the osmoregulatory pathway. It is interesting to note that both SCT and its receptor (SCTR) gene are activated upon hyperosmolality in the kidney. However, the precise molecular mechanisms underlying the induction of the SCTR gene expression in response to changes in osmolality have yet to be clarified. Detailed DNA sequence analysis of the promoter regions of the SCTR gene reveals the presence of multiple osmotic response elements (ORE). The ORE is the binding site of a key osmosensitive transactivator, namely, the nuclear factor of activated T-cells 5 (NFAT5). SCTR and NFAT5 are co-expressed in the kidney cortex and medulla collecting duct cells. We therefore hypothesize that NFAT5 is responsible for modulating SCTR expression in hypertonic environments. In this study, we found hypertonicity stimulates the promoter activities and endogenous gene expression of SCTR in mouse kidney cortex collecting duct cells (M1) and inner medulla collecting duct cells (mIMCD3). The overexpression and silencing of NFAT5 further confirmed it to be responsible for the up-regulation of the SCTR gene under hypertonic conditions. A significant increase in the interaction between NFAT5 and the SCTR promoter was also observed following chromatin immunoprecipitation assay. In vivo, osmotic stress up-regulates the SCTR gene in the kidney cortex and medulla of wild-type mice, but does not do so in NFAT5(+/-) animals. Hence, this study provides comprehensive information on how NFAT5 regulates SCTR expression in different osmotic environments. PMID:27080132

  12. Synergistic inhibition of T-cell activation by a cell-permeable ZAP-70 mutant and ctCTLA-4

    Kim, Kyun-Do [Department of Biotechnology, Yonsei University, Seodaemun-Gu, Shinchon-Dong 134, Seoul 120-749 (Korea, Republic of); Choi, Je-Min; Chae, Wook-Jin [Department of Immunobiology, Yale University School of Medicine, New Haven CT 06520 (United States); Lee, Sang-Kyou, E-mail: sjrlee@yonsei.ac.kr [Department of Biotechnology, Yonsei University, Seodaemun-Gu, Shinchon-Dong 134, Seoul 120-749 (Korea, Republic of); ForHumanTech Co., Ltd., Kowoon Institute of Technology Innovation, Bldg. 706, Suwon (Korea, Republic of)

    2009-04-10

    T-cell activation requires TcR-mediated and co-stimulatory signals. ZAP-70 participates in the initial step of TcR signal transduction, while a co-receptor, CTLA-4, inhibits T-cell activation. In previous studies, the overexpression of a ZAP-70 mutant (ZAP-70-Y319F) inhibited the TcR-induced activation of NFAT and IL-2 production, while Hph-1-ctCTLA-4 prevented allergic inflammation. To develop an effective immunosuppressive protein drug that blocks both TcR-mediated and co-stimulatory signaling pathways, a fusion protein of ZAP-70-Y319F and the Hph-1 protein transduction domain was generated. Hph-1-ZAP-70-Y319F inhibited the phosphorylation of ZAP-70-Tyr{sup 319}, LAT-Tyr{sup 191}, and p44/42 MAPK induced by TcR stimulation, NFAT- and AP-1-mediated gene transcription, and the induction of CD69 expression and IL-2 secretion. Hph-1-ZAP-70-Y319F and Hph-1-ctCTLA-4 synergistically inhibited signaling events during T-cell activation. This is the first report to demonstrate the synergistic inhibition of signals transmitted via TcR and its co-stimulatory receptor by cell-permeable forms of intracellular signal mediators.

  13. Hyperoxia Inhibits T Cell Activation in Mice

    Hughes-Fulford, M.; Meissler, J.; Aguayo, E. T.; Globus, R.; Aguado, J.; Candelario, T.

    2013-02-01

    , spleens were removed and the splenocytes were isolated and kept as individual biological samples. We have also examined transcription factors (JASPAR) and pathways of the immune system to help us understand the mechanism of regulation. Results: Our recent mouse immunology experiment aboard STS-131 suggests that the early T cell immune response was inhibited in animals that have been exposed to spaceflight, even 24 hours after return to earth. Moreover, recent experiments in hyperoxic mice show that many of the same genes involved in early T cell activation were altered. Specifically, expression of IL-2Rα, Cxcl2, TNFα, FGF2, LTA and BCL2 genes are dysregulated in mice exposed to hyperoxia. Conclusions: If these hyperoxia-induced changes of gene expression in early T cell activation are additive to the changes seen in the microgravity of spaceflight, there could be an increased infection risk to EVA astronauts, which should be addressed prior to conducting a Mars or other long-term mission.

  14. NFAT2 mediates high glucose-induced glomerular podocyte apoptosis through increased Bax expression

    Background: Hyperglycemia promotes podocyte apoptosis and plays a key role in the pathogenesis of diabetic nephropathy. However, the mechanisms that mediate hyperglycemia-induced podocyte apoptosis is still far from being fully understood. Recent studies reported that high glucose activate nuclear factor of activated T cells (NFAT) in vascular smooth muscle or pancreatic β-cells. Here, we sought to determine if hyperglycemia activates NFAT2 in cultured podocyte and whether this leads to podocyte apoptosis. Meanwhile, we also further explore the mechanisms of NFAT2 activation and NFAT2 mediates high glucose-induced podocyte apoptosis. Methods: Immortalized mouse podocytes were cultured in media containing normal glucose (NG), or high glucose (HG) or HG plus cyclosporine A (a pharmacological inhibitor of calcinerin) or 11R-VIVIT (a special inhibitor of NFAT2). The activation of NFAT2 in podocytes was detected by western blotting and immunofluorescence assay. The role of NFAT2 in hyperglycemia-induced podocyte apoptosis was further evaluated by observing the inhibition of NFAT2 activation by 11R-VIVIT using flow cytometer. Intracellular Ca2+ was monitored in HG-treated podcocytes using Fluo-3/AM. The mRNA and protein expression of apoptosis gene Bax were measured by real time-qPCR and western blotting. Results: HG stimulation activated NFAT2 in a time- and dose-dependent manner in cultured podocytes. Pretreatment with cyclosporine A (500 nM) or 11R-VIVIT (100 nM) completely blocked NFAT2 nuclear accumulation. Meanwhile, the apoptosis effects induced by HG were also abrogated by concomitant treatment with 11R-VIVIT in cultured podocytes. We further found that HG also increased [Ca2+]i, leading to activation of calcineurin, and subsequent increased nuclear accumulation of NFAT2 and Bax expression in cultured podocytes. Conclusion: Our results identify a new finding that HG-induced podocyte apoptosis is mediated by calcineurin/NFAT2/Bax signaling pathway, which may

  15. Mechanism of nuclear factor of activated T-cells mediated FasL expression in corticosterone -treated mouse Leydig tumor cells

    Wang Qian

    2008-06-01

    Full Text Available Abstract Background Fas and FasL is important mediators of apoptosis. We have previously reported that the stress levels of corticosterone (CORT, glucocorticoid in rat increase expression of Fas/FasL and activate Fas/FasL signal pathway in rat Leydig cells, which consequently leads to apoptosis. Moreover, our another study showed that nuclear factor of activated T-cells (NFAT may play a potential role in up-regulation of FasL during CORT-treated rat Leydig cell. It is not clear yet how NFAT is involved in CORT-induced up-regulation of FasL. The aim of the present study is to investigate the molecular mechanisms of NFAT-mediated FasL expression in CORT-treated Leydig cells. Results Western blot analysis showed that NFAT2 expression is present in mouse Leydig tumor cell (mLTC-1. CORT-induced increase in FasL expression in mLTC-1 was ascertained by Western Blot analysis and CORT-induced increase in apoptotic frequency of mLTC-1 cells was detected by FACS with annexin-V labeling. Confocal imaging of NFAT2-GFP in mLTC-1 showed that high level of CORT stimulated NFAT translocation from the cytoplasm to the nucleus. RNA interference-mediated knockdown of NFAT2 significantly attenuated CORT-induced up-regulation of FasL expression in mLTC. These results corroborated our previous finding that NFAT2 is involved in CORT-induced FasL expression in rat Leydig cells and showed that mLTC-1 is a suitable model for investigating the mechanism of CORT-induced FasL expression. The analysis of reporter constructs revealed that the sequence between -201 and +71 of mouse FasL gene is essential for CORT-induced FasL expression. The mutation analysis demonstrated that CORT-induced FasL expression is mediated via an NFAT binding element located in the -201 to +71 region. Co-transfection studies with an NFAT2 expression vector and reporter construct containing -201 to +71 region of FasL gene showed that NFAT2 confer a strong inducible activity to the FasL promoter at its

  16. Improved anti-leukemia activities of adoptively transferred T cells expressing bispecific T-cell engager in mice.

    Liu, X; Barrett, D M; Jiang, S; Fang, C; Kalos, M; Grupp, S A; June, C H; Zhao, Y

    2016-01-01

    Despite the impressive clinical efficacy of T cells engineered to express chimeric antigen receptors (CAR-Ts), the current applications of CAR-T cell therapy are limited by major treatment-related toxicity. Thus, safer yet effective alternative approaches must be developed. In this study, we compared CD19 bispecific T-cell engager (BiTE)-transferred T cells that had been transfected by RNA electroporation with CD19 CAR RNA-transferred T cells both in vitro and in an aggressive Nalm6 leukemia mouse model. BiTEs were secreted from the transferred T cells and enabled both the transferred and bystander T cells to specifically recognize CD19(+) cell lines, with increased tumor killing ability, prolonged functional persistence, increased cytokine production and potent proliferation compared with the CAR-T cells. More interestingly, in comparison with CD3/CD28 bead-stimulated T cells, T cells that were expanded by a rapid T-cell expansion protocol (REP) showed enhanced anti-tumor activities for both CAR and BiTE RNA-electroporated T cells both in vitro and in a Nalm6 mouse model (P<0.01). Furthermore, the REP T cells with BiTE RNAs showed greater efficacy in the Nalm6 leukemia model compared with REP T cells with CAR RNA (P<0.05) and resulted in complete leukemia remission. PMID:27258611

  17. NFAT5 regulates transcription of the mouse telomerase reverse transcriptase gene

    Fujiki, Tsukasa; Udono, Miyako; Kotake, Yojiro; Yamashita, Makiko; Shirahata, Sanetaka; Katakura, Yoshinori, E-mail: katakura.yoshinori.528@m.kyushu-u.ac.jp

    2010-12-10

    We aimed to clarify the transcription-regulation mechanisms of the mouse telomerase reverse transcriptase gene (mTERT). First, we searched for the promoter region required for transcriptional activation of mTERT and identified an enhancer cis-element (named mTERT-EE) located between - 200 and - 179 bp of the mouse TERT gene (mTERT). EMSA results suggested that nuclear factor of activated T cells (NFAT) member proteins bind to mTERT-EE. We then identified NFAT5 as the factor binding to mTERT-EE and found that it activates the transcription of the mTERT core promoter. The results that siRNA directed against NFAT5 significantly reduced mTERT expression and mTERT core promoter activity and that the expressions of NFAT5 and mTERT were well correlated in various mouse tissues except liver suggest that NFAT5 dominantly and directly regulates mTERT expression. To clarify their functionality further, we investigated the effect of hypertonic stress, a known stimulus affecting the expression and transcriptional activity of NFAT5, on mTERT expression. The result indicated that hypertonic stress activates mTERT transcription via the activation and recruitment of NFAT5 to the mTERT promoter. These results provide useful information about the transcription-regulation mechanisms of mTERT.

  18. Elevated Intracellular Calcium Increases Ferritin H Expression Through an NFAT-Independent Posttranscriptional Mechanism Involving mRNA Stabilization

    MacKenzie, Elizabeth L.; Tsuji, Yoshiaki

    2008-01-01

    An increase in intracellular Ca2+ is one of the initiating events in T cell activation. A calcium-mediated signaling cascade in T cells involves activation of calcineurin and the dephosphorylation and translocation of Nuclear Factor of Activated T-cells (NFAT), resulting in the transcriptional activation of target genes such as IL-2. In the present study, we found that increased intracellular calcium leads to induction of the antioxidant protein ferritin H. We previously reported that the fer...

  19. Relations of transcription expression of IL-2 with nuclear factor of activated T cells as well as changes of C-Fos and C-Jun after trauma

    罗艳; 梁华平; 胡承香; 徐祥; 王正国

    2002-01-01

    Objective: To observe the relations among expression of interleukin-2 (IL-2) in spleen lymphocytes, DNA binding activity of nuclear factor of activated T cells (NFAT) and expression of the partly family members C-Fos, C-Jun after trauma. Methods: A murine closed trauma model was used, animals were sacrificed 6, 12 hours and 1, 4, 7, 10, 14 days, respectively after injury. Spleen lymphocytes were isolated from injured mice and stimulated with concanavalin-A. The culture supernatants were harvested and assayed for IL-2 activity. Total RNA was extracted from spleen lymphocytes and assayed for IL-2 mRNA. Nuclear protein was extracted, and the DNA binding activity of NFAT was measured using an electrophoretic mobility shift assay (EMSA), the expressions of C-Fos, C-Jun protein determined by Western blot analysis. Results: The expressions of IL-2 activity and IL-2 mRNA in spleen lymphocytes were decreased in injured mice compared with those in control mice, and the most obvious decrease appeared on the 4th day after injury. The DNA binding activity of NFAT decreased gradually and reached the minimum that was only 41% of the control on the 4th day after injury, which was closely associated with the decline of IL-2 activity and IL-2 mRNA. An decrease in the expression of C-Fos on the 1st and 4th day after injury, trauma had no significant effect on the C-Jun expression.Conclusions: These results suggest that the inhibition of IL-2 expression is partly due to the impairment in the activation of NFAT in injured mice; and the decline in the DNA binding activity of NFAT is partly due to trauma block in the C-Fos expression.

  20. Relations of transcription expression of IL—2 with nuclear factor of activated T cells as well as changes of C—Fos and C—Jun after trauma

    罗艳; 梁华平; 等

    2002-01-01

    Objective:To observe the relations among expression of interleukin-2(IL-2)in spleen lymphocytes,DNA binding activity of nuclear factor of activated T cells(NFAT)and expression of the partly family members C-Fos,C-Jun after trauma.Methods:A murine closed trauma model was used,animals were sacrificed6,12hours and 1,4,7,10,14days,respectively after injury,Spleen lymplocytes were isolated from injured mice and stimulated with concanavalin-A,The culture supernatants were harvested and assayed for IL-2activity,Total RNA was extracted from spleen lymphocytes and assayed for IL-2mRNA.Nuclear protein was extracted,and the DNA binding activity of NFAT was measured using an electrophoretic mobility shift assay(EMSA),the expressions of C-Fos,C-Jun protein determined by Western blot analysis.Results:The expressions of IL-2 activity and IL-2mRNA in spleen lymphocytes were decreased in injured mice compared with those in control mice,and the most obvious decrease appeared on the 4th day after injury,The DNA binding activity of NFAT decreased gradually and reached the minimum that was only41%of the control on the 4th day after injury,which was cloely associated with the decline of IL-2activity and IL-2mRNA.An decrease in the expression ofC-Fos on the lst and 4th day after injury,trauma had no significant effect on the C-Jun expression.Conclusions:These results suggest that the inhibition of IL-2 expression is partly due to the impairment in the activation of NFAT in injured mice;and the decline in the DNA binding activity of NFAT is partly due to trauma block in the C-Fos expression.

  1. Annexin A7 deficiency potentiates cardiac NFAT activity promoting hypertrophic signaling

    Highlights: • Cardiac Anxa7 expression was up-regulated following TAC. • The hypertrophic response following TAC was augmented in Anxa7-deficient mice. • Silencing of Anxa7 increased indicators of HL-1 cardiomyocytes hypertrophy. • Silencing of Anxa7 induced Nfatc1 nuclear translocation. • Silencing of Anxa7 enhanced NFAT-dependent transcriptional activity. - Abstract: Annexin A7 (Anxa7) is a cytoskeletal protein interacting with Ca2+ signaling which in turn is a crucial factor for cardiac remodeling following cardiac injury. The present study explored whether Anxa7 participates in the regulation of cardiac stress signaling. To this end, mice lacking functional Anxa7 (anxa7−/−) and wild-type mice (anxa7+/+) were investigated following pressure overload by transverse aortic constriction (TAC). In addition, HL-1 cardiomyocytes were silenced with Anxa7 siRNA and treated with isoproterenol. Transcript levels were determined by quantitative RT-PCR, transcriptional activity by luciferase reporter assay and protein abundance by Western blotting and confocal microscopy. As a result, TAC treatment increased the mRNA and protein levels of Anxa7 in wild-type mice. Moreover, TAC increased heart weight to body weight ratio and the cardiac mRNA levels of αSka, Nppb, Col1a1, Col3a1 and Rcan1, effects more pronounced in anxa7−/− mice than in anxa7+/+ mice. Silencing of Anxa7 in HL-1 cardiomyocytes significantly increased nuclear localization of Nfatc1. Furthermore, Anxa7 silencing increased NFAT-dependent transcriptional activity as well as αSka, Nppb, and Rcan1 mRNA levels both, under control conditions and following β-adrenergic stimulation by isoproterenol. These observations point to an important role of annexin A7 in the regulation of cardiac NFAT activity and hypertrophic response following cardiac stress conditions

  2. Annexin A7 deficiency potentiates cardiac NFAT activity promoting hypertrophic signaling

    Voelkl, Jakob; Alesutan, Ioana; Pakladok, Tatsiana; Viereck, Robert; Feger, Martina; Mia, Sobuj [Department of Physiology, University of Tübingen, Tübingen (Germany); Schönberger, Tanja [Department of Cardiology and Cardiovascular Medicine, University of Tübingen, Tübingen (Germany); Noegel, Angelika A. [Center for Biochemistry, Institute of Biochemistry I, University of Cologne, Köln (Germany); Gawaz, Meinrad [Department of Cardiology and Cardiovascular Medicine, University of Tübingen, Tübingen (Germany); Lang, Florian, E-mail: florian.lang@uni-tuebingen.de [Department of Physiology, University of Tübingen, Tübingen (Germany)

    2014-02-28

    Highlights: • Cardiac Anxa7 expression was up-regulated following TAC. • The hypertrophic response following TAC was augmented in Anxa7-deficient mice. • Silencing of Anxa7 increased indicators of HL-1 cardiomyocytes hypertrophy. • Silencing of Anxa7 induced Nfatc1 nuclear translocation. • Silencing of Anxa7 enhanced NFAT-dependent transcriptional activity. - Abstract: Annexin A7 (Anxa7) is a cytoskeletal protein interacting with Ca{sup 2+} signaling which in turn is a crucial factor for cardiac remodeling following cardiac injury. The present study explored whether Anxa7 participates in the regulation of cardiac stress signaling. To this end, mice lacking functional Anxa7 (anxa7{sup −/−}) and wild-type mice (anxa7{sup +/+}) were investigated following pressure overload by transverse aortic constriction (TAC). In addition, HL-1 cardiomyocytes were silenced with Anxa7 siRNA and treated with isoproterenol. Transcript levels were determined by quantitative RT-PCR, transcriptional activity by luciferase reporter assay and protein abundance by Western blotting and confocal microscopy. As a result, TAC treatment increased the mRNA and protein levels of Anxa7 in wild-type mice. Moreover, TAC increased heart weight to body weight ratio and the cardiac mRNA levels of αSka, Nppb, Col1a1, Col3a1 and Rcan1, effects more pronounced in anxa7{sup −/−} mice than in anxa7{sup +/+} mice. Silencing of Anxa7 in HL-1 cardiomyocytes significantly increased nuclear localization of Nfatc1. Furthermore, Anxa7 silencing increased NFAT-dependent transcriptional activity as well as αSka, Nppb, and Rcan1 mRNA levels both, under control conditions and following β-adrenergic stimulation by isoproterenol. These observations point to an important role of annexin A7 in the regulation of cardiac NFAT activity and hypertrophic response following cardiac stress conditions.

  3. PLZF(+) Innate T Cells Support the TGF-β-Dependent Generation of Activated/Memory-Like Regulatory T Cells.

    Kang, Byung Hyun; Park, Hyo Jin; Park, Hi Jung; Lee, Jae-Ii; Park, Seong Hoe; Jung, Kyeong Cheon

    2016-06-30

    PLZF-expressing invariant natural killer T cells and CD4 T cells are unique subsets of innate T cells. Both are selected via thymocyte-thymocyte interaction, and they contribute to the generation of activated/memory-like CD4 and CD8 T cells in the thymus via the production of IL-4. Here, we investigated whether PLZF(+) innate T cells also affect the development and function of Foxp3(+) regulatory CD4 T cells. Flow cytometry analysis of the thymus and spleen from both CIITA transgenic C57BL/6 and wild-type BALB/c mice, which have abundant PLZF(+) CD4 T cells and invariant natural killer T cells, respectively, revealed that Foxp3(+) T cells in these mice exhibited a CD103(+) activated/memory-like phenotype. The frequency of CD103(+) regulatory T cells was considerably decreased in PLZF(+) cell-deficient CIITA(Tg)Plzf(lu/lu) and BALB/c.CD1d(-/-) mice as well as in an IL-4-deficient background, such as in CIITA(Tg)IL-4(-/-) and BALB/c.lL-4(-/-) mice, indicating that the acquisition of an activated/memory-like phenotype was dependent on PLZF(+) innate T cells and IL-4. Using fetal thymic organ culture, we further demonstrated that IL-4 in concert with TGF-β enhanced the acquisition of the activated/memory-like phenotype of regulatory T cells. In functional aspects, the activated/memory-like phenotype of Treg cells was directly related to their suppressive function; regulatory T cells of CIITA(Tg)PIV(-/-) mice more efficiently suppressed ovalbumin-induced allergic airway inflammation compared with their counterparts from wild-type mice. All of these findings suggest that PLZF(+) innate T cells also augmented the generation of activated/memory-like regulation via IL-4 production. PMID:27101876

  4. Functional Implications of Plasma Membrane Condensation for T Cell Activation

    Rentero, Carles; Zech, Tobias; Quinn, Carmel M.; Engelhardt, Karin; Williamson, David; Grewal, Thomas; Jessup, Wendy; Harder, Thomas; Gaus, Katharina

    2008-01-01

    The T lymphocyte plasma membrane condenses at the site of activation but the functional significance of this receptor-mediated membrane reorganization is not yet known. Here we demonstrate that membrane condensation at the T cell activation sites can be inhibited by incorporation of the oxysterol 7-ketocholesterol (7KC), which is known to prevent the formation of raft-like liquid-ordered domains in model membranes. We enriched T cells with 7KC, or cholesterol as control, to assess the importa...

  5. TRESK channel as a potential target to treat T-cell mediated immune dysfunction

    Han, Jaehee [Medical Research Center for Neural Dysfunction, Department of Physiology, Institute of Health Sciences, Gyeongsang National University, School of Medicine, Jinju 660-751 (Korea, Republic of); Kang, Dawon, E-mail: dawon@gnu.ac.kr [Medical Research Center for Neural Dysfunction, Department of Physiology, Institute of Health Sciences, Gyeongsang National University, School of Medicine, Jinju 660-751 (Korea, Republic of)

    2009-12-25

    In this review, we propose that TRESK background K{sup +} channel could serve as a potential therapeutic target for T-cell mediated immune dysfunction. TRESK has many immune function-related properties. TRESK is abundantly expressed in the thymus, the spleen, and human leukemic T-lymphocytes. TRESK is highly activated by Ca{sup 2+}, calcineurin, acetylcholine, and histamine which induce hypertrophy, whereas TRESK is inhibited by immunosuppressants, such as cyclosporin A and FK506. Cyclosporine A and FK506 target the binding site of nuclear factor of activated T-cells (NFAT) to inhibit calcineurin. Interestingly, TRESK possesses an NFAT-like docking site that is present at its intracellular loop. Calcineurin has been found to interact with TRESK via specific NFAT-like docking site. When the T-cell is activated, calcineurin can bind to the NFAT-docking site of TRESK. The activation of both TRESK and NFAT via Ca{sup 2+}-calcineurin-NFAT/TRESK pathway could modulate the transcription of new genes in addition to regulating several aspects of T-cell function.

  6. TRESK channel as a potential target to treat T-cell mediated immune dysfunction

    In this review, we propose that TRESK background K+ channel could serve as a potential therapeutic target for T-cell mediated immune dysfunction. TRESK has many immune function-related properties. TRESK is abundantly expressed in the thymus, the spleen, and human leukemic T-lymphocytes. TRESK is highly activated by Ca2+, calcineurin, acetylcholine, and histamine which induce hypertrophy, whereas TRESK is inhibited by immunosuppressants, such as cyclosporin A and FK506. Cyclosporine A and FK506 target the binding site of nuclear factor of activated T-cells (NFAT) to inhibit calcineurin. Interestingly, TRESK possesses an NFAT-like docking site that is present at its intracellular loop. Calcineurin has been found to interact with TRESK via specific NFAT-like docking site. When the T-cell is activated, calcineurin can bind to the NFAT-docking site of TRESK. The activation of both TRESK and NFAT via Ca2+-calcineurin-NFAT/TRESK pathway could modulate the transcription of new genes in addition to regulating several aspects of T-cell function.

  7. DMPD: Activation of lymphokine genes in T cells: role of cis-acting DNA elements thatrespond to T cell activation signals. [Dynamic Macrophage Pathway CSML Database

    Full Text Available 1492121 Activation of lymphokine genes in T cells: role of cis-acting DNA elements ...html) (.csml) Show Activation of lymphokine genes in T cells: role of cis-acting ...DNA elements thatrespond to T cell activation signals. PubmedID 1492121 Title Activation of lymphokine genes in T cells: role

  8. Canine CD4+CD8+ double positive T cells in peripheral blood have features of activated T cells.

    Bismarck, Doris; Schütze, Nicole; Moore, Peter; Büttner, Mathias; Alber, Gottfried; Buttlar, Heiner v

    2012-10-15

    In dogs a CD4(+)CD8(+) double positive T cell subpopulation exists that has not been phenotypically defined yet. We demonstrate that canine CD4(+)CD8(+) T cells are mature CD1a(-) and TCRαβ(+) T cells. To analyse the activation potential of CD4(+)CD8(+) T cells, PBMC from dogs vaccinated against canine distemper virus (CDV) were re-stimulated with CDV. Upon antigen-specific stimulation, the CD4(+)CD8(+) T cell fraction increases and consists nearly exclusively of proliferated cells. Similarly, other features of activated effector/memory T cells such as up-regulation of CD25 and MHC-II as well as down-regulation of CD62L (L-selectin) were observed in CD4(+)CD8(+) T cells after stimulation. Canine CD4(+)CD8(+) T cells are less abundant, but more heterogeneous than porcine ones, comprising a small proportion expressing the β chain of CD8 in addition to the CD8α chain, like human CD4(+)CD8(+) T cells. In summary, this analysis provides the basis for functional characterisation of the in vivo relevance of CD4(+)CD8(+) T cells in T-cell mediated immunity. PMID:22789871

  9. NFAT5 regulates HIV-1 in primary monocytes via a highly conserved long terminal repeat site.

    Shahin Ranjbar

    2006-12-01

    Full Text Available To replicate, HIV-1 capitalizes on endogenous cellular activation pathways resulting in recruitment of key host transcription factors to its viral enhancer. RNA interference has been a powerful tool for blocking key checkpoints in HIV-1 entry into cells. Here we apply RNA interference to HIV-1 transcription in primary macrophages, a major reservoir of the virus, and specifically target the transcription factor NFAT5 (nuclear factor of activated T cells 5, which is the most evolutionarily divergent NFAT protein. By molecularly cloning and sequencing isolates from multiple viral subtypes, and performing DNase I footprinting, electrophoretic mobility shift, and promoter mutagenesis transfection assays, we demonstrate that NFAT5 functionally interacts with a specific enhancer binding site conserved in HIV-1, HIV-2, and multiple simian immunodeficiency viruses. Using small interfering RNA to ablate expression of endogenous NFAT5 protein, we show that the replication of three major HIV-1 viral subtypes (B, C, and E is dependent upon NFAT5 in human primary differentiated macrophages. Our results define a novel host factor-viral enhancer interaction that reveals a new regulatory role for NFAT5 and defines a functional DNA motif conserved across HIV-1 subtypes and representative simian immunodeficiency viruses. Inhibition of the NFAT5-LTR interaction may thus present a novel therapeutic target to suppress HIV-1 replication and progression of AIDS.

  10. Transgelin-2 in B-Cells Controls T-Cell Activation by Stabilizing T Cell - B Cell Conjugates

    Chae, Myoung-Won; Kim, Hye-Ran; Kim, Chang-Hyun; Jun, Chang-Duk; Park, Zee-Yong

    2016-01-01

    The immunological synapse (IS), a dynamic and organized junction between T-cells and antigen presenting cells (APCs), is critical for initiating adaptive immunity. The actin cytoskeleton plays a major role in T-cell reorganization during IS formation, and we previously reported that transgelin-2, an actin-binding protein expressed in T-cells, stabilizes cortical F-actin, promoting T-cell activation in response to antigen stimulation. Transgelin-2 is also highly expressed in B-cells, although no specific function has been reported. In this study, we found that deficiency in transgelin-2 (TAGLN2-/-) in B-cells had little effect on B-cell development and activation, as measured by the expression of CD69, MHC class II molecules, and CD80/86. Nevertheless, in B-cells, transgelin-2 accumulated in the IS during the interaction with T-cells. These results led us to hypothesize that transgelin-2 may also be involved in IS stability in B-cells, thereby influencing T-cell function. Notably, we found that transgelin-2 deficiency in B-cells reduced T-cell activation, as determined by the release of IL-2 and interferon-γ and the expression of CD69. Furthermore, the reduced T-cell activation was correlated with reduced B-cell–T-cell conjugate formation. Collectively, these results suggest that actin stability in B-cells during IS formation is critical for the initiation of adaptive T-cell immunity. PMID:27232882

  11. Nuclear Factor of Activated T cells (NFAT): key regulator of cardiac hypertrophy and skeletal muscle adaptation

    Bourajjaj, M.

    2008-01-01

    Despite significant progress in the prevention and treatment of cardiovascular diseases, heart failure is still a leading cause of morbidity and mortality in industrial countries. Sustained cardiac hypertrophy, which is defined as an increase in heart size resulting from an increase in cardiomyocyte

  12. T Cell Epitope Immunotherapy Induces a CD4+ T Cell Population with Regulatory Activity

    Verhoef Adrienne

    2005-01-01

    Full Text Available Background Synthetic peptides, representing CD4+ T cell epitopes, derived from the primary sequence of allergen molecules have been used to down-regulate allergic inflammation in sensitised individuals. Treatment of allergic diseases with peptides may offer substantial advantages over treatment with native allergen molecules because of the reduced potential for cross-linking IgE bound to the surface of mast cells and basophils. Methods and Findings In this study we address the mechanism of action of peptide immunotherapy (PIT in cat-allergic, asthmatic patients. Cell-division-tracking dyes, cell-mixing experiments, surface phenotyping, and cytokine measurements were used to investigate immunomodulation in peripheral blood mononuclear cells (PBMCs after therapy. Proliferative responses of PBMCs to allergen extract were significantly reduced after PIT. This was associated with modified cytokine profiles generally characterised by an increase in interleukin-10 and a decrease in interleukin-5 production. CD4+ cells isolated after PIT were able to actively suppress allergen-specific proliferative responses of pretreatment CD4neg PBMCs in co-culture experiments. PIT was associated with a significant increase in surface expression of CD5 on both CD4+ and CD8+ PBMCs. Conclusion This study provides evidence for the induction of a population of CD4+ T cells with suppressor/regulatory activity following PIT. Furthermore, up-regulation of cell surface levels of CD5 may contribute to reduced reactivity to allergen.

  13. NFAT5 Contributes to Osmolality-Induced MCP-1 Expression in Mesothelial Cells

    Christoph Küper

    2012-01-01

    Full Text Available Increased expression of the C-C chemokine monocyte chemoattractant protein-1 (MCP-1 in mesothelial cells in response to high glucose concentrations and/or high osmolality plays a crucial role in the development of peritoneal fibrosis during continuous ambulatory peritoneal dialysis (CAPD. Recent studies suggest that in kidney cells osmolality-induced MCP-1 upregulation is mediated by the osmosensitive transcription factor, nuclear factor of activated T cells 5 (NFAT5. The present study addressed the question of whether activation of NFAT5 by hyperosmolality, as present in PD fluids, contributes to MCP-1 expression in the mesothelial cell line Met5A. Hyperosmolality, induced by addition of glucose, NaCl, or mannitol to the growth medium, increased NFAT5 activity and stimulated MCP-1 expression in Met5A cells. siRNA-mediated knockdown of NFAT5 attenuated osmolality-induced MCP-1 upregulation substantially. Hyperosmolality also induced activation of nuclear factor-κB (NF-κB. Accordingly, pharmacological inhibition of NF-κB significantly decreased osmolality-induced MCP-1 expression. Taken together, these results indicate that high osmolalities activate the transcription factor NFAT5 in mesothelial cells. NFAT5 in turn upregulates MCP-1, likely in combination with NF-κB, and thus may participate in the development of peritoneal fibrosis during CAPD.

  14. Activated human CD4 T cells express transporters for both cysteine and cystine

    Levring, Trine Bøegh; Hansen, Ann Kathrine; Nielsen, Bodil Lisbeth;

    2012-01-01

    Because naïve T cells are unable to import cystine due to the absence of cystine transporters, it has been suggested that T cell activation is dependent on cysteine generated by antigen presenting cells. The aim of this study was to determine at which phases during T cell activation exogenous...... cystine/cysteine is required and how T cells meet this requirement. We found that early activation of T cells is independent of exogenous cystine/cysteine, whereas T cell proliferation is strictly dependent of uptake of exogenous cystine/cysteine. Naïve T cells express no or very low levels of both...... cystine and cysteine transporters. However, we found that these transporters become strongly up-regulated during T cell activation and provide activated T cells with the required amount of cystine/cysteine needed for T cell proliferation. Thus, T cells are equipped with mechanisms that allow T cell...

  15. NFAT Signaling in Osteoblasts Regulates the Hematopoietic Niche in the Bone Microenvironment

    Cheryl L. Sesler

    2013-01-01

    Full Text Available Osteoblasts support hematopoietic cell development, including B lymphopoiesis. We have previously shown that the nuclear factor of activated T cells (NFAT negatively regulates osteoblast differentiation and bone formation. Interestingly, in smooth muscle, NFAT has been shown to regulate the expression of vascular cellular adhesion molecule-1 (VCAM-1, a mediator of cell adhesion and signaling during leukocyte development. To examine whether NFAT signaling in osteoblasts regulates hematopoietic development in vivo, we generated a mouse model expressing dominant-negative NFAT driven by the 2.3 kb fragment of the collagen-αI promoter to disrupt NFAT activity in osteoblasts (dnNFATOB. Bone histomorphometry showed that dnNFATOB mice have significant increases in bone volume (44% and mineral apposition rate (131% and decreased trabecular thickness (18%. In the bone microenvironment, dnNFATOB mice displayed a significant increase (87% in Lineage−cKit+Sca-1+ (LSK cells and significant decreases in B220+CD19−IgM− pre-pro-B cells (41% and B220+CD19+IgM+ immature B cells (40%. Concurrent with these findings, LSK cell differentiation into B220+ cells was inhibited when cocultured on differentiated primary osteoblasts harvested from dnNFATOB mice. Gene expression and protein levels of VCAM-1 in osteoblasts decreased in dnNFATOB mice compared to controls. These data suggest that osteoblast-specific NFAT activity mediates early B lymphopoiesis, possibly by regulating VCAM-1 expression on osteoblasts.

  16. Oligomeric Procyanidins Interfere with Glycolysis of Activated T Cells. A Novel Mechanism for Inhibition of T Cell Function

    Masao Goto

    2015-10-01

    Full Text Available Procyanidins, which are flavonoids that are found in a variety of plant species, reduce or prevent immune disorders, such as allergy and autoimmune diseases, through an unknown mechanism. In the present study, we investigated the effects of procyanidins on the T cell receptor (TCR-mediated responses of CD4+ T cells in vitro. Apple procyanidins strongly suppressed the proliferation of splenic CD4+ T cells that were stimulated by an anti-CD3ε antibody, as well as splenocytes stimulated by antigen, but did not alter interleukin (IL-2 secretion from these cells. Furthermore, we found that oligomeric procyanidins strongly suppressed, in a degree of polymerization dependent manner, the proliferation of activated CD4+ T cells, as well as their production of effector cytokines, including glycolysis associated-cytokines, without affecting IL-2 secretion. Additionally, we investigated the inhibitory effects of oligomeric procyanidins on the glycolytic activity of activated CD4+ T cells. We show that pentameric procyanidin suppressed L-lactate production and glucose uptake in activated CD4+ T cells. These results suggest that oligomeric procyanidins suppress the functions of activated CD4+ T cells by interfering with glycolysis.

  17. Antigen-specific T cell activation independently of the MHC: chimeric antigen receptor (CAR-redirected T cells.

    Hinrich eAbken

    2013-11-01

    Full Text Available Adoptive T cell therapy has recently shown powerful in initiating a lasting anti-tumor response with spectacular therapeutic success in some cases. Specific T cell therapy, however, is limited since a number of cancer cells are not recognized by T cells due to various mechanisms including the limited availability of tumor-specific T cells and deficiencies in antigen processing or major histocompatibility complex (MHC expression of cancer cells. To make adoptive cell therapy applicable for the broad variety of cancer entities, patient's T cells are engineered ex vivo with pre-defined specificity by a recombinant chimeric antigen receptor (CAR which consists in the extracellular part of an antibody-derived domain for binding with a tumor-associated antigen and in the intracellular part of a TCR-derived signaling moiety for T cell activation. The specificity of CAR mediated T cell recognition is defined by the antibody domain, is independent of MHC presentation and can be extended to any target for which an antibody is available. We discuss the advantages and limitations of MHC-independent T cell targeting by an engineered CAR and review most significant progress recently made in early stage clinical trials to treat cancer.

  18. HIV-1 latency in actively dividing human T cell lines

    Berkhout Ben

    2008-04-01

    Full Text Available Abstract Background Eradication of HIV-1 from an infected individual cannot be achieved by current drug regimens. Viral reservoirs established early during the infection remain unaffected by anti-retroviral therapy and are able to replenish systemic infection upon interruption of the treatment. Therapeutic targeting of viral latency will require a better understanding of the basic mechanisms underlying the establishment and long-term maintenance of HIV-1 in resting memory CD4 T cells, the most prominent reservoir of transcriptional silent provirus. However, the molecular mechanisms that permit long-term transcriptional control of proviral gene expression in these cells are still not well understood. Exploring the molecular details of viral latency will provide new insights for eventual future therapeutics that aim at viral eradication. Results We set out to develop a new in vitro HIV-1 latency model system using the doxycycline (dox-inducible HIV-rtTA variant. Stable cell clones were generated with a silent HIV-1 provirus, which can subsequently be activated by dox-addition. Surprisingly, only a minority of the cells was able to induce viral gene expression and a spreading infection, eventhough these experiments were performed with the actively dividing SupT1 T cell line. These latent proviruses are responsive to TNFα treatment and alteration of the DNA methylation status with 5-Azacytidine or genistein, but not responsive to the regular T cell activators PMA and IL2. Follow-up experiments in several T cell lines and with wild-type HIV-1 support these findings. Conclusion We describe the development of a new in vitro model for HIV-1 latency and discuss the advantages of this system. The data suggest that HIV-1 proviral latency is not restricted to resting T cells, but rather an intrinsic property of the virus.

  19. Nef-mediated enhancement of cellular activation and human immunodeficiency virus type 1 replication in primary T cells is dependent on association with p21-activated kinase 2

    Olivieri Kevin C

    2011-08-01

    Full Text Available Abstract Background The HIV-1 accessory protein Nef is an important determinant of lentiviral pathogenicity that contributes to disease progression by enhancing viral replication and other poorly understood mechanisms. Nef mediates diverse functions including downmodulation of cell surface CD4 and MHC Class I, enhancement of viral infectivity, and enhancement of T cell activation. Nef interacts with a multiprotein signaling complex that includes Src family kinases, Vav1, CDC42, and activated PAK2 (p21-activated kinase 2. Although previous studies have attempted to identify a biological role for the Nef-PAK2 signaling complex, the importance of this complex and its constituent proteins in Nef function remains unclear. Results Here, we show that Nef mutants defective for PAK2-association, but functional for CD4 and MHC Class I downmodulation and infectivity enhancement, are also defective for the ability to enhance viral replication in primary T cells that are infected and subsequently activated by sub-maximal stimuli (1 μg/ml PHA-P. In contrast, these Nef mutants had little or no effect on HIV-1 replication in T cells activated by stronger stimuli (2 μg/ml PHA-P or anti-CD3/CD28-coated beads. Viruses bearing wild-type Nefs, but not Nef mutants defective for PAK2 association, enhanced NFAT and IL2 receptor promoter activity in Jurkat cells. Moreover, expression of wild-type Nefs, but not mutant Nefs defective for PAK2 association, was sufficient to enhance responsiveness of primary CD4 and CD8 T cells to activating stimuli in Nef-expressing and bystander cells. siRNA knockdown of PAK2 in Jurkat cells reduced NFAT activation induced by anti-CD3/CD28 stimulation both in the presence and absence of Nef, and expression of a PAK2 dominant mutant inhibited Nef-mediated enhancement of CD25 expression. Conclusion Nef-mediated enhancement of cellular activation and viral replication in primary T cells is dependent on PAK2 and on the strength of the

  20. Activated Human T Cells Secrete Exosomes That Participate in IL-2 Mediated Immune Response Signaling

    Wahlgren, Jessica; Tanya De L Karlson; Glader, Pernilla; Telemo, Esbjörn; Valadi, Hadi

    2012-01-01

    It has previously been shown that nano-meter sized vesicles (30–100 nm), exosomes, secreted by antigen presenting cells can induce T cell responses thus showing the potential of exosomes to be used as immunological tools. Additionally, activated CD3+ T cells can secrete exosomes that have the ability to modulate different immunological responses. Here, we investigated what effects exosomes originating from activated CD3+ T cells have on resting CD3+ T cells by studying T cell proliferation, c...

  1. A self-inactivating retrovector incorporating the IL-2 promoter for activation-induced transgene expression in genetically engineered T-cells

    Lejeune Laurence

    2006-11-01

    Full Text Available Abstract Background T-cell activation leads to signaling pathways that ultimately result in induction of gene transcription from the interleukin-2 (IL-2 promoter. We hypothesized that the IL-2 promoter or its synthetic derivatives can lead to T-cell specific, activation-induced transgene expression. Our objective was to develop a retroviral vector for stable and activation-induced transgene expression in T-lymphocytes. Results First, we compared the transcriptional potency of the full-length IL-2 promoter with that of a synthetic promoter composed of 3 repeats of the Nuclear Factor of Activated T-Cells (NFAT element following activation of transfected Jurkat T-cells expressing the large SV40 T antigen (Jurkat TAg. Although the NFAT3 promoter resulted in a stronger induction of luciferase reporter expression post stimulation, the basal levels of the IL-2 promoter-driven reporter expression were much lower indicating that the IL-2 promoter can serve as a more stringent activation-dependent promoter in T-cells. Based on this data, we generated a self-inactivating retroviral vector with the full-length human IL-2 promoter, namely SINIL-2pr that incorporated the enhanced green fluorescent protein (EGFP fused to herpes simplex virus thymidine kinase as a reporter/suicide "bifunctional" gene. Subsequently, Vesicular Stomatitis Virus-G Protein pseudotyped retroparticles were generated for SINIL-2pr and used to transduce the Jurkat T-cell line and the ZAP-70-deficient P116 cell line. Flow cytometry analysis showed that EGFP expression was markedly enhanced post co-stimulation of the gene-modified cells with 1 μM ionomycin and 10 ng/ml phorbol 12-myristate 13-acetate (PMA. This activation-induced expression was abrogated when the cells were pretreated with 300 nM cyclosporin A. Conclusion These results demonstrate that the SINIL-2pr retrovector leads to activation-inducible transgene expression in Jurkat T-cell lines. We propose that this design can be

  2. Ca(2+)-activated K+ channels in human leukemic T cells

    1992-01-01

    Using the patch-clamp technique, we have identified two types of Ca(2+)- activated K+ (K(Ca)) channels in the human leukemic T cell line. Jurkat. Substances that elevate the intracellular Ca2+ concentration ([Ca2+]i), such as ionomycin or the mitogenic lectin phytohemagglutinin (PHA), as well as whole-cell dialysis with pipette solutions containing elevated [Ca2+]i, activate a voltage-independent K+ conductance. Unlike the voltage-gated (type n) K+ channels in these cells, the majority of K(C...

  3. Establishment of human T cell clones exhibiting natural killer-like activity

    Alam, Shahabuddin; Katakura, Yoshinori; Shirahata, Sanetaka

    1999-01-01

    We have succeeded in establishing a method to reproducibly immortalize human T cells by oncogene(s) transfection (Alam, 1997). This study was based on our previous discoveries that these immortalized T cell lines contained T cells which showed cytotoxicity against K562 cells in MHC-nonrestricted manner. Then we attempted to obtain human T cell clones exhibiting natural killer-like activity. Here, we tried to establish clones from these immortalized T cell lines by limiting dilution after stim...

  4. Role of the T cell receptor ligand affinity in T cell activation by bacterial superantigens

    Andersen, P S; Geisler, C; Buus, S; Mariuzza, R A; Karjalainen, K

    2001-01-01

    the SEC3 variants correlated with enhanced binding without any optimum in the binding range covered by native TCR ligands. Comparable studies using anti-TCR antibodies of known affinity confirmed these observations. By comparing the biological potency of the two sets of ligands, we found a significant...... correlation between ligand affinity and ligand potency indicating that it is the density of receptor-ligand complexes in the T cell contact area that determines TCR signaling strength....

  5. Suplatast tosilate alleviates nasal symptoms through the suppression of nuclear factor of activated T-cells-mediated IL-9 gene expression in toluene-2,4-diisocyanate-sensitized rats.

    Mizuguchi, Hiroyuki; Orimoto, Naoki; Kadota, Takuya; Kominami, Takahiro; Das, Asish K; Sawada, Akiho; Tamada, Misaki; Miyagi, Kohei; Adachi, Tsubasa; Matsumoto, Mayumi; Kosaka, Tomoya; Kitamura, Yoshiaki; Takeda, Noriaki; Fukui, Hiroyuki

    2016-03-01

    Histamine H1 receptor (H1R) gene is upregulated in patients with pollinosis; its expression level is highly correlated with the nasal symptom severity. Antihistamines are widely used as allergy treatments because they inhibit histamine signaling by blocking H1R or suppressing H1R signaling as inverse agonists. However, long-term treatment with antihistamines does not completely resolve toluene-2,4-diisocyanate (TDI)-induced nasal symptoms, although it can decrease H1R gene expression to the basal level, suggesting additional signaling is responsible for the pathogenesis of the allergic symptoms. Here, we show that treatment with suplatast tosilate in combination with antihistamines markedly alleviates nasal symptoms in TDI-sensitized rats. Suplatast suppressed TDI-induced upregulation of IL-9 gene expression. Suplatast also suppressed ionomycin/phorbol-12-myristate-13-acetate-induced upregulation of IL-2 gene expression in Jurkat cells, in which calcineurin (CN)/nuclear factor of activated T-cells (NFAT) signaling is known to be involved. Immunoblot analysis demonstrated that suplatast inhibited binding of NFAT to DNA. Furthermore, suplatast suppressed ionomycin-induced IL-9 mRNA upregulation in RBL-2H3 cells, in which CN/NFAT signaling is also involved. These data suggest that suplatast suppressed NFAT-mediated IL-9 gene expression in TDI-sensitized rats and this might be the underlying mechanism of the therapeutic effects of combined therapy of suplatast with antihistamine. PMID:26874672

  6. Mast cells enhance T cell activation: Importance of mast cell-derived TNF

    Nakae, Susumu; Suto, Hajime; Kakurai, Maki; Sedgwick, Jonathon D.; Tsai, Mindy; Galli, Stephen J.

    2005-05-01

    Mast cells are not only important effector cells in immediate hypersensitivity reactions and immune responses to pathogens but also can contribute to T cell-mediated disorders. However, the mechanisms by which mast cells might influence T cells in such settings are not fully understood. We find that mast cells can enhance proliferation and cytokine production in multiple T cell subsets. Mast cell-dependent enhancement of T cell activation can be promoted by FcRI-dependent mast cell activation, TNF production by both mast cells and T cells, and mast cell-T cell contact. However, at high concentrations of cells, mast cells can promote T cell activation independent of IgE or TNF. Finally, mast cells also can promote T cell activation by means of soluble factors. These findings identify multiple mechanisms by which mast cells can influence T cell proliferation and cytokine production. allergy | asthma | autoimmunity | cytokines | immune response

  7. Activation-Induced Cell Death in T Cells and Autoimmunity

    JianZhang; XuemeiXu; YongLiu

    2004-01-01

    Activation-induced cell death (AICD), which results from the interaction between Fas and Fas ligand, is responsible for maintaining tolerance to self-antigen. A defect in AICD may lead to development of autoimmunity. During the last several years, much progress has been made in understanding the mechanism(s) of AICD and its potential role in the pathogenesis of autoimmune diseases. In this review, we summarize the most recent progress on the regulation of the susceptibility of T cells to AICD and its possible involvement in autoimmune diseases.

  8. Expression of recombination-activating genes and T cell receptor gene recombination in the human T cell leukemia cell line

    ZOU Hong-yun; MA Li; MENG Min-jie; YAO Xin-sheng; LIN Ying; WU Zhen-qiang; HE Xiao-wei; WANG Ju-fang; WANG Xiao-ning

    2007-01-01

    Background Recent studies have suggested that mature T cells can change their specificity through reexpression of recombination-activating genes (RAG) and RAG-mediated V(D)J recombination. This process is named receptor revision and has been observed in mature peripheral T cells from transgenic mice and human donors. However, whether the receptor revision in mature T cells is a random or orientated process remains poorly understood. Here we used the Jurkat human T cell line, which represents a mature stage of T cell development, as a model to investigate the regulation of T cell receptor (TCR) gene recombination.Methods TCR Dβ-Jβ signal joint T cell receptor excision DNA circles (sjTRECs) were determined by nested and seminested PCR. Double-strand DNA breaks at recombination signal sequences (RSSs) in the TCRVβ chain locus were detected by ligation-mediated-PCR. Further analysis of the complementarity-determining region 3 (CDR3) size of the TCRVβ chain was examined by the TCR GeneScan technique.Results RAG1, RAG2, and three crucial components of the nonhomologous DNA end-joining (NHEJ) pathway were readily detected in Jurkat. Characteristics of junctional diversity of Dβ2-Jβ2 signal joints and ds RSS breaks associated with the Dβ25' and Dβ 23' sites were detected in DNA from Jurkat cells. CDR3 size and the gene sequences of the TCRVβ chain did not change during cell proliferation.Conclusions RAG1 and RAG2 and ongoing TCR gene recombination are coexpressed in Jurkat cells, but the ongoing recombination process may not play a role in modification of the TCR repertoire. However, the results suggest that Jurkat could be used as a model for studying the regulation of RAGs and V(D)J recombination and as a "special" model of the coexistence of TCR gene rearrangements and "negative" receptor revision.

  9. Nod2 Activates NF-kB in CD4+ T Cells but Its Expression Is Dispensable for T Cell-Induced Colitis

    Zanello, Galliano; Goethel, Ashleigh; Forster, Katharina; Geddes, Kaoru; Philpott, Dana J.; Croitoru, Kenneth

    2013-01-01

    Although the etiology of Crohn's disease (CD) remains elusive this disease is characterized by T cell activation that leads to chronic inflammation and mucosal damage. A potential role for maladaptation between the intestinal microbiota and the mucosal immune response is suggested by the fact that mutations in the pattern recognition receptor Nod2 are associated with higher risks for developing CD. Although Nod2 deletion in CD4+ T cells has been shown to impair the induction of colitis in the murine T cell transfer model, the analysis of T cell intrinsic Nod2 function in T cell differentiation and T cell-mediated immunity is inconsistent between several studies. In addition, the role of T cell intrinsic Nod2 in regulatory T cell (Treg) development and function during colitis remain to be analyzed. In this study, we show that Nod2 expression is higher in activated/memory CD4+ T cells and its expression was inducible after T cell receptor (TCR) ligation. Nod2 stimulation with muramyl dipeptide (MDP) led to a nuclear accumulation of c-Rel NF-kB subunit. Although functionally active in CD4+ T cells, the deletion of Nod2 did not impair the induction and the prevention of colitis in the T cell transfer model. Moreover, Nod2 deletion did not affect the development of Foxp3+ Treg cells in the spleen of recipient mice and Nod2 deficient CD4 T cells expressing the OVA specific transgenic TCR were able to differentiate in Foxp3+ Treg cells after OVA feeding. In vitro, CD25+ Nod2 deficient T cells suppressed T cell proliferation as well as wild type counter parts and T cell stimulation with MDP did not affect the proliferation and the cytokine secretion of T cells. In conclusion, our data indicate that Nod2 is functional in murine CD4+ T cells but its expression is dispensable for the T cell regulation of colitis. PMID:24324812

  10. Human retinal pigment epithelial cell-induced apoptosis in activated T cells

    Jørgensen, A; Wiencke, A K; la Cour, M;

    1998-01-01

    apoptosis was detected by 7-amino-actinomycin D and annexin V staining. RESULTS: Retinal pigment epithelial cells expressed FasL and induced apoptosis in activated Fas+ T cells. Blocking of Fas-FasL interaction with antibody strongly inhibited RPE-mediated T-cell apoptosis. Retinal pigment epithelial cells...... human retinal pigment epithelial (RPE) cells can induce apoptosis in activated T cells. METHODS: Fas ligand (FasL) expression was detected by flow cytometry and immunohistochemistry. Cultured RPE cells were cocultured with T-cell lines and peripheral blood lymphocytes for 6 hours to 2 days. Induction of...... induced apoptosis in several activated T-cell populations and T-cell lines, including T-cell antigen receptor (TCR)-CD3-negative T-cell lines. In contrast, RPE cells induced little or no apoptosis in resting peripheral T cells. Major histocompatibility complex (MHC) class II monoclonal antibodies, which...

  11. CD4+ and CD8+ T cell activation are associated with HIV DNA in resting CD4+ T cells.

    Leslie R Cockerham

    Full Text Available The association between the host immune environment and the size of the HIV reservoir during effective antiretroviral therapy is not clear. Progress has also been limited by the lack of a well-accepted assay for quantifying HIV during therapy. We examined the association between multiple measurements of HIV and T cell activation (as defined by markers including CD38, HLA-DR, CCR5 and PD-1 in 30 antiretroviral-treated HIV-infected adults. We found a consistent association between the frequency of CD4+ and CD8+ T cells expressing HLA-DR and the frequency of resting CD4+ T cells containing HIV DNA. This study highlights the need to further examine this relationship and to better characterize the biology of markers commonly used in HIV studies. These results may also have implications for reactivation strategies.

  12. Myeloid-derived Suppressor Cells Inhibit T Cell Activation by Depleting Cystine and Cysteine

    Minu K Srivastava; Sinha, Pratima; Clements, Virginia K.; Rodriguez, Paulo; Ostrand-Rosenberg, Suzanne

    2009-01-01

    Myeloid-derived suppressor cells (MDSC) are present in most cancer patients and are potent inhibitors of T-cell-mediated anti-tumor immunity. Their inhibitory activity is attributed to production of arginase, reactive oxygen species, inducible nitric oxide synthase, and IL-10. We now report that MDSC also block T cell activation by sequestering cystine and limiting the availability of cysteine. Cysteine is an essential amino acid for T cell activation because T cells lack cystathionase, which...

  13. CD4 T cell activation and disease activity at onset of multiple sclerosis

    Jensen, J; Langkilde, Annika Reynberg; Fenst, C; Nicolaisen, M. S.; Roed, H. G.; Christensen, M; Sellebjerg, F

    We studied CD4 T cell activation in patients with clinically isolated syndromes (CIS) suggesting an initial attack of multiple sclerosis. The percentage of blood CD26+ CD4 T cells was increased in these patients, and correlated with magnetic resonance imaging disease activity and clinical disease...

  14. Human retinal pigment epithelial cells inhibit proliferation and IL2R expression of activated T cells

    Kaestel, Charlotte G; Jørgensen, Annette; Nielsen, Mette;

    2002-01-01

    The purpose of this study was to characterize the effects of human retinal pigment epithelial (RPE) cells on activated T cells. Activated T cells were cocultured with adult and foetal human RPE cells whereafter apoptosis and proliferation were determined by flow cytometry and (3)H...... addition to use of TCR negative T cell lines. The expression of IL2R-alpha -beta and -gamma chains of activated T cells was analysed by flow cytometry after incubation of T cells alone or with RPE cells. Human RPE cells were found to inhibit the proliferation of activated T cells by a cell contact......-dependent mechanism. The RPE cells inhibitory abilities were not affected by blocking of any of the tested surface molecules. The inhibition of the T cells' proliferation correlates with a decreased expression of IL2R-beta and -gamma chains. The T cells regain their ability to proliferate and increase their IL2R...

  15. Imaging and Analysis of OT1 T Cell Activation on Lipid Bilayers

    sprotocols

    2015-01-01

    Authors: Peter Beemiller, Jordan Jacobelli & Matthew Krummel ### Abstract Supported lipid bilayers are frequently used to study cell membrane protein dynamics during immune synapse formation by T cells. Here we describe methods for the imaging and analysis of OT1+ T cell activation and T-cell receptor (TCR) dynamics on lipid bilayers. ### Introduction T cells are activated at immune synapses when TCRs bind agonist ligands on antigen presenting cells (APCs). Glass cover...

  16. Activated human T cells secrete exosomes that participate in IL-2 mediated immune response signaling.

    Jessica Wahlgren

    Full Text Available It has previously been shown that nano-meter sized vesicles (30-100 nm, exosomes, secreted by antigen presenting cells can induce T cell responses thus showing the potential of exosomes to be used as immunological tools. Additionally, activated CD3⁺ T cells can secrete exosomes that have the ability to modulate different immunological responses. Here, we investigated what effects exosomes originating from activated CD3⁺ T cells have on resting CD3⁺ T cells by studying T cell proliferation, cytokine production and by performing T cell and exosome phenotype characterization. Human exosomes were generated in vitro following CD3⁺ T cell stimulation with anti-CD28, anti-CD3 and IL-2. Our results show that exosomes purified from stimulated CD3⁺ T cells together with IL-2 were able to generate proliferation in autologous resting CD3⁺ T cells. The CD3⁺ T cells stimulated with exosomes together with IL-2 had a higher proportion of CD8⁺ T cells and had a different cytokine profile compared to controls. These results indicate that activated CD3⁺ T cells communicate with resting autologous T cells via exosomes.

  17. Cutting Edge: Engineering Active IKKβ in T Cells Drives Tumor Rejection.

    Evaristo, César; Spranger, Stefani; Barnes, Sarah E; Miller, Michelle L; Molinero, Luciana L; Locke, Frederick L; Gajewski, Thomas F; Alegre, Maria-Luisa

    2016-04-01

    Acquired dysfunction of tumor-reactive T cells is one mechanism by which tumors can evade the immune system. Identifying and correcting pathways that contribute to such dysfunction should enable novel anticancer therapy design. During cancer growth, T cells show reduced NF-κB activity, which is required for tumor rejection. Impaired T cell-intrinsic NF-κB may create a vicious cycle conducive to tumor progression and further T cell dysfunction. We hypothesized that forcing T cell-intrinsic NF-κB activation might break this cycle and induce tumor elimination. NF-κB was activated in T cells by inducing the expression of a constitutively active form of the upstream activator IκB kinase β (IKKβ). T cell-restricted constitutively active IKKβ augmented the frequency of functional tumor-specific CD8(+) T cells and improved tumor control. Transfer of constitutively active IKKβ-transduced T cells also boosted endogenous T cell responses that controlled pre-established tumors. Our results demonstrate that driving T cell-intrinsic NF-κB can result in tumor control, thus identifying a pathway with potential clinical applicability. PMID:26903482

  18. Chemokines: a new dendritic cell signal for T cell activation

    Christoph A Thaiss

    2011-08-01

    Full Text Available Dendritic cells (DCs are the main inducers and regulators of cytotoxic T lymphocyte (CTL responses against viruses and tumors. One checkpoint to avoid misguided CTL activation, which might damage healthy cells of the body, is the necessity for multiple activation signals, involving both antigenic as well as additional signals that reflect the presence of pathogens. DCs provide both signals when activated by ligands of pattern recognition receptors and licensed by helper lymphocytes. Recently, it has been established that such T cell licensing can be facilitated by CD4+ T helper cells (classical licensing or by NKT cells (alternative licensing. Licensing regulates the DC/CTL cross-talk at multiple layers. Direct recruitment of CTLs through chemokines released by licensed DCs has recently emerged as a common theme and has a crucial impact on the efficiency of CTL responses. Here, we discuss recent advances in our understanding of DC licensing for cross-priming and implications for the temporal and spatial regulation underlying this process. Future vaccination strategies will benefit from a deeper insight into the mechanisms that govern CTL activation.

  19. O-GlcNAc signaling is essential for NFAT-mediated transcriptional reprogramming during cardiomyocyte hypertrophy

    Facundo, Heberty T.; Brainard, Robert E.; Watson, Lewis J.; Ngoh, Gladys A.; Hamid, Tariq; Prabhu, Sumanth D.; Jones, Steven P.

    2012-01-01

    The regulation of cardiomyocyte hypertrophy is a complex interplay among many known and unknown processes. One specific pathway involves the phosphatase calcineurin, which regulates nuclear translocation of the essential cardiac hypertrophy transcription factor, nuclear factor of activated T-cells (NFAT). Although metabolic dysregulation is frequently described during cardiac hypertrophy, limited insights exist regarding various accessory pathways. One metabolically derived signal, beta-O-lin...

  20. The antihistamine olopatadine regulates T cell activation in palladium allergy.

    Iguchi, Naohiko; Takeda, Yuri; Sato, Naoki; Ukichi, Kenichirou; Katakura, Akira; Ueda, Kyosuke; Narushima, Takayuki; Higuchi, Shigehito; Ogasawara, Kouetsu

    2016-06-01

    Because of its corrosion resistance palladium (Pd) has been widely used in many consumer products ranging from fashion accessories to dental materials. Recently, however, an increase in Pd allergy cases has been reported. Metal allergy is categorized as a Type IV allergy, which is characterized as a delayed-type hypersensitivity reaction in which T cells are known to play an important role; however, the precise mechanism of their action remains unclear. Here we defined the relationship between histamine and the Pd allergic reaction specifically with respect to T cell responses. To verify the effects of histamine on T cells, we examined whether there is a change in IFN-γ production following stimulation of histamine or the antihistamine, olopatadine hydrochloride (OLP), in vitro. In addition, we assessed whether OLP administration affected the degree of footpad swelling or IFN-γ production during the Pd allergy response in mice. We found that histamine stimulation increased IFN-γ production in T cells, specifically enhancing IFN-γ production in CD8(+) T cells compared with CD4(+) T cells. Interestingly, OLP suppressed the production of IFN-γ in CD8(+) T cells, and this compound inhibited footpad swelling and IFN-γ production in mice with Pd allergy. These results suggest that histamine promotes the Type IV allergic reaction and thus, the histamine 1 receptor (H1R) might be useful therapeutic target for treatment of metal allergy. PMID:27035718

  1. Transcriptomic analysis of mouse EL4 T cells upon T cell activation and in response to protein synthesis inhibition via cycloheximide treatment.

    Lim, Pek Siew; Hardy, Kristine; Peng, Kaiman; Shannon, Frances M

    2016-03-01

    T cell activation involves the recognition of a foreign antigen complexed to the major histocompatibility complex on the antigen presenting T cell to the T cell receptor. This leads to activation of signaling pathways, which ultimately leads to induction of key cytokine genes responsible for eradication of foreign antigens. We used the mouse EL4 T cell as a model system to study genes that are induced as a result of T cell activation using phorbol myristate acetate (PMA) and calcium ionomycin (I) as stimuli. We were also interested to examine the importance of new protein synthesis in regulating the expression of genes involved in T cell activation. Thus we have pre-treated mouse EL4 T cells with cycloheximide, a protein synthesis inhibitor, and left the cells unstimulated or stimulated with PMA/I for 4 h. We performed microarray expression profiling of these cells to correlate the gene expression with chromatin state of T cells upon T cell activation [1]. Here, we detail further information and analysis of the microarray data, which shows that T cell activation leads to differential expression of genes and inducible genes can be further classified as primary and secondary response genes based on their protein synthesis dependency. The data is available in the Gene Expression Omnibus under accession number GSE13278. PMID:26981393

  2. Resistance of CD45RA- T cells to apoptosis and functional impairment, and activation of tumor-antigen specific T cells during radiation therapy of prostate cancer.

    Tabi, Zsuzsanna; Spary, Lisa K; Coleman, Sharon; Clayton, Aled; Mason, Malcolm D; Staffurth, John

    2010-07-15

    The effect of radiation therapy (RT) to the pelvis on circulating T cells was studied in prostate cancer (PCa) patients to provide a baseline for a more informed design of combination radioimmunotherapy. Peripheral blood samples taken from 12 PCa patients with locally advanced tumor before, during, and after hypofractionated RT were analyzed for T cell phenotype and function. There was significantly more loss of naive and early memory compared with more differentiated T cells during RT. The proportions of annexin-V(+) and Fas-expressing T cells were elevated in patients during RT and in PBMC irradiated in vitro ( 2-fold in the presence of an IkappaB-kinase inhibitor, indicating a protective effect via this pathway. T cell proliferation was impaired during RT with IL-2-dependent recovery post-RT. Recall T cell responses to common viral Ags, measured by IFN-gamma production, were little affected by RT. In vitro irradiation of healthy donor PBMCs resulted in a significantly increased frequency of responding T cells, due at least partly to the preferential elimination of CD45RA(+) T cells. Most importantly, antitumor CD4(+) and CD8(+) T cell responses were detectable after, but not before or during RT. The results indicate that generating tumor-specific T cell responses before RT and boosting their activity post-RT are ways likely to amplify the frequency and function of antitumor T cells, with implications for scheduling immunotherapy in PCa. PMID:20548027

  3. Activated human γδ T cells induce peptide-specific CD8+ T-cell responses to tumor-associated self-antigens.

    Altvater, Bianca; Pscherer, Sibylle; Landmeier, Silke; Kailayangiri, Sareetha; Savoldo, Barbara; Juergens, Heribert; Rossig, Claudia

    2012-03-01

    Specific cellular immunotherapy of cancer requires efficient generation and expansion of cytotoxic T lymphocytes (CTLs) that recognize tumor-associated self-antigens. Here, we investigated the capacity of human γδ T cells to induce expansion of CD8+ T cells specific for peptides derived from the weakly immunogenic tumor-associated self-antigens PRAME and STEAP1. Coincubation of aminobisphosphonate-stimulated human peripheral blood-derived γδ T cells (Vγ9+Vδ2+), loaded with HLA-A*02-restricted epitopes of PRAME, with autologous peripheral blood CD8+ T cells stimulated the expansion of peptide-specific cytolytic effector memory T cells. Moreover, peptide-loaded γδ T cells efficiently primed antigen-naive CD45RA+ CD8+ T cells against PRAME peptides. Direct comparisons with mature DCs revealed equal potency of γδ T cells and DCs in inducing primary T-cell responses and peptide-specific T-cell activation and expansion. Antigen presentation by γδ T-APCs was not able to overcome the limited capacity of peptide-specific T cells to interact with targets expressing full-length antigen. Importantly, T cells with regulatory phenotype (CD4+ CD25hiFoxP3+) were lower in cocultures with γδ T cells compared to DCs. In summary, bisphosphonate-activated γδ T cells permit generation of CTLs specific for weakly immunogenic tumor-associated epitopes. Exploiting this strategy for effective immunotherapy of cancer requires strategies that enhance the avidity of CTL responses to allow for efficient targeting of cancer. PMID:21928126

  4. Malignant T cells express lymphotoxin alpha and drive endothelial activation in cutaneous T cell lymphoma

    Lauenborg, Britt; Christensen, Louise; Ralfkiaer, Ulrik;

    2015-01-01

    internal organs and blood. Yet, little is known about the mechanism of the CTCL dissemination. Here, we show that CTCL cells express LTα in situ and that LTα expression is driven by aberrantly activated JAK3/STAT5 pathway. Importantly, via TNF receptor 2, LTα functions as an autocrine factor by stimulating...

  5. 缬沙坦对高糖诱导肾小球足细胞活化T细胞核因子2活化及细胞凋亡的影响%Effects of valsartan on high glucose-induced NFAT2 activation and apoptosis in podocytes

    李锐钊; 章斌; 张丽; 史伟; 梁馨苓; 刘双信; 王文健

    2013-01-01

    AIM To investigate the effects of valsartan on nuclear factor 2 of activated T cells (NFAT2)activation and apoptosis induced by high glucose in podocytes.METHODS The podocytes were treated with various conditions,including different concentrations of glucose (10-40 mmol· L-1),different incubation time (0.5-48 h),valsartan,NFAT2 inhibitor (11 R-VIVI) and NFAT2 activator (ionomycin).The protein levels of NFAT2 in the nucleus of podocytes were detected by western blot and podocyte apoptosis was evaluated by flow cytometry.RESULTS NFAT2 was significantly activated in podocytes when treated with high glucose (20 mmol·L-1) for 2 hours.Ionomycin also increased the expression of NFAT2 in nucleus of podocyte (P > 0.05).NFAT2 nuclear accumulation was significantly inhibited in the podocytes pretreatment with valsartan (0.1 mmol·L-1) or 11R-VIVIT compared with high glucose treated alone groups (P < 0.01),without difference between valsartan and 11R-VIVIT groups (P > 0.05).Apoptosis rate was markedly increased in podocytes treated with high glucose ((21.47 ± 6.24) %) at 48 h compared with that treated with normal glucose ((12.27 ±1.31) %,P < 0.05).High glucose-induced apoptosis was also significantly abrogated by concomitant treatment with valsartan or 11R-VIVIT (P < 0.01),again no significant difference between valsartan and 11R-VIVIT groups (P > 0.05).CONCLUSION Valsartan inhibits podocyte high glucose-induce apoptosis via decreasing nuclear NFAT2 activation.%目的 探讨缬沙坦对高糖诱导肾小球足细胞活化T细胞核因子2 (NFAT2)活化及细胞凋亡的影响.方法 将分化成熟的足细胞分组给予不同糖浓度、不同干预时间、NFAT2抑制剂11R-VIVIT、NFAT2活化剂ionomycin等刺激,采用Western blot检测足细胞胞核中NFAT2的表达水平、流式细胞仪检测足细胞凋亡情况.结果 高糖能刺激足细胞NFAT2活化入核,20 mmol· L-1葡萄糖作用2h时NFAT2活化入核达到高峰;这与Ionomycin作用相似,两者刺激下的NFAT

  6. Towards immunotherapy with redirected T cells in a large animal model: Ex vivo activation, expansion, and genetic modification of canine T cells

    Mata, Melinda; Vera, Juan; Gerken, Claudia; Rooney, Cliona M; Miller, Tasha; Pfent, Catherine; Wang, Lisa L.; Wilson-Robles, Heather M.; Gottschalk, Stephen

    2014-01-01

    Adoptive transfer of T cells expressing chimeric antigen receptors (CARs) has shown promising anti-tumor activity in early phase clinical studies, especially for hematological malignancies. However, most preclinical models do not reliably mimic human disease. We reasoned that developing an adoptive T-cell therapy approach for spontaneous osteosarcoma (OS) occurring in dogs would more closely reproduce the condition in human cancer. To generate CAR-expressing canine T cells we developed expans...

  7. Activation of Human T Cells in Hypertension: Studies of Humanized Mice and Hypertensive Humans.

    Itani, Hana A; McMaster, William G; Saleh, Mohamed A; Nazarewicz, Rafal R; Mikolajczyk, Tomasz P; Kaszuba, Anna M; Konior, Anna; Prejbisz, Aleksander; Januszewicz, Andrzej; Norlander, Allison E; Chen, Wei; Bonami, Rachel H; Marshall, Andrew F; Poffenberger, Greg; Weyand, Cornelia M; Madhur, Meena S; Moore, Daniel J; Harrison, David G; Guzik, Tomasz J

    2016-07-01

    Emerging evidence supports an important role for T cells in the genesis of hypertension. Because this work has predominantly been performed in experimental animals, we sought to determine whether human T cells are activated in hypertension. We used a humanized mouse model in which the murine immune system is replaced by the human immune system. Angiotensin II increased systolic pressure to 162 versus 116 mm Hg for sham-treated animals. Flow cytometry of thoracic lymph nodes, thoracic aorta, and kidney revealed increased infiltration of human leukocytes (CD45(+)) and T lymphocytes (CD3(+) and CD4(+)) in response to angiotensin II infusion. Interestingly, there was also an increase in the memory T cells (CD3(+)/CD45RO(+)) in the aortas and lymph nodes. Prevention of hypertension using hydralazine and hydrochlorothiazide prevented the accumulation of T cells in these tissues. Studies of isolated human T cells and monocytes indicated that angiotensin II had no direct effect on cytokine production by T cells or the ability of dendritic cells to drive T-cell proliferation. We also observed an increase in circulating interleukin-17A producing CD4(+) T cells and both CD4(+) and CD8(+) T cells that produce interferon-γ in hypertensive compared with normotensive humans. Thus, human T cells become activated and invade critical end-organ tissues in response to hypertension in a humanized mouse model. This response likely reflects the hypertensive milieu encountered in vivo and is not a direct effect of the hormone angiotensin II. PMID:27217403

  8. Transcription regulates HIF-1α expression in CD4(+) T cells.

    Bollinger, Thomas; Bollinger, Annalena; Gies, Sydney; Feldhoff, Lea; Solbach, Werner; Rupp, Jan

    2016-01-01

    The transcription factor hypoxia inducible factor-1α (HIF-1α) mediates the metabolic adaptation of cells to hypoxia and T-helper cell fate. However, HIF-1α regulation in CD4(+) T cells (T cells) remains elusive. Here we observed that depletion of oxygen (O2⩽2%) alone was not sufficient to induce HIF-1α expression in T cells. However, when hypoxic T cells were stimulated, HIF-1α was expressed and this was dependent on nuclear factor-κB- and nuclear factor of activated T cell (NFAT)-mediated transcriptional upregulation of Hif-1α mRNA. HIF-1α upregulation could be blocked by drugs inhibiting NF-κB, NFAT or mammalian target of rapamycin precluding CD4(+) T-cell stimulation or translation in T cells, as well as by blocking transcription. CD3, CD28, phorbol-12-myristat-13-acetat (PMA) or ionomycin-stimulated T cells did not express HIF-1α under normoxic conditions. In conclusion, regulation of HIF-1α expression in CD4(+) T cells in hypoxia gravely relies on its transcriptional upregulation and subsequent enhanced protein stabilization. PMID:26150319

  9. Opposing roles for RhoH GTPase during T-cell migration and activation

    Baker, Christina M; Comrie, William A; Hyun, Young-Min; Chung, Hung-Li; Fedorchuk, Christine A; Lim, Kihong; Brakebusch, Cord; McGrath, James L; Waugh, Richard E; Meier-Schellersheim, Martin; Kim, Minsoo

    2012-01-01

    T cells spend the majority of their time perusing lymphoid organs in search of cognate antigen presented by antigen presenting cells (APCs) and then quickly recirculate through the bloodstream to another lymph node. Therefore, regulation of a T-cell response is dependent upon the ability of cells...... activation as well as prolonged T:APC conjugates. RT-PCR analyses of activated CD4(+) T cells and live images of T-cell migration and immunological synapse (IS) formation revealed that functions of RhoH took place primarily at the levels of transcription and intracellular distribution. Thus, we conclude that...

  10. Activated human gammadelta T cells as stimulators of specific CD8+ T-cell responses to subdominant Epstein Barr virus epitopes: potential for immunotherapy of cancer.

    Landmeier, Silke; Altvater, Bianca; Pscherer, Sibylle; Juergens, Heribert; Varnholt, Lena; Hansmeier, Anna; Bollard, Catherine M; Moosmann, Andreas; Bisping, Guido; Rossig, Claudia

    2009-04-01

    The efficacy of current cancer vaccines is limited by the functional heterogeneity and poor availability and expansion of professional antigen-presenting cells (APCs). Besides their potent innate effector properties, gammadelta T cells have been suggested to be involved in the initiation and maintenance of adaptive immune responses. Here, we investigated the capacity of human gammadelta T cells to induce expansion of virus-specific T cells to Epstein Barr virus (EBV) antigens. Aminobisphosphonate-stimulated human peripheral blood-derived gammadelta T cells (Vgamma2+Vdelta2+) acquired a dual phenotype characteristic for both APCs and effector memory T cells. Coincubation of activated gammadelta T cells pulsed with human leukocyte antigen-restricted epitopes of either the highly stimulatory EBV lytic cycle antigen Bam H1 Z fragment leftward open reading frame or the tumor-associated latent EBV antigen latent membrane protein 2a (LMP2a) with autologous peripheral blood lymphocytes induced selective expansion of peptide-specific, fully functional CD3CD8 cytolytic effector memory T cells. Furthermore, gammadelta T APCs efficiently processed and presented endogenous antigen, as demonstrated by the capacity of LMP2a gene-transduced gammadelta T cells to induce expansion of T cells with broad specificity for various LMP2a peptides. The capacity of autologous gammadelta T cells to induce LMP2a-specific autologous cytotoxic T lymphocytes was confirmed in 2 patients with Hodgkin lymphoma. In summary, bisphosphonate-activated human gammadelta T cells stimulate expansion of cytotoxic effector T cells specific for both subdominant and dominant viral epitopes and thus show promise as a novel source of efficient APCs for immunotherapy of viral and malignant disease. PMID:19242369

  11. Activated human γδ T cells as stimulators of specific CD8+ T cell responses to subdominant Epstein Barr virus (EBV) epitopes: Potential for immunotherapy of cancer

    Landmeier, Silke; Altvater, Bianca; Pscherer, Sibylle; Juergens, Heribert; Varnholt, Lena; Hansmeier, Anna; Bollard, Catherine M.; Moosmann, Andreas; Bisping, Guido; Rossig, Claudia

    2011-01-01

    The efficacy of current cancer vaccines is limited by the functional heterogeneity and poor availability and expansion of professional antigen-presenting cells (APCs). Besides their potent innate effector properties, γδ T cells have been suggested to be involved in the initiation and maintenance of adaptive immune responses. Here, we investigated the capacity of human γδ T cells to induce expansion of virus-specific T cells to Epstein Barr virus (EBV) antigens. Aminobisphosphonate-stimulated human peripheral blood-derived γδ T cells (Vγ9+Vδ2+) acquired a dual phenotype characteristic for both APCs and effector memory T cells. Coincubation of activated γδ T cells pulsed with HLA-restricted epitopes of either the highly stimulatory EBV lytic cycle antigen BZLF-1 or the tumor-associated latent EBV antigen LMP2a with autologous peripheral blood lymphocytes induced selective expansion of peptide-specific, fully functional CD3+CD8+ cytolytic effector memory T cells. Furthermore, γδ T-APCs efficiently processed and presented endogenous antigen, as demonstrated by the capacity of LMP2a gene-transduced γδ T cells to induce expansion of T cells with broad specificity for various LMP2a peptides. The capacity of autologous γδ T cells to induce LMP2a-specific autologous CTLs was confirmed in two patients with Hodgkin lymphoma. In summary, bisphosphonate-activated human γδ T cells stimulate expansion of cytotoxic effector T cells specific for both subdominant and dominant viral epitopes and thus show promise as a novel source of efficient APCs for immunotherapy of viral and malignant disease. PMID:19242369

  12. From the cradle to the grave: activities of GATA-3 throughout T cell development and differentiation

    Hosoya, Tomonori; Maillard, Ivan; Engel, James Douglas

    2010-01-01

    GATA family transcription factors play multiple vital roles in hematopoiesis in many cell lineages, and in particular, T cells require GATA-3 for execution of several developmental steps. Transcriptional activation of the Gata3 gene is observed throughout T-cell development and differentiation in stage-specific fashion. GATA-3 has been described as a master regulator of T-helper 2 (Th2) cell differentiation in mature CD4+ T cells. During T-cell development in the thymus, its roles in the CD4 ...

  13. Synergy between the T3/Ti complex and Tp44 in human T cell activation

    Weiss, A.; Manger, B.; Shields, R.; Imboden, J.

    1986-03-05

    Ligands, including antigen, which interact with the T3/T cell antigen receptor (Ti) induce an increase in cytoplasmic free calcium ((Ca/sup + +/)/sub i/). However, such an increase in (Ca/sup + +/)/sub i/ is not sufficient to result in T cell activation. Activation can occur if such ligands, which increase (Ca/sup + +/)/sub i/, are added in the presence of phorbol myristate acetate (PMA). This suggests that interactions involving other T cell surface receptors might function in a manner analogous to PMA and play an important role in T cell activation. Using the human leukemic T cell line Jurkat, they demonstrated that 9.3, a monoclonal antibody specific for a 90 kD homodimer expressed on human T cells (Tp44), synergized with some ligands interacting with T3/Ti on Jurkat in the induction of IL-2 production from Jurkat. Thus, 9.3 synergized with PHA, ConA, or sepharose beads coupled with anti-T3 or anti-Ti but not with calcium ionophores or soluble anti-T3 or anti-Ti. Furthermore, at high concentrations only, 9.3 could also synergize with PMA in the activation of Jurkat and T3/Ti negative mutants of Jurkat. No detectable activation of protein kinase C was induced by 9.3 alone. These studies suggest that T cell activation may result from the simultaneous triggering of several distinct T cell surface molecules.

  14. Inhibition of T cell proliferation by selective block of Ca(2+)-activated K(+) channels

    Jensen, B S; Odum, Niels; Jorgensen, N K;

    1999-01-01

    established. The recent cloning of the Ca(2+)-activated, intermediate-conductance K(+) channel (IK channel) has enabled a detailed investigation of the role of this highly Ca(2+)-sensitive K(+) channel in the calcium signaling and subsequent regulation of T cell proliferation. The role IK channels play in T...... cell activation and proliferation has been investigated by using various blockers of IK channels. The Ca(2+)-activated K(+) current in human T cells is shown by the whole-cell voltage-clamp technique to be highly sensitive to clotrimazole, charybdotoxin, and nitrendipine, but not to ketoconazole...... inhibited after block of IK channels by clotrimazole. Clotrimazole and cyclosporin A act synergistically to inhibit T cell proliferation, which confirms that block of IK channels affects the process downstream from T cell receptor activation. We suggest that IK channels constitute another target for immune...

  15. Peripheral blood T cell activation after radioiodine treatment for Graves' disease

    Wei-Ping Teng; Stark, R.; Borysiewicz, L.K.; Weetman, A.P. (Department of Medicine, University of Cambridge Clinical School, Level 5, Addenbrooke' s Hospital, Cambridge (UK)); Munro, A.J. (Department of Clinical Oncology, Hammersmith Hospital, London (UK)); McHardy Young, S. (Department of Medicine, Central Middlesex Hospital, London (UK))

    1990-01-01

    Radioiodine therapy for Graves' thyrotoxicosis produces a rise in thyroid autoantibodies in the first three months after treatment, but little is known of its effects on T cells. We have therefore followed the changes in T cell subsets in sequential samples from 23 patients with Graves' disease treated with radioiodine, using dualcolour flow cytometry. In the first month after treatment there was a significant rise in activated T cells, identified by the markers HLA-DR(la) and CDw26/Tal (p<0.025 in both cases). CD45RO-positive T cells, which are the primed population containing memory cells, also increased (p<0.025), but there was no change in CD45R-positive, resting T cells or in the CD4 to CD8 (helper to cytotoxic/suppressor) ratio. Vicia villosa-binding T cells, containing the contrasuppressor population, showed a more variable response, but the trend was to an overall increase from pre-treatment values (p<0.025). The changes did not appear to be related to antithyroid drug treatment, since they were seen irrespective of whether patients continued such therapy. These results suggest that T cell activation and enhanced contrasuppressor activity may in part be responsible for the rise in autoantibodies after radioiodine. The T cell changes could also contribute to the worsening of ophthalmopathy seen in some radioiodine-treated patients. (author).

  16. Versatile strategy for controlling the specificity and activity of engineered T cells.

    Ma, Jennifer S Y; Kim, Ji Young; Kazane, Stephanie A; Choi, Sei-Hyun; Yun, Hwa Young; Kim, Min Soo; Rodgers, David T; Pugh, Holly M; Singer, Oded; Sun, Sophie B; Fonslow, Bryan R; Kochenderfer, James N; Wright, Timothy M; Schultz, Peter G; Young, Travis S; Kim, Chan Hyuk; Cao, Yu

    2016-01-26

    The adoptive transfer of autologous T cells engineered to express a chimeric antigen receptor (CAR) has emerged as a promising cancer therapy. Despite impressive clinical efficacy, the general application of current CAR-T--cell therapy is limited by serious treatment-related toxicities. One approach to improve the safety of CAR-T cells involves making their activation and proliferation dependent upon adaptor molecules that mediate formation of the immunological synapse between the target cancer cell and T-cell. Here, we describe the design and synthesis of structurally defined semisynthetic adaptors we refer to as "switch" molecules, in which anti-CD19 and anti-CD22 antibody fragments are site-specifically modified with FITC using genetically encoded noncanonical amino acids. This approach allows the precise control over the geometry and stoichiometry of complex formation between CD19- or CD22-expressing cancer cells and a "universal" anti-FITC-directed CAR-T cell. Optimization of this CAR-switch combination results in potent, dose-dependent in vivo antitumor activity in xenograft models. The advantage of being able to titrate CAR-T-cell in vivo activity was further evidenced by reduced in vivo toxicity and the elimination of persistent B-cell aplasia in immune-competent mice. The ability to control CAR-T cell and cancer cell interactions using intermediate switch molecules may expand the scope of engineered T-cell therapy to solid tumors, as well as indications beyond cancer therapy. PMID:26759368

  17. Bromelain treatment reduces CD25 expression on activated CD4+ T cells in vitro✩

    Secor, Eric R.; Singh, Anurag; Guernsey, Linda A.; McNamara, Jeff T.; Zhan, Lijun; Maulik, Nilanjana; Thrall, Roger S.

    2009-01-01

    Bromelain (Br), an extract from pineapple stem with cysteine protease activity, exerts anti-inflammatory effects in a number of inflammatory models. We have previously shown that Br treatment decreased activated CD4+ T cells and has a therapeutic role in an ovalbumin-induced murine model of allergic airway disease. The current study was designed to determine the effect of Br on CD4+ T cell activation, specifically the expression of CD25 in vitro. CD25 is up regulated upon T cell activation, f...

  18. Surfactant protein A integrates activation signal strength to differentially modulate T cell proliferation.

    Mukherjee, Sambuddho; Giamberardino, Charles; Thomas, Joseph; Evans, Kathy; Goto, Hisatsugu; Ledford, Julie G; Hsia, Bethany; Pastva, Amy M; Wright, Jo Rae

    2012-02-01

    Pulmonary surfactant lipoproteins lower the surface tension at the alveolar-airway interface of the lung and participate in host defense. Previous studies reported that surfactant protein A (SP-A) inhibits lymphocyte proliferation. We hypothesized that SP-A-mediated modulation of T cell activation depends upon the strength, duration, and type of lymphocyte activating signals. Modulation of T cell signal strength imparted by different activating agents ex vivo and in vivo in different mouse models and in vitro with human T cells shows a strong correlation between strength of signal (SoS) and functional effects of SP-A interactions. T cell proliferation is enhanced in the presence of SP-A at low SoS imparted by exogenous mitogens, specific Abs, APCs, or in homeostatic proliferation. Proliferation is inhibited at higher SoS imparted by different doses of the same T cell mitogens or indirect stimuli such as LPS. Importantly, reconstitution with exogenous SP-A into the lungs of SP-A(-/-) mice stimulated with a strong signal also resulted in suppression of T cell proliferation while elevating baseline proliferation in unstimulated T cells. These signal strength and SP-A-dependent effects are mediated by changes in intracellular Ca(2+) levels over time, involving extrinsic Ca(2+)-activated channels late during activation. These effects are intrinsic to the global T cell population and are manifested in vivo in naive as well as memory phenotype T cells. Thus, SP-A appears to integrate signal thresholds to control T cell proliferation. PMID:22219327

  19. Novel function of perforin in negatively regulating CD4+T cell activation by affecting calcium signaling

    Enguang Bi; Kairui Mao; Jia Zou; Yuhan Zheng; Bing Sun; Chunjian Huang; Yu Hu; Xiaodong Wu; Weiwen Deng; Guomei Lin; Zhiduo Liu; Lin Tian; Shuhui Sun

    2009-01-01

    Perforin is a pore-forming protein engaged mainly in mediating target T cell death and is employed by cytotoxic Tlymphocytes (CTLs) and natural killer cells. However, whether it also plays a role in conventional CD4+ T cell func-tion remains unclear. Here we report that in perforin-deficient (PKO) mice, CD4+ T cells are hyperproliferative in response to T cell receptor (TCR) stimulation. This feature of hyperproliferation is accompanied by the enhancement both in cell division and in IL-2 secretion. It seems that the perforin deficiency does not influence T cell development in thymus spleen and lymph node. In vivo, perforin deficiency results in increased antigen-specific T cell prolifera-tion and antibody production. Furthermore, PKO mice are more susceptible to experimental autoimmune uveitis. To address the molecular mechanism, we found that after TCR stimulation, CD44 T cells from PKO mice display an increased intracellular calcium flux and subsequently enhance activation of transcription factor NFATI. Our results indicate that perforin plays a negative role in regulating CD4+ T cell activation and immune response by affecting TCR-dependent Ca2+ signaling.

  20. Early phenotypic activation of circulating helper memory T cells in scleroderma: correlation with disease activity.

    Fiocco, U; Rosada, M; L. Cozzi; Ortolani, C; Silvestro, G.; A. Ruffatti; Cozzi, E; Gallo, C; S. Todesco

    1993-01-01

    OBJECTIVES--The differential expression of several accessory/activation molecules (CD26, CD29, CD45RA, CD25, MLR4, HLA-DR) on peripheral blood CD4+ and CD8+ T lymphocytes in patients with scleroderma was compared with that in controls and patients with other connective systemic diseases to look for evidence of the involvement of T cells in the disease process of scleroderma. METHODS--The two colour expression of surface molecules by circulating T cells was analysed with a panel of monoclonal ...

  1. A global transcriptional view of apoptosis in human T-cell activation

    Windgassen Dirk

    2008-10-01

    Full Text Available Abstract Background T-cell activation is an essential step of immune response. The process of proper T-cell activation is strictly monitored and regulated by apoptosis signaling. Yet, regulation of apoptosis, an integral and crucial facet during the process of T-cell activation, is not well understood. Methods In this study, a Gene-Ontology driven global gene expression analysis coupled with protein abundance and activity assays identified genes and pathways associated with regulation of apoptosis in primary human CD3+ T cells and separately CD4+ and CD8+ T cells. Results We identified significantly regulated apoptotic genes in several protein families, such as BCL2 proteins, CASPASE proteins, and TNF receptors, and detailed their transcriptional kinetics during the T-cell activation process. Transcriptional patterns of a few select genes (BCL2A1, BBC3 and CASP3 were validated at the protein level. Many of these apoptotic genes are involved in NF-κB signaling pathway, including TNFRSF10A, TNFRSF10B, TRAF4, TRAF1, TRAF3, and TRAF6. Upregulation of NF-κB and IκB family genes (REL, RELA, and RELB, NFKBIA, NFKBIE and NFKB1 at 48 to 96 hours, supported by the increase of phosphorylated RELA (p65, suggests that the involvement of the NF-κB complex in the process of T-cell proliferation is not only regulated at the protein level but also at the transcriptional level. Examination of genes involved in MAP kinase signalling pathway, important in apoptosis, suggests an induction of p38 and ERK1 cascades in T-cell proliferation (at 48 to 96 hours, which was explored using phosphorylation assays for p38 (MAPK14 and ERK1 (MAPK3. An immediate and short-lived increase of AP-1 activity measured by DNA-binding activity suggests a rapid and transient activation of p38 and/or JNK cascades upon T-cell activation. Conclusion This comparative genome-scale, transcriptional analysis of T-cell activation in the CD4+ and CD8+ subsets and the mixed CD3+ population

  2. Exhaustion of Activated CD8 T Cells Predicts Disease Progression in Primary HIV-1 Infection.

    Hoffmann, Matthias; Pantazis, Nikos; Martin, Genevieve E; Hickling, Stephen; Hurst, Jacob; Meyerowitz, Jodi; Willberg, Christian B; Robinson, Nicola; Brown, Helen; Fisher, Martin; Kinloch, Sabine; Babiker, Abdel; Weber, Jonathan; Nwokolo, Nneka; Fox, Julie; Fidler, Sarah; Phillips, Rodney; Frater, John

    2016-07-01

    The rate at which HIV-1 infected individuals progress to AIDS is highly variable and impacted by T cell immunity. CD8 T cell inhibitory molecules are up-regulated in HIV-1 infection and associate with immune dysfunction. We evaluated participants (n = 122) recruited to the SPARTAC randomised clinical trial to determine whether CD8 T cell exhaustion markers PD-1, Lag-3 and Tim-3 were associated with immune activation and disease progression. Expression of PD-1, Tim-3, Lag-3 and CD38 on CD8 T cells from the closest pre-therapy time-point to seroconversion was measured by flow cytometry, and correlated with surrogate markers of HIV-1 disease (HIV-1 plasma viral load (pVL) and CD4 T cell count) and the trial endpoint (time to CD4 count approaches. PMID:27415828

  3. 5-Azacytidine Promotes an Inhibitory T-Cell Phenotype and Impairs Immune Mediated Antileukemic Activity

    Thomas Stübig

    2014-01-01

    Full Text Available Demethylating agent, 5-Azacytidine (5-Aza, has been shown to be active in treatment of myeloid malignancies. 5-Aza enhances anticancer immunity, by increasing expression of tumor-associated antigens. However, the impact of 5-Aza immune responses remains poorly understood. Here, T-cell mediated tumor immunity effects of 5-Aza, are investigated in vitro and in vivo. T-cells from healthy donors were treated with 5-Aza and analyzed by qRT-PCR and flow cytometry for changes in gene expression and phenotype. Functionality was assessed by a tumor lysis assay. Peripheral blood from patients treated with 5-Aza after alloSCT was monitored for changes in T-cell subpopulations. 5-Aza treatment resulted in a decrease in CD8+ T-cells, whereas CD4+ T-cells increased. Furthermore, numbers of IFN-γ+ T-helper 1 cells (Th1 were reduced, while Treg-cells showed substantial increase. Additionally, CD8+ T-cells exhibited limited killing capacity against leukemic target cells. In vivo data confirm the increase of Treg compartment, while CD8+ T-effector cell numbers were reduced. 5-Aza treatment results in a shift from cytotoxic to regulatory T-cells with a functional phenotype and a major reduction in proinflammatory Th1-cells, indicating a strong inhibition of tumor-specific T-cell immunity by 5-Aza.

  4. 5-Azacytidine Promotes an Inhibitory T-Cell Phenotype and Impairs Immune Mediated Antileukemic Activity

    Stübig, Thomas; Luetkens, Tim; Hildebrandt, York; Atanackovic, Djordje; Binder, Thomas M. C.; Fehse, Boris; Kröger, Nicolaus

    2014-01-01

    Demethylating agent, 5-Azacytidine (5-Aza), has been shown to be active in treatment of myeloid malignancies. 5-Aza enhances anticancer immunity, by increasing expression of tumor-associated antigens. However, the impact of 5-Aza immune responses remains poorly understood. Here, T-cell mediated tumor immunity effects of 5-Aza, are investigated in vitro and in vivo. T-cells from healthy donors were treated with 5-Aza and analyzed by qRT-PCR and flow cytometry for changes in gene expression and phenotype. Functionality was assessed by a tumor lysis assay. Peripheral blood from patients treated with 5-Aza after alloSCT was monitored for changes in T-cell subpopulations. 5-Aza treatment resulted in a decrease in CD8+ T-cells, whereas CD4+ T-cells increased. Furthermore, numbers of IFN-γ+ T-helper 1 cells (Th1) were reduced, while Treg-cells showed substantial increase. Additionally, CD8+ T-cells exhibited limited killing capacity against leukemic target cells. In vivo data confirm the increase of Treg compartment, while CD8+ T-effector cell numbers were reduced. 5-Aza treatment results in a shift from cytotoxic to regulatory T-cells with a functional phenotype and a major reduction in proinflammatory Th1-cells, indicating a strong inhibition of tumor-specific T-cell immunity by 5-Aza. PMID:24757283

  5. 5-azacytidine promotes an inhibitory T-cell phenotype and impairs immune mediated antileukemic activity.

    Stübig, Thomas; Badbaran, Anita; Luetkens, Tim; Hildebrandt, York; Atanackovic, Djordje; Binder, Thomas M C; Fehse, Boris; Kröger, Nicolaus

    2014-01-01

    Demethylating agent, 5-Azacytidine (5-Aza), has been shown to be active in treatment of myeloid malignancies. 5-Aza enhances anticancer immunity, by increasing expression of tumor-associated antigens. However, the impact of 5-Aza immune responses remains poorly understood. Here, T-cell mediated tumor immunity effects of 5-Aza, are investigated in vitro and in vivo. T-cells from healthy donors were treated with 5-Aza and analyzed by qRT-PCR and flow cytometry for changes in gene expression and phenotype. Functionality was assessed by a tumor lysis assay. Peripheral blood from patients treated with 5-Aza after alloSCT was monitored for changes in T-cell subpopulations. 5-Aza treatment resulted in a decrease in CD8+ T-cells, whereas CD4+ T-cells increased. Furthermore, numbers of IFN-γ + T-helper 1 cells (Th1) were reduced, while Treg-cells showed substantial increase. Additionally, CD8+ T-cells exhibited limited killing capacity against leukemic target cells. In vivo data confirm the increase of Treg compartment, while CD8+ T-effector cell numbers were reduced. 5-Aza treatment results in a shift from cytotoxic to regulatory T-cells with a functional phenotype and a major reduction in proinflammatory Th1-cells, indicating a strong inhibition of tumor-specific T-cell immunity by 5-Aza. PMID:24757283

  6. Surfactant Protein A integrates activation signal strength to differentially modulate T cell proliferation

    Mukherjee, Sambuddho; Giamberardino, Charles; Thomas, Joseph; Evans, Kathy; GOTO, HISATSUGU; Ledford, Julie G.; Hsia, Bethany; Pastva, Amy M.; Wright, Jo Rae

    2012-01-01

    Pulmonary surfactant lipoproteins lower the surface tension at the alveolar:airway interface of the lung and participate in host defense. Previous studies reported that surfactant protein A (SP-A) inhibits lymphocyte proliferation. We hypothesized that SP-A mediated modulation of T cell activation depends upon the strength, duration and type of lymphocyte activating signals. Modulation of T cell signal strength imparted by different activating agents ex and in vivo in different mouse models, ...

  7. Correlated expression of T cell growth factor dependence, sensitivity to Vicia villosa lectin, and cytolytic activity in hybrids between cytolytic T cells and T lymphomas

    1982-01-01

    Somatic cell fusion between cytolytically active, T cell growth factor- (TCGF) dependent murine T cell lines (CTL lines) and noncytolytic, TCGF- independent murine T lymphoma lines has yielded two types of somatic cell hybrids (5): cytolytic hybrids, growth of which is dependent on TCGF, and hybrids with very weak or undetectable cytolytic activity which grow at the same rate with or without TCGF. Here we report that the former can produce stable variants that resemble the latter type. Some o...

  8. Dendritic cell, monocyte and T cell activation and response to glatiramer acetate in multiple sclerosis

    Sellebjerg, F; Hesse, D; Limborg, S;

    2012-01-01

    patients, the latter followed prospectively for one year. Gd-enhanced magnetic resonance imaging (MRI) studies were conducted in all patients. Disease activity was assessed as relapses. Results: The median percentage of DCs expressing CD40 was 10% in untreated MS patients and 5.9% in GA-treated patients...... associated with regulatory, naïve or central memory T cell populations, but CD4+ T cell activation was not related with relapse risk. Conclusions: MS patients treated with GA show prominent changes in circulating antigen-presenting cells and CD4+ T cells. Expression of CD40 on DCs is significantly lower and...

  9. Alternative splicing of MALT1 controls signalling and activation of CD4+ T cells

    Meininger, Isabel; Griesbach, Richard A.; Hu, Desheng; Gehring, Torben; Seeholzer, Thomas; Bertossi, Arianna; Kranich, Jan; Oeckinghaus, Andrea; Eitelhuber, Andrea C; Greczmiel, Ute; Gewies, Andreas; Schmidt-Supprian, Marc; Ruland, Jürgen; Brocker, Thomas; Heissmeyer, Vigo

    2016-01-01

    MALT1 channels proximal T-cell receptor (TCR) signalling to downstream signalling pathways. With MALT1A and MALT1B two conserved splice variants exist and we demonstrate here that MALT1 alternative splicing supports optimal T-cell activation. Inclusion of exon7 in MALT1A facilitates the recruitment of TRAF6, which augments MALT1 scaffolding function, but not protease activity. Naive CD4+ T cells express almost exclusively MALT1B and MALT1A expression is induced by TCR stimulation. We identify...

  10. Multiple mechanisms regulate c-myc gene expression during normal T cell activation.

    Lindsten, T; June, C H; Thompson, C. B.

    1988-01-01

    Quiescent normal human T cells express low levels of steady-state c-myc mRNA as a result of low constitutive promoter utilization, a block to transcriptional elongation within the gene, and rapid degradation of c-myc mRNA in the cytoplasm. Following the activation of the T cell receptor (TCR)/CD3 complex, quiescent T cells are induced to express c-myc mRNA. Two intracellular pathways, one involving protein kinase C activation and the other mediated by increased intracellular calcium concentra...

  11. Activated T cells sustain myeloid-derived suppressor cell-mediated immune suppression

    Damuzzo, Vera; Francescato, Samuela; Pozzuoli, Assunta; Berizzi, Antonio; Mocellin, Simone; Rossi, Carlo Riccardo; Bronte, Vincenzo; Mandruzzato, Susanna

    2016-01-01

    The expansion of myeloid derived suppressor cells (MDSCs), a suppressive population able to hamper the immune response against cancer, correlates with tumor progression and overall survival in several cancer types. We have previously shown that MDSCs can be induced in vitro from precursors present in the bone marrow and observed that these cells are able to actively proliferate in the presence of activated T cells, whose activation level is critical to drive the suppressive activity of MDSCs. Here we investigated at molecular level the mechanisms involved in the interplay between MDSCs and activated T cells. We found that activated T cells secrete IL-10 following interaction with MDSCs which, in turn, activates STAT3 phosphorylation on MDSCs then leading to B7-H1 expression. We also demonstrated that B7-H1+ MDSCs are responsible for immune suppression through a mechanism involving ARG-1 and IDO expression. Finally, we show that the expression of ligands B7-H1 and MHC class II both on in vitro-induced MDSCs and on MDSCs in the tumor microenvironment of cancer patients is paralleled by an increased expression of their respective receptors PD-1 and LAG-3 on T cells, two inhibitory molecules associated with T cell dysfunction. These findings highlight key molecules and interactions responsible for the extensive cross-talk between MDSCs and activated T cells that are at the basis of immune suppression. PMID:26700461

  12. Malignant T cells exhibit CD45 resistant Stat3 activation and proliferation in cutaneous

    Krejsgaard, Thorbjørn Frej; Helvad, Rikke; Ralfkiaer, Elisabeth;

    2010-01-01

    transcription (Stat) activation and cytokine-induced proliferation in lymphocytes. Consequently, CD45 dysregulation could be implicated in aberrant Jak/Stat activation and proliferation in lymphoproliferative diseases. Despite high expression of the CD45 ligand, Galectin-1, in skin lesions from cutaneous T-cell...... lymphoma (CTCL), the malignant T cells exhibit constitutive activation of the Jak3/Stat3 signalling pathway and uncontrolled proliferation. We show that CD45 expression is down-regulated on malignant T cells when compared to non-malignant T cells established from CTCL skin lesions. Moreover, CD45 cross......CD45 is a protein tyrosine phosphatase, which is well-known for regulating antigen receptor signalling in T and B cells via its effect on Src kinases. It has recently been shown that CD45 can also dephosphorylate Janus kinases (Jaks) and thereby regulate Signal transducer and activator of...

  13. Unexpected T cell regulatory activity of anti-histone H1 autoantibody: Its mode of action in regulatory T cell-dependent and -independent manners

    Highlights: ► Anti-histone H1 autoantibody (anti-H1) acts on T cells to inhibit their activation. ► Anti-H1 suppresses T cell activation in Treg cell-dependent and -independent manners. ► Suboptimal dose of anti-H1 enhances suppressor function of Treg cells. ► High dose of anti-H1 directly inhibits T cell receptor signaling. -- Abstract: Induction of anti-nuclear antibodies against DNA or histones is a hallmark of autoimmune disorders, but their actual contribution to disease predisposition remains to be clarified. We have previously reported that autoantibodies against histone H1 work as a critical graft survival factor in a rat model of tolerogeneic liver transplantation. Here we show that an immunosuppressive anti-histone H1 monoclonal antibody (anti-H1 mAb) acts directly on T cells to inhibit their activation in response to T cell receptor (TCR) ligation. Intriguingly, the T cell activation inhibitory activity of anti-H1 mAb under suboptimal dosages required regulatory T (Treg) cells, while high dose stimulation with anti-H1 mAb triggered a Treg cell-independent, direct negative regulation of T cell activation upon TCR cross-linking. In the Treg cell-dependent mode of immunosuppressive action, anti-H1 mAb did not induce the expansion of CD4+Foxp3+ Treg cells, but rather potentiated their regulatory capacity. These results reveal a previously unappreciated T cell regulatory role of anti-H1 autoantibody, whose overproduction is generally thought to be pathogenic in the autoimmune settings

  14. Unexpected T cell regulatory activity of anti-histone H1 autoantibody: Its mode of action in regulatory T cell-dependent and -independent manners

    Takaoka, Yuki [Department of Molecular Biotechnology, Graduate School of Advanced Sciences of Matter, Hiroshima University, Higashi-Hiroshima (Japan); Kawamoto, Seiji, E-mail: skawa@hiroshima-u.ac.jp [Department of Molecular Biotechnology, Graduate School of Advanced Sciences of Matter, Hiroshima University, Higashi-Hiroshima (Japan); Katayama, Akiko [Department of Molecular Biotechnology, Graduate School of Advanced Sciences of Matter, Hiroshima University, Higashi-Hiroshima (Japan); Nakano, Toshiaki [Liver Transplantation Program, Chang Gung Memorial Hospital-Kaohsiung Medical Center, Chang Gung University College of Medicine, Kaohsiung, Taiwan (China); Yamanaka, Yasushi; Takahashi, Miki [Department of Molecular Biotechnology, Graduate School of Advanced Sciences of Matter, Hiroshima University, Higashi-Hiroshima (Japan); Shimada, Yayoi; Chiang, Kuei-Chen [Kazusa Institute for Drug Discovery, Josai International University, Kisarazu (Japan); Ohmori, Naoya [Kazusa Institute for Drug Discovery, Josai International University, Kisarazu (Japan); Faculty of Nursing, Josai International University, Togane (Japan); Aki, Tsunehiro [Department of Molecular Biotechnology, Graduate School of Advanced Sciences of Matter, Hiroshima University, Higashi-Hiroshima (Japan); Goto, Takeshi; Sato, Shuji [Kazusa Institute for Drug Discovery, Josai International University, Kisarazu (Japan); Faculty of Nursing, Josai International University, Togane (Japan); Goto, Shigeru [Liver Transplantation Program, Chang Gung Memorial Hospital-Kaohsiung Medical Center, Chang Gung University College of Medicine, Kaohsiung, Taiwan (China); Iwao Hospital, Yufuin (Japan); Chen, Chao-Long [Liver Transplantation Program, Chang Gung Memorial Hospital-Kaohsiung Medical Center, Chang Gung University College of Medicine, Kaohsiung, Taiwan (China); Ono, Kazuhisa [Department of Molecular Biotechnology, Graduate School of Advanced Sciences of Matter, Hiroshima University, Higashi-Hiroshima (Japan)

    2013-02-08

    Highlights: ► Anti-histone H1 autoantibody (anti-H1) acts on T cells to inhibit their activation. ► Anti-H1 suppresses T cell activation in Treg cell-dependent and -independent manners. ► Suboptimal dose of anti-H1 enhances suppressor function of Treg cells. ► High dose of anti-H1 directly inhibits T cell receptor signaling. -- Abstract: Induction of anti-nuclear antibodies against DNA or histones is a hallmark of autoimmune disorders, but their actual contribution to disease predisposition remains to be clarified. We have previously reported that autoantibodies against histone H1 work as a critical graft survival factor in a rat model of tolerogeneic liver transplantation. Here we show that an immunosuppressive anti-histone H1 monoclonal antibody (anti-H1 mAb) acts directly on T cells to inhibit their activation in response to T cell receptor (TCR) ligation. Intriguingly, the T cell activation inhibitory activity of anti-H1 mAb under suboptimal dosages required regulatory T (Treg) cells, while high dose stimulation with anti-H1 mAb triggered a Treg cell-independent, direct negative regulation of T cell activation upon TCR cross-linking. In the Treg cell-dependent mode of immunosuppressive action, anti-H1 mAb did not induce the expansion of CD4{sup +}Foxp3{sup +} Treg cells, but rather potentiated their regulatory capacity. These results reveal a previously unappreciated T cell regulatory role of anti-H1 autoantibody, whose overproduction is generally thought to be pathogenic in the autoimmune settings.

  15. Hepatitis C virus core protein regulates p300/CBP co-activation function. Possible role in the regulation of NF-AT1 transcriptional activity

    Hepatitis C virus (HCV) core is a viral structural protein; it also participates in some cellular processes, including transcriptional regulation. However, the mechanisms of core-mediated transcriptional regulation remain poorly understood. Oncogenic virus proteins often target p300/CBP, a known co-activator of a wide variety of transcription factors, to regulate the expression of cellular and viral genes. Here we demonstrate, for the first time, that HCV core protein interacts with p300/CBP and enhances both its acetyl-transferase and transcriptional activities. In addition, we demonstrate that nuclear core protein activates the NH2-terminal transcription activation domain (TAD) of NF-AT1 in a p300/CBP-dependent manner. We propose a model in which core protein regulates the co-activation function of p300/CBP and activates NF-AT1, and probably other p300/CBP-regulated transcription factors, by a novel mechanism involving the regulation of the acetylation state of histones and/or components of the transcriptional machinery

  16. Surface markers of cloned human T cells with various cytolytic activities

    1981-01-01

    Human T cells stimulated in secondary allogeneic mixed lymphocyte culture (MLC) were cloned under limiting conditions in microculture systems using T cell growth factor and irradiated allogeneic cells. Clones with lytic activity against either phytohemagglutinin-induced blast cells bearing the stimulating alloantigen(s) (cytotoxic T lymphocyte [CTL] activity), L1210 mouse lymphoma cells coated with rabbit antibody (antibody-dependent cell-mediated cytotoxicity [ADCC]), or K562 human target ce...

  17. Interleukin-13-induced MUC5AC expression is regulated by a PI3K–NFAT3 pathway in mouse tracheal epithelial cells

    Highlights: • IL-13 specifically induced NFAT3 activation in mouse tracheal epithelial cells. • CsA and LY294002 significantly blocked IL-13-induced MUC5AC production. • The PI3K–NFAT3 pathway is positively involved in IL-13-induced MUC5AC production. - Abstract: Interleukin-13 (IL-13) plays a critical role in asthma mucus overproduction, while the mechanisms underlying this process are not fully elucidated. Previous studies showed that nuclear factor of activated T cells (NFAT) is involved in the pathogenesis of asthma, but whether it can directly regulate IL-13-induced mucus (particularly MUC5AC) production is still not clear. Here we showed that IL-13 specifically induced NFAT3 activation through promoting its dephosphorylation in air–liquid interface (ALI) cultures of mouse tracheal epithelial cells (mTECs). Furthermore, both Cyclosporin A (CsA, a specific NFAT inhibitor) and LY294002 (a Phosphoinositide 3-kinase (PI3K) inhibitor) significantly blocked IL-13-induced MUC5AC mRNA and protein production through the inhibition of NFAT3 activity. We also confirmed that CsA could not influence the forkhead Box A2 (Foxa2) and mouse calcium dependent chloride channel 3 (mClca3) expression in IL-13-induced MUC5AC production, which both are known to be important in IL-13-stimulated mucus expression. Our study is the first to demonstrate that the PI3K–NFAT3 pathway is positively involved in IL-13-induced mucus production, and provided novel insights into the molecular mechanism of asthma mucus hypersecretion

  18. Interleukin-13-induced MUC5AC expression is regulated by a PI3K–NFAT3 pathway in mouse tracheal epithelial cells

    Yan, Fugui; Li, Wen; Zhou, Hongbin; Wu, Yinfang; Ying, Songmin; Chen, Zhihua [Department of Respiratory and Critical Care Medicine, Second Affiliated Hospital of Zhejiang University School of Medicine, Hangzhou, Zhejiang (China); Shen, Huahao, E-mail: huahaoshen@163.com [Department of Respiratory and Critical Care Medicine, Second Affiliated Hospital of Zhejiang University School of Medicine, Hangzhou, Zhejiang (China); State Key Lab. of Respiratory Disease (SKLRS) (China)

    2014-03-28

    Highlights: • IL-13 specifically induced NFAT3 activation in mouse tracheal epithelial cells. • CsA and LY294002 significantly blocked IL-13-induced MUC5AC production. • The PI3K–NFAT3 pathway is positively involved in IL-13-induced MUC5AC production. - Abstract: Interleukin-13 (IL-13) plays a critical role in asthma mucus overproduction, while the mechanisms underlying this process are not fully elucidated. Previous studies showed that nuclear factor of activated T cells (NFAT) is involved in the pathogenesis of asthma, but whether it can directly regulate IL-13-induced mucus (particularly MUC5AC) production is still not clear. Here we showed that IL-13 specifically induced NFAT3 activation through promoting its dephosphorylation in air–liquid interface (ALI) cultures of mouse tracheal epithelial cells (mTECs). Furthermore, both Cyclosporin A (CsA, a specific NFAT inhibitor) and LY294002 (a Phosphoinositide 3-kinase (PI3K) inhibitor) significantly blocked IL-13-induced MUC5AC mRNA and protein production through the inhibition of NFAT3 activity. We also confirmed that CsA could not influence the forkhead Box A2 (Foxa2) and mouse calcium dependent chloride channel 3 (mClca3) expression in IL-13-induced MUC5AC production, which both are known to be important in IL-13-stimulated mucus expression. Our study is the first to demonstrate that the PI3K–NFAT3 pathway is positively involved in IL-13-induced mucus production, and provided novel insights into the molecular mechanism of asthma mucus hypersecretion.

  19. Interferon-alpha administration enhances CD8+ T cell activation in HIV infection.

    Maura Manion

    Full Text Available BACKGROUND: Type I interferons play important roles in innate immune defense. In HIV infection, type I interferons may delay disease progression by inhibiting viral replication while at the same time accelerating disease progression by contributing to chronic immune activation. METHODS: To investigate the effects of type I interferons in HIV-infection, we obtained cryopreserved peripheral blood mononuclear cell samples from 10 subjects who participated in AIDS Clinical Trials Group Study 5192, a trial investigating the activity of systemic administration of IFNα for twelve weeks to patients with untreated HIV infection. Using flow cytometry, we examined changes in cell cycle status and expression of activation antigens by circulating T cells and their maturation subsets before, during and after IFNα treatment. RESULTS: The proportion of CD38+HLA-DR+CD8+ T cells increased from a mean of 11.7% at baseline to 24.1% after twelve weeks of interferon treatment (p = 0.006. These frequencies dropped to an average of 20.1% six weeks after the end of treatment. In contrast to CD8+ T cells, the frequencies of activated CD4+ T cells did not change with administration of type I interferon (mean percentage of CD38+DR+ cells = 2.62% at baseline and 2.17% after 12 weeks of interferon therapy. As plasma HIV levels fell with interferon therapy, this was correlated with a "paradoxical" increase in CD8+ T cell activation (p<0.001. CONCLUSION: Administration of type I interferon increased expression of the activation markers CD38 and HLA DR on CD8+ T cells but not on CD4+ T cells of HIV+ persons. These observations suggest that type I interferons may contribute to the high levels of CD8+ T cell activation that occur during HIV infection.

  20. Epigenetic mechanisms, T-cell activation, and CCR5 genetics interact to regulate T-cell expression of CCR5, the major HIV-1 coreceptor.

    Gornalusse, German G; Mummidi, Srinivas; Gaitan, Alvaro A; Jimenez, Fabio; Ramsuran, Veron; Picton, Anabela; Rogers, Kristen; Manoharan, Muthu Saravanan; Avadhanam, Nymisha; Murthy, Krishna K; Martinez, Hernan; Molano Murillo, Angela; Chykarenko, Zoya A; Hutt, Richard; Daskalakis, Demetre; Shostakovich-Koretskaya, Ludmila; Abdool Karim, Salim; Martin, Jeffrey N; Deeks, Steven G; Hecht, Frederick; Sinclair, Elizabeth; Clark, Robert A; Okulicz, Jason; Valentine, Fred T; Martinson, Neil; Tiemessen, Caroline Tanya; Ndung'u, Thumbi; Hunt, Peter W; He, Weijing; Ahuja, Sunil K

    2015-08-25

    T-cell expression levels of CC chemokine receptor 5 (CCR5) are a critical determinant of HIV/AIDS susceptibility, and manifest wide variations (i) between T-cell subsets and among individuals and (ii) in T-cell activation-induced increases in expression levels. We demonstrate that a unifying mechanism for this variation is differences in constitutive and T-cell activation-induced DNA methylation status of CCR5 cis-regulatory regions (cis-regions). Commencing at an evolutionarily conserved CpG (CpG -41), CCR5 cis-regions manifest lower vs. higher methylation in T cells with higher vs. lower CCR5 levels (memory vs. naïve T cells) and in memory T cells with higher vs. lower CCR5 levels. HIV-related and in vitro induced T-cell activation is associated with demethylation of these cis-regions. CCR5 haplotypes associated with increased vs. decreased gene/surface expression levels and HIV/AIDS susceptibility magnify vs. dampen T-cell activation-associated demethylation. Methylation status of CCR5 intron 2 explains a larger proportion of the variation in CCR5 levels than genotype or T-cell activation. The ancestral, protective CCR5-HHA haplotype bears a polymorphism at CpG -41 that is (i) specific to southern Africa, (ii) abrogates binding of the transcription factor CREB1 to this cis-region, and (iii) exhibits a trend for overrepresentation in persons with reduced susceptibility to HIV and disease progression. Genotypes lacking the CCR5-Δ32 mutation but with hypermethylated cis-regions have CCR5 levels similar to genotypes heterozygous for CCR5-Δ32. In HIV-infected individuals, CCR5 cis-regions remain demethylated, despite restoration of CD4+ counts (≥800 cells per mm(3)) with antiretroviral therapy. Thus, methylation content of CCR5 cis-regions is a central epigenetic determinant of T-cell CCR5 levels, and possibly HIV-related outcomes. PMID:26307764

  1. Foxp3 Inhibits HDAC1 Activity to Modulate Gene Expression in Human T cells

    Holmes, Derek; Gao, Jianmei; Su, Lishan

    2011-01-01

    We have previously reported that HIV-1 preferentially infects Foxp3+ Treg cells in vitro and in vivo, and Foxp3 enhances the HIV-1 LTR expression through epigenetic mechanisms in T cells. We report here that histone deacetylase inhibitor (HDACi) failed to further enhance HIV gene expression in FoxP3+ T cells. We discovered that Foxp3 inhibited cellular HDAC activity in T cells, and mutations in the forkhead domain that ablate Foxp3 function also abolished its ability to inhibit HDAC. When co-...

  2. Inhibition of Ly-6A antigen expression prevents T cell activation

    1990-01-01

    Antisense oligonucleotides complementary to the 5' end of the mRNA encoding the Ly-6A protein were used to block the expression of that protein. Using this approach we could inhibit the expression of Ly-6A by 60-80% in antigen-primed lymph node (LN) T cells as well as in the D10 T cell clone. Inhibition of Ly-6 expression resulted in the inability to restimulate in vitro, antigen-primed T cells. It also blocked the activation of normal spleen cells by Con A, monoclonal antibody (mAb) to CD3, ...

  3. Loss of receptor on tuberculin-reactive T-cells marks active pulmonary tuberculosis.

    Mathias Streitz

    Full Text Available BACKGROUND: Tuberculin-specific T-cell responses have low diagnostic specificity in BCG vaccinated populations. While subunit-antigen (e.g. ESAT-6, CFP-10 based tests are useful for diagnosing latent tuberculosis infection, there is no reliable immunological test for active pulmonary tuberculosis. Notably, all existing immunological tuberculosis-tests are based on T-cell response size, whereas the diagnostic potential of T-cell response quality has never been explored. This includes surface marker expression and functionality of mycobacterial antigen specific T-cells. METHODOLOGY/PRINCIPAL FINDINGS: Flow-cytometry was used to examine over-night antigen-stimulated T-cells from tuberculosis patients and controls. Tuberculin and/or the relatively M. tuberculosis specific ESAT-6 protein were used as stimulants. A set of classic surface markers of T-cell naïve/memory differentiation was selected and IFN-gamma production was used to identify T-cells recognizing these antigens. The percentage of tuberculin-specific T-helper-cells lacking the surface receptor CD27, a state associated with advanced differentiation, varied considerably between individuals (from less than 5% to more than 95%. Healthy BCG vaccinated individuals had significantly fewer CD27-negative tuberculin-reactive CD4 T-cells than patients with smear and/or culture positive pulmonary tuberculosis, discriminating these groups with high sensitivity and specificity, whereas individuals with latent tuberculosis infection exhibited levels in between. CONCLUSIONS/SIGNIFICANCE: Smear and/or culture positive pulmonary tuberculosis can be diagnosed by a rapid and reliable immunological test based on the distribution of CD27 expression on peripheral blood tuberculin specific T-cells. This test works very well even in a BCG vaccinated population. It is simple and will be of great utility in situations where sputum specimens are difficult to obtain or sputum-smear is negative. It will also help

  4. Tissue signatures influence the activation of intrahepatic CD8+ T cells against malaria sporozoites

    Alexandre eMorrot

    2014-08-01

    Full Text Available Plasmodium sporozoites and liver stages express antigens that are targeted to the MHC-Class I antigen-processing pathway. After the introduction of Plasmodium sporozoites by Anopheles mosquitoes, bone marrow-derived dendritic cells in skin-draining lymph nodes are the first cells to cross-present parasite antigens and elicit specific CD8+ T cells. One of these antigens is the immunodominant circumsporozoite protein (CSP. The CD8+ T cell-mediated protective immune response against CSP is dependent on the interleukin loop involving IL-4 receptor expression on CD8+ cells and IL-4 secretion by CD4+ T cell helpers. In a few days, these CD8+ T cells re-circulate to secondary lymphoid organs and the liver. In the liver, the hepatic sinusoids are enriched with cells, such as dendritic, sinusoidal endothelial and Kupffer cells, that are able to cross-present MHC class I antigens to intrahepatic CD8+ T cells. Specific CD8+ T cells actively find infected hepatocytes and target intra-cellular parasites through mechanisms that are both Interferon-g-dependent and -independent. Immunity is mediated by CD8+ T effector or effector-memory cells and, when present in high numbers, these cells can provide sterilizing immunity. Human vaccination trials with recombinant formulations or attenuated sporozoites have yet to achieve the high numbers of specific effector T cells that are required for sterilizing immunity. In spite of the limited number of specific CD8+ T cells, attenuated sporozoites provided multiple times by the endovenous route provided a high degree of protective immunity. These observations highlight that CD8+ T cells may be useful for improving antibody-mediated protective immunity to pre-erythrocytic stages of malaria parasites.

  5. HELIGMOSOMOIDES POLYGYRUS BAKERI INFECTION ACTIVATES COLONIC FOXP3+ T CELLS ENHANCING THEIR CAPACITY TO PREVENT COLITIS

    Hang, Long; Blum, Arthur M.; Setiawan, Tommy; Urban, Joseph P.; Stoyanoff, Korynn M.; Weinstock, Joel V.

    2013-01-01

    Helminthic infections protect mice from colitis in murine models of inflammatory bowel disease and also may protect people. Helminths like H. bakeri (Hpb) can induce Tregs. Experiments explored if Hpb infection could protect mice from colitis through activation of colonic Treg and examined mechanisms of action. We showed that Hpb infection increased the number of T cells expressing Foxp3 in the colon. More importantly, Foxp3+/IL10- and Foxp3+/IL10+ T cell subsets isolated from the colon of Hp...

  6. Functionally active T cells land T cell precursors in the thymus of newborn mice irradiated in fetal stage of development

    Mice were irradiated in dose of 2 Gy in 14 or 17 days of gestation. Irradiation retarded the increase of cell number in developing thymuses but in the day of birth the number of thymocytes was normalized. In normal development SC-1+ cells (T cell precursors) disappeared from the thymus immediately before the birth. After the irradiation they persisted in the newborn thymus. Mitogenic responses of newborn thymocytes on the action of thymic peptides and T cell mitogens were decreased after the fetal irradiation (adult irradiation enhances mitogenic response of thymocytes)

  7. Peripheral blood T cell activation after radioiodine treatment for graves' disease

    Radioiodine therapy for Graves' thyrotoxicosis produces a rise in thyroid autoantibodies in the first three months after treatment, but little is known of its effects on T cells. We have therefore followed the changes in T cells subsets in sequential samples from 23 patients with Graves' disease treated with radioiodine, using dual-colour flow cytometry. In the first month after treatment there was a significant rise in activated T cells, identified by the markers HLA-DR (Ia) and CDW 26/Ta1 (P<0.025 in both case). CD45RO-positive T cells, which are the prime population containing memory cells, also increased (P<0.025), but there was no change in CD45R-positive, resting cells or in the CD4/CD8 (helper to cytotoxic/suppressor) ratio. Vicia villosa-binding T cells, containing the contra-suppressor population, showed a more variable response, but the trend was to an overall increase from pre-treatment values (P<0.025). The change did not appear to be related to antithyroid drugs treatment, since they were seen irrespective of whether patients convinced such therapy. These results suggest that T cell activation and enhanced contra-suppressor activity may in part be responsible for the rise in autoantibodies after radioiodine therapy

  8. Angiopoietin 2 stimulates TIE2-expressing monocytes to suppress T cell activation and to promote regulatory T cell expansion.

    Coffelt, Seth B; Chen, Yung-Yi; Muthana, Munitta; Welford, Abigail F; Tal, Andrea O; Scholz, Alexander; Plate, Karl H; Reiss, Yvonne; Murdoch, Craig; De Palma, Michele; Lewis, Claire E

    2011-04-01

    Angiopoietin 2 (ANGPT2) is a proangiogenic cytokine whose expression is often upregulated by endothelial cells in tumors. Expression of its receptor, TIE2, defines a highly proangiogenic subpopulation of myeloid cells in circulation and tumors called TIE2-expressing monocytes/macrophages (TEMs). Genetic depletion of TEMs markedly reduces tumor angiogenesis in various tumor models, emphasizing their essential role in driving tumor progression. Previously, we demonstrated that ANGPT2 augments the expression of various proangiogenic genes, the potent immunosuppressive cytokine, IL-10, and a chemokine for regulatory T cells (Tregs), CCL17 by TEMs in vitro. We now show that TEMs also express higher levels of IL-10 than TIE2(-) macrophages in tumors and that ANGPT2-stimulated release of IL-10 by TEMs suppresses T cell proliferation, increases the ratio of CD4(+) T cells to CD8(+) T cells, and promotes the expansion of CD4(+)CD25(high)FOXP3(+) Tregs. Furthermore, syngeneic murine tumors expressing high levels of ANGPT2 contained not only high numbers of TEMs but also increased numbers of Tregs, whereas genetic depletion of tumor TEMs resulted in a marked reduction in the frequency of Tregs in tumors. Taken together, our data suggest that ANGPT2-stimulated TEMs represent a novel, potent immunosuppressive force in tumors. PMID:21368233

  9. The Dendritic Cell Synapse: A Life Dedicated to T Cell Activation.

    Benvenuti, Federica

    2016-01-01

    T-cell activation within immunological synapses is a complex process whereby different types of signals are transmitted from antigen-presenting cells to T cells. The molecular strategies developed by T cells to interpret and integrate these signals have been systematically dissected in recent years and are now in large part understood. On the other side of the immune synapse, dendritic cells (DCs) participate actively in synapse formation and maintenance by remodeling of membrane receptors and intracellular content. However, the details of such changes have been only partially characterized. The DCs actin cytoskeleton has been one of the first systems to be identified as playing an important role in T-cell priming and some of the underlying mechanisms have been elucidated. Similarly, the DCs microtubule cytoskeleton undergoes major spatial changes during synapse formation that favor polarization of the DCs subcellular space toward the interacting T cell. Recently, we have begun to investigate the trafficking machinery that controls polarized delivery of endosomal vesicles at the DC-T immune synapse with the aim of understanding the functional relevance of polarized secretion of soluble factors during T-cell priming. Here, we will review the current knowledge of events occurring in DCs during synapse formation and discuss the open questions that still remain unanswered. PMID:27014259

  10. Prolonged activation of virus-specific CD8+T cells after acute B19 infection.

    2005-12-01

    Full Text Available BACKGROUND: Human parvovirus B19 (B19 is a ubiquitous and clinically significant pathogen, causing erythema infectiosum, arthropathy, transient aplastic crisis, and intrauterine fetal death. The phenotype of CD8+ T cells in acute B19 infection has not been studied previously. METHODS AND FINDINGS: The number and phenotype of B19-specific CD8+ T cell responses during and after acute adult infection was studied using HLA-peptide multimeric complexes. Surprisingly, these responses increased in magnitude over the first year post-infection despite resolution of clinical symptoms and control of viraemia, with T cell populations specific for individual epitopes comprising up to 4% of CD8+ T cells. B19-specific T cells developed and maintained an activated CD38+ phenotype, with strong expression of perforin and CD57 and downregulation of CD28 and CD27. These cells possessed strong effector function and intact proliferative capacity. Individuals tested many years after infection exhibited lower frequencies of B19-specific cytotoxic T lymphocytes, typically 0.05%-0.5% of CD8+ T cells, which were perforin, CD38, and CCR7 low. CONCLUSION: This is the first example to our knowledge of an "acute" human viral infection inducing a persistent activated CD8+ T cell response. The likely explanation--analogous to that for cytomegalovirus infection--is that this persistent response is due to low-level antigen exposure. CD8+ T cells may contribute to the long-term control of this significant pathogen and should be considered during vaccine development.

  11. Switch-mediated activation and retargeting of CAR-T cells for B-cell malignancies.

    Rodgers, David T; Mazagova, Magdalena; Hampton, Eric N; Cao, Yu; Ramadoss, Nitya S; Hardy, Ian R; Schulman, Andrew; Du, Juanjuan; Wang, Feng; Singer, Oded; Ma, Jennifer; Nunez, Vanessa; Shen, Jiayin; Woods, Ashley K; Wright, Timothy M; Schultz, Peter G; Kim, Chan Hyuk; Young, Travis S

    2016-01-26

    Chimeric antigen receptor T (CAR-T) cell therapy has produced impressive results in clinical trials for B-cell malignancies. However, safety concerns related to the inability to control CAR-T cells once infused into the patient remain a significant challenge. Here we report the engineering of recombinant antibody-based bifunctional switches that consist of a tumor antigen-specific Fab molecule engrafted with a peptide neo-epitope, which is bound exclusively by a peptide-specific switchable CAR-T cell (sCAR-T). The switch redirects the activity of the bio-orthogonal sCAR-T cells through the selective formation of immunological synapses, in which the sCAR-T cell, switch, and target cell interact in a structurally defined and temporally controlled manner. Optimized switches specific for CD19 controlled the activity, tissue-homing, cytokine release, and phenotype of sCAR-T cells in a dose-titratable manner in a Nalm-6 xenograft rodent model of B-cell leukemia. The sCAR-T-cell dosing regimen could be tuned to provide efficacy comparable to the corresponding conventional CART-19, but with lower cytokine levels, thereby offering a method of mitigating cytokine release syndrome in clinical translation. Furthermore, we demonstrate that this methodology is readily adaptable to targeting CD20 on cancer cells using the same sCAR-T cell, suggesting that this approach may be broadly applicable to heterogeneous and resistant tumor populations, as well as other liquid and solid tumor antigens. PMID:26759369

  12. Vaginal immunization to elicit primary T-cell activation and dissemination.

    Elena Pettini

    Full Text Available Primary T-cell activation at mucosal sites is of utmost importance for the development of vaccination strategies. T-cell priming after vaginal immunization, with ovalbumin and CpG oligodeoxynucleotide adjuvant as model vaccine formulation, was studied in vivo in hormone-synchronized mice and compared to the one induced by the nasal route. Twenty-four hours after both vaginal or nasal immunization, antigen-loaded dendritic cells were detected within the respective draining lymph nodes. Vaginal immunization elicited a strong recruitment of antigen-specific CD4(+ T cells into draining lymph nodes that was more rapid than the one observed following nasal immunization. T-cell clonal expansion was first detected in iliac lymph nodes, draining the genital tract, and proliferated T cells disseminated towards distal lymph nodes and spleen similarly to what observed following nasal immunization. T cells were indeed activated by the antigen encounter and acquired homing molecules essential to disseminate towards distal lymphoid organs as confirmed by the modulation of CD45RB, CD69, CD44 and CD62L marker expression. A multi-type Galton Watson branching process, previously used for in vitro analysis of T-cell proliferation, was applied to model in vivo CFSE proliferation data in draining lymph nodes 57 hours following immunization, in order to calculate the probabilistic decision of a cell to enter in division, rest in quiescence or migrate/die. The modelling analysis indicated that the probability of a cell to proliferate was higher following vaginal than nasal immunization. All together these data show that vaginal immunization, despite the absence of an organized mucosal associated inductive site in the genital tract, is very efficient in priming antigen-specific CD4(+ T cells and inducing their dissemination from draining lymph nodes towards distal lymphoid organs.

  13. Vitamin D receptor activation and downregulation of renin-angiotensin system attenuate morphine-induced T cell apoptosis

    Chandel, Nirupama; Sharma, Bipin; Salhan, Divya; Husain, Mohammad; Malhotra, Ashwani; Buch, Shilpa; Singhal, Pravin C.

    2012-01-01

    Opiates have been reported to induce T cell loss. We evaluated the role of vitamin D receptor (VDR) and the activation of the renin-angiotensin system (RAS) in morphine-induced T cell loss. Morphine-treated human T cells displayed downregulation of VDR and the activation of the RAS. On the other hand, a VDR agonist (EB1089) enhanced T cell VDR expression both under basal and morphine-stimulated states. Since T cells with silenced VDR displayed the activation of the RAS, whereas activation of ...

  14. Tob1 plays a critical role in the activation of encephalitogenic T cells in CNS autoimmunity

    Schulze-Topphoff, Ulf; Casazza, Simona; Varrin-Doyer, Michel; Pekarek, Kara; Sobel, Raymond A.; Hauser, Stephen L.; Oksenberg, Jorge R.; Zamvil, Scott S.

    2013-01-01

    Reliable biomarkers corresponding to disease progression or therapeutic responsiveness in multiple sclerosis (MS) have not been yet identified. We previously reported that low expression of the antiproliferative gene TOB1 in CD4+ T cells of individuals presenting with an initial central nervous system (CNS) demyelinating event (a clinically isolated syndrome), correlated with high risk for progression to MS. We report that experimental autoimmune encephalomyelitis (EAE) in Tob1−/− mice was associated with augmented CNS inflammation, increased infiltrating CD4+ and CD8+ T cell counts, and increased myelin-reactive Th1 and Th17 cells, with reduced numbers of regulatory T cells. Reconstitution of Rag1−/− mice with Tob1−/− CD4+ T cells recapitulated the aggressive EAE phenotype observed in Tob1−/− mice. Furthermore, severe spontaneous EAE was observed when Tob1−/− mice were crossed to myelin oligodendrocyte glycoprotein–specific T cell receptor transgenic (2D2) mice. Collectively, our results reveal a critical role for Tob1 in adaptive T cell immune responses that drive development of EAE, thus providing support for the development of Tob1 as a biomarker for demyelinating disease activity. PMID:23797093

  15. Metabolic reprogramming of alloantigen-activated T cells after hematopoietic cell transplantation.

    Nguyen, Hung D; Chatterjee, Shilpak; Haarberg, Kelley M K; Wu, Yongxia; Bastian, David; Heinrichs, Jessica; Fu, Jianing; Daenthanasanmak, Anusara; Schutt, Steven; Shrestha, Sharad; Liu, Chen; Wang, Honglin; Chi, Hongbo; Mehrotra, Shikhar; Yu, Xue-Zhong

    2016-04-01

    Alloreactive donor T cells are the driving force in the induction of graft-versus-host disease (GVHD), yet little is known about T cell metabolism in response to alloantigens after hematopoietic cell transplantation (HCT). Here, we have demonstrated that donor T cells undergo metabolic reprograming after allogeneic HCT. Specifically, we employed a murine allogeneic BM transplant model and determined that T cells switch from fatty acid β-oxidation (FAO) and pyruvate oxidation via the tricarboxylic (TCA) cycle to aerobic glycolysis, thereby increasing dependence upon glutaminolysis and the pentose phosphate pathway. Glycolysis was required for optimal function of alloantigen-activated T cells and induction of GVHD, as inhibition of glycolysis by targeting mTORC1 or 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 3 (PFKFB3) ameliorated GVHD mortality and morbidity. Together, our results indicate that donor T cells use glycolysis as the predominant metabolic process after allogeneic HCT and suggest that glycolysis has potential as a therapeutic target for the control of GVHD. PMID:26950421

  16. CD8+ T cell activation predominate early immune responses to hypercholesterolemia in Apoe-/- mice

    Björkbacka Harry

    2010-12-01

    Full Text Available Abstract Background It is well established that adaptive immune responses induced by hypercholesterolemia play an important role in the development of atherosclerosis, but the pathways involved remain to be fully characterized. In the present study we assessed immune responses to hypercholesterolemia induced by feeding Apoe-/- mice a high-fat diet for 4 or 8 weeks. Results The primary immune response in lymph nodes draining the aortic root was an increased expression of interferon (IFN-γ in CD8+CD28+ T cells, while an activation of IFN-γ expression in CD4+ T cells was observed only after 8 weeks of high-fat diet. Contrarily, spleen CD4+ T cells responded with a higher expression of IL-10. Spleen CD8+ T cells expressed both IFN-γ and IL-10 and showed enhanced proliferation when exposed to Concanavalin A. Plasma levels of IgG and IgM against oxidized LDL did not change, but the level of apolipoprotein B/IgM immune complexes was increased. Conclusion Hypercholesterolemia leads to unopposed activation of Th1 immune responses in lymph nodes draining atherosclerotic lesions, whereas Th1 activation in the spleen is balanced by a concomitant activation of Th2 cells. The activation of CD8+ T cells implies that hypercholesterolemia is associated with formation of cell autoantigens.

  17. T-cell activation promotes tumorigenesis in inflammation-associated cancer

    Lairmore Michael

    2009-12-01

    Full Text Available Abstract Chronic inflammation has long been associated with a wide range of malignancies, is now widely accepted as a risk factor for development of cancer, and has been implicated as a promoter of a variety of cancers including hematopoietic malignancies. We have described a mouse model uniquely suited to examine the link between inflammation and lymphoma in which the Tax oncogene, expressed in activated T and NK cells, perpetuates chronic inflammation that begins as microscopic intraepithelial lesions and develops into inflammatory nodules, subcutaneous tumors, and large granular lymphocytic leukemia. The use of bioluminescent imaging in these mice has expanded our ability to interrogate aspects of inflammation and tumorigenesis non-invasively. Here we demonstrate that bioluminescence induction in these mice correlated with inflammation resulting from wounding, T cell activation, and exposure to chemical agents. In experiments in which long-term effects of inflammation on disease outcome were monitored, the development of lymphoma was promoted by an inflammatory stimulus. Finally we demonstrated that activation of T-cells in T-cell receptor (TCR transgenic TAX-LUC animals dramatically exacerbated the development of subcutaneous TCR- CD16+ LGL tumors. The role of activated T-cells and acquired immunity in inflammation-associated cancers is broadly applicable to hematopoietic malignancies, and we propose these mice will be of use in dissecting mechanisms by which activated T-cells promote lymphomagenesis in vivo.

  18. CIP2A Promotes T-Cell Activation and Immune Response to Listeria monocytogenes Infection

    Cvrljevic, Anna; Khan, Mohd Moin; Treise, Irina; Adler, Thure; Aguilar-Pimentel, Juan Antonio; Au-Yeung, Byron; Sittig, Eleonora; Laajala, Teemu Daniel; Chen, Yiling; Oeder, Sebastian; Calzada-Wack, Julia; Horsch, Marion; Aittokallio, Tero; Busch, Dirk H.; Ollert, Markus W.; Neff, Frauke; Beckers, Johannes; Gailus-Durner, Valerie; Fuchs, Helmut; de Angelis, Martin Hrabě; Chen, Zhi; Lahesmaa, Riitta; Westermarck, Jukka

    2016-01-01

    The oncoprotein Cancerous Inhibitor of Protein Phosphatase 2A (CIP2A) is overexpressed in most malignancies and is an obvious candidate target protein for future cancer therapies. However, the physiological importance of CIP2A-mediated PP2A inhibition is largely unknown. As PP2A regulates immune responses, we investigated the role of CIP2A in normal immune system development and during immune response in vivo. We show that CIP2A-deficient mice (CIP2AHOZ) present a normal immune system development and function in unchallenged conditions. However when challenged with Listeria monocytogenes, CIP2AHOZ mice display an impaired adaptive immune response that is combined with decreased frequency of both CD4+ T-cells and CD8+ effector T-cells. Importantly, the cell autonomous effect of CIP2A deficiency for T-cell activation was confirmed. Induction of CIP2A expression during T-cell activation was dependent on Zap70 activity. Thus, we reveal CIP2A as a hitherto unrecognized mediator of T-cell activation during adaptive immune response. These results also reveal CIP2AHOZ as a possible novel mouse model for studying the role of PP2A activity in immune regulation. On the other hand, the results also indicate that CIP2A targeting cancer therapies would not cause serious immunological side-effects. PMID:27100879

  19. Transcriptional Regulation of TMP21 by NFAT

    Xia Kun

    2011-03-01

    Full Text Available Abstract Background TMP21 is a member of the p24 cargo protein family, which is involved in protein transport between the Golgi apparatus and ER. Alzheimer's Disease (AD is the most common neurodegenerative disorder leading to dementia and deposition of amyloid β protein (Aβ is the pathological feature of AD pathogenesis. Knockdown of TMP21 expression by siRNA causes a sharp increase in Aβ production; however the underlying mechanism by which TMP21 regulates Aβ generation is unknown, and human TMP21 gene expression regulation has not yet been studied. Results In this report we have cloned a 3.3-kb fragment upstream of the human TMP21 gene. The transcription start site (TSS of the human TMP21 gene was identified. A series of nested deletions of the 5' flanking region of the human TMP21 gene were subcloned into the pGL3-basic luciferase reporter plasmid. We identified the -120 to +2 region as containing the minimal sequence necessary for TMP21 gene promoter activity. Gel shift assays revealed that the human TMP21 gene promoter contains NFAT response elements. Expression of NFAT increased TMP21 gene expression and inhibition of NFAT by siRNA reduced TMP21 gene expression. Conclusion NFAT plays a very important role in the regulation of human TMP21 gene expression. This study demonstrates that the human TMP21 gene expression is transcriptionally regulated by NFAT signaling.

  20. Otud7b facilitates T cell activation and inflammatory responses by regulating Zap70 ubiquitination.

    Hu, Hongbo; Wang, Hui; Xiao, Yichuan; Jin, Jin; Chang, Jae-Hoon; Zou, Qiang; Xie, Xiaoping; Cheng, Xuhong; Sun, Shao-Cong

    2016-03-01

    Signal transduction from the T cell receptor (TCR) is crucial for T cell-mediated immune responses and, when deregulated, also contributes to the development of autoimmunity. How TCR signaling is regulated is incompletely understood. In this study, we demonstrate a ubiquitin-dependent mechanism in which the deubiquitinase Otud7b has a crucial role in facilitating TCR signaling. Upon TCR ligation, Otud7b is rapidly recruited to the tyrosine kinase Zap70, a central mediator of TCR-proximal signaling. Otud7b deficiency attenuates the activation of Zap70 and its downstream pathways and impairs T cell activation and differentiation, rendering mice refractory to T cell-mediated autoimmune and inflammatory responses. Otud7b facilitated Zap70 activation by deubiquitinating Zap70, thus preventing the association of Zap70 with the negative-regulatory phosphatases Sts1 and Sts2. These findings establish Otud7b as a positive regulator of TCR-proximal signaling and T cell activation, highlighting the importance of deubiquitination in regulating Zap70 function. PMID:26903241

  1. Activation of a suppressor T-cell pathway by interferon.

    Aune, T M; Pierce, C. W.

    1982-01-01

    In addition to antiviral activities, murine fibroblast (type I) interferon (IFN-beta) suppresses immune responses. The mechanism(s) by which IFN-beta suppresses antibody responses by murine spleen cells to sheep erythrocytes in vitro has been investigated. IFN-beta-mediated suppression is partially or completely prevented by catalase, 2-mercaptoethanol, and certain peroxidase substrates (ascorbic acid, potassium iodide, and tyrosine). These same reagents also block suppression by mediators fr...

  2. T Cells

    T Cells - National Multiple Sclerosis Society Skip to navigation Skip to content Menu Navigation National Multiple Sclerosis Society Sign ... Is MS? Definition of MS T Cells T Cells Share Smaller Text Larger Text Print In this ...

  3. Antigen Presentation and T-Cell Activation Are Critical for RBP4-Induced Insulin Resistance.

    Moraes-Vieira, Pedro M; Castoldi, Angela; Aryal, Pratik; Wellenstein, Kerry; Peroni, Odile D; Kahn, Barbara B

    2016-05-01

    Adipose tissue (AT) inflammation contributes to impaired insulin action, which is a major cause of type 2 diabetes. RBP4 is an adipocyte- and liver-derived protein with an important role in insulin resistance, metabolic syndrome, and AT inflammation. RBP4 elevation causes AT inflammation by activating innate immunity, which elicits an adaptive immune response. RBP4-overexpressing mice (RBP4-Ox) are insulin resistant and glucose intolerant and have increased AT macrophages and T-helper 1 cells. We show that high-fat diet-fed RBP4(-/-) mice have reduced AT inflammation and improved insulin sensitivity versus wild type. We also elucidate the mechanism for RBP4-induced macrophage antigen presentation and subsequent T-cell activation. In RBP4-Ox, AT macrophages display enhanced c-Jun N-terminal kinase, extracellular signal-related kinase, and p38 phosphorylation. Inhibition of these pathways and of NF-κB reduces activation of macrophages and CD4 T cells. MyD88 is an adaptor protein involved in proinflammatory signaling. In macrophages from MyD88(-/-) mice, RBP4 fails to stimulate secretion of tumor necrosis factor, IL-12, and IL-6 and CD4 T-cell activation. In vivo blockade of antigen presentation by treating RBP4-Ox mice with CTLA4-Ig, which blocks costimulation of T cells, is sufficient to reduce AT inflammation and improve insulin resistance. Thus, MyD88 and downstream mitogen-activated protein kinase and NF-κB pathways are necessary for RBP4-induced macrophage antigen presentation and subsequent T-cell activation. Also, blocking antigen presentation with CTLA4-Ig improves RBP4-induced insulin resistance and macrophage-induced T-cell activation. PMID:26936962

  4. Downregulation of CD4+CD25+ regulatory T cells may underlie enhanced Th1 immunity caused by immunization with activated autologous T cells

    Qi Cao; Dangsheng Li; Ningli Li; Li Wang; Fang Du; Huiming Sheng; Yan Zhang; Juanjuan Wu; Baihua Shen; Tianwei Shen; Jingwu Zhang

    2007-01-01

    Regulatory T cells (Treg) play important roles in immune system homeostasis, and may also be involved in tumor immunotolerance by suppressing Thl immune response which is involved in anti-tumor immunity. We have previously reported that immunization with attenuated activated autologous T cells leads to enhanced anti-tumor immunity and upregulated Thl responses in vivo. However, the underlying molecular mechanisms are not well understood. Here we show that Treg function was significantly downregulated in mice that received immunization of attenuated activated autologous T cells. We found that Foxp3 expression decreased in CD4+CD25+ T cells from the immunized mice. Moreover, CD4+CD25+Foxp3+ Treg obtained from immunized mice exhibited diminished immunosuppression ability compared to those from naive mice. Further analysis showed that the serum of immunized mice contains a high level of anti-CD25 antibody (about 30 ng/ml,/K0.01 vs controls). Consistent with a role of anti-CD25 response in the down-regulation of Treg, adoptive transfer of serum from immunized mice to naive mice led to a significant decrease in Treg population and function in recipient mice. The triggering of anti-CD25 response in immunized mice can be explained by the fact that CD25 was induced to a high level in the ConA activated autologous T cells used for immunization. Our results demonstrate for the first time that immunization with attenuated activated autologous T cells evokes anti-CD25 antibody production, which leads to impeded CD4+CD25+Foxp3+ Treg expansion and function in vivo. We suggest that dampened Treg function likely contributes to enhanced Thl response in immunized mice and is at least part of the mechanism underlying the boosted anti-tumor immunity.

  5. Small molecule inhibition of 6-phosphofructo-2-kinase suppresses t cell activation

    Telang Sucheta

    2012-05-01

    Full Text Available Abstract Background T cell activation is associated with a rapid increase in intracellular fructose-2,6-bisphosphate (F2,6BP, an allosteric activator of the glycolytic enzyme, 6-phosphofructo-1-kinase. The steady state concentration of F2,6BP in T cells is dependent on the expression of the bifunctional 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatases (PFKFB1-4 and the fructose-2,6-bisphosphatase, TIGAR. Of the PFKFB family of enzymes, PFKFB3 has the highest kinase:bisphosphatase ratio and has been demonstrated to be required for T cell proliferation. A small molecule antagonist of PFKFB3, 3-(3-pyridinyl-1-(4-pyridinyl-2-propen-1-one (3PO, recently has been shown to reduce F2,6BP synthesis, glucose uptake and proliferation in transformed cells. We hypothesized that the induction of PFKFB3 expression may be required for the stimulation of glycolysis in T cells and that exposure to the PFKFB3 antagonist, 3PO, would suppress T cell activation. Methods We examined PFKFB1-4 and TIGAR expression and F2,6BP concentration in purified CD3+ T cells stimulated with microbead-conjugated agonist antibodies specific for CD3 and the co-stimulatory receptor, CD28. We then determined the effect of 3PO on anti-CD3/anti-CD28-induced T cell activation, F2,6BP synthesis, 2-[1-14C]-deoxy-d-glucose uptake, lactate secretion, TNF-α secretion and proliferation. Finally, we examined the effect of 3PO administration on the development of delayed type hypersensitivity to methylated BSA and on imiquimod-induced psoriasis in mice. Results We found that purified human CD3+ T cells express PFKFB2, PFKFB3, PFKFB4 and TIGAR, and that anti-CD3/anti-CD28 conjugated microbeads stimulated a >20-fold increase in F2,6BP with a coincident increase in protein expression of the PFKFB3 family member and a decrease in TIGAR protein expression. We then found that exposure to the PFKFB3 small molecule antagonist, 3PO (1–10 μM, markedly attenuated the stimulation of F2,6BP

  6. Expression of CD11c Is Associated with Unconventional Activated T Cell Subsets with High Migratory Potential

    Cantero, Jon; Tarrats, Antoni; Fernández, Marco Antonio; Sumoy, Lauro; Rodolosse, Annie; McSorley, Stephen J.

    2016-01-01

    CD11c is an α integrin classically employed to define myeloid dendritic cells. Although there is little information about CD11c expression on human T cells, mouse models have shown an association of CD11c expression with functionally relevant T cell subsets. In the context of genital tract infection, we have previously observed increased expression of CD11c in circulating T cells from mice and women. Microarray analyses of activated effector T cells expressing CD11c derived from naïve mice demonstrated enrichment for natural killer (NK) associated genes. Here we find that murine CD11c+ T cells analyzed by flow cytometry display markers associated with non-conventional T cell subsets, including γδ T cells and invariant natural killer T (iNKT) cells. However, in women, only γδ T cells and CD8+ T cells were enriched within the CD11c fraction of blood and cervical tissue. These CD11c+ cells were highly activated and had greater interferon (IFN)-γ secretory capacity than CD11c- T cells. Furthermore, circulating CD11c+ T cells were associated with the expression of multiple adhesion molecules in women, suggesting that these cells have high tissue homing potential. These data suggest that CD11c expression distinguishes a population of circulating T cells during bacterial infection with innate capacity and mucosal homing potential. PMID:27119555

  7. Preferential killing of cancer cells and activated human T cells using ZnO nanoparticles

    Nanoparticles are increasingly being recognized for their potential utility in biological applications including nanomedicine. Here we examine the response of normal human cells to ZnO nanoparticles under different signaling environments and compare it to the response of cancerous cells. ZnO nanoparticles exhibit a strong preferential ability to kill cancerous T cells (∼28-35 x) compared to normal cells. Interestingly, the activation state of the cell contributes toward nanoparticle toxicity, as resting T cells display a relative resistance while cells stimulated through the T cell receptor and CD28 costimulatory pathway show greater toxicity in direct relation to the level of activation. Mechanisms of toxicity appear to involve the generation of reactive oxygen species, with cancerous T cells producing higher inducible levels than normal T cells. In addition, nanoparticles were found to induce apoptosis and the inhibition of reactive oxygen species was found to be protective against nanoparticle induced cell death. The novel findings of cell selective toxicity, towards potential disease causing cells, indicate a potential utility of ZnO nanoparticles in the treatment of cancer and/or autoimmunity

  8. Preferential killing of cancer cells and activated human T cells using ZnO nanoparticles

    Hanley, Cory; Layne, Janet; Feris, Kevin; Wingett, Denise [Department of Biological Sciences, Boise State University, Boise, ID 83725 (United States); Punnoose, Alex; Reddy, K M; Coombs, Isaac; Coombs, Andrew [Department of Physics, Boise State University, Boise, ID 83725 (United States)], E-mail: denisewingett@boisestate.edu

    2008-07-23

    Nanoparticles are increasingly being recognized for their potential utility in biological applications including nanomedicine. Here we examine the response of normal human cells to ZnO nanoparticles under different signaling environments and compare it to the response of cancerous cells. ZnO nanoparticles exhibit a strong preferential ability to kill cancerous T cells ({approx}28-35 x) compared to normal cells. Interestingly, the activation state of the cell contributes toward nanoparticle toxicity, as resting T cells display a relative resistance while cells stimulated through the T cell receptor and CD28 costimulatory pathway show greater toxicity in direct relation to the level of activation. Mechanisms of toxicity appear to involve the generation of reactive oxygen species, with cancerous T cells producing higher inducible levels than normal T cells. In addition, nanoparticles were found to induce apoptosis and the inhibition of reactive oxygen species was found to be protective against nanoparticle induced cell death. The novel findings of cell selective toxicity, towards potential disease causing cells, indicate a potential utility of ZnO nanoparticles in the treatment of cancer and/or autoimmunity.

  9. Asynchronous combinatorial action of four regulatory factors activates Bcl11b for T cell commitment.

    Kueh, Hao Yuan; Yui, Mary A; Ng, Kenneth K H; Pease, Shirley S; Zhang, Jingli A; Damle, Sagar S; Freedman, George; Siu, Sharmayne; Bernstein, Irwin D; Elowitz, Michael B; Rothenberg, Ellen V

    2016-08-01

    During T cell development, multipotent progenitors relinquish competence for other fates and commit to the T cell lineage by turning on Bcl11b, which encodes a transcription factor. To clarify lineage commitment mechanisms, we followed developing T cells at the single-cell level using Bcl11b knock-in fluorescent reporter mice. Notch signaling and Notch-activated transcription factors collaborate to activate Bcl11b expression irrespectively of Notch-dependent proliferation. These inputs work via three distinct, asynchronous mechanisms: an early locus 'poising' function dependent on TCF-1 and GATA-3, a stochastic-permissivity function dependent on Notch signaling, and a separate amplitude-control function dependent on Runx1, a factor already present in multipotent progenitors. Despite their necessity for Bcl11b expression, these inputs act in a stage-specific manner, providing a multitiered mechanism for developmental gene regulation. PMID:27376470

  10. Activation of resting human T cells requires prolonged stimulation of protein kinase C

    Berry, N.; Ase, K.; Kishimoto, A.; Nishizuka, Y. (Kobe Univ. School of Medicine (Japan))

    1990-03-01

    Purified resting human T cells can be induced to express the {alpha} subunit of the interleukin 2 receptor and to proliferate by treatment with 12-0-tetradecanoylphorbol-13-acetate plus ionomycin but not with 1,2-dioctanoylglycerol plus ionomycin. Determination of the translocation of protein kinase C showed that 12-0-tetradecanoylphorbol-13-acetate plus ionomycin caused a prolonged membrane association of the enzyme for more than 4 hr, whereas 1,2-dioctanoylglycerol plus ionomycin induced a transient membrane association, which was maximal at 20 min. Delivery of multiple additions of 1,2-dioctanoylglycerol plus ionomycin to the T cells resulted in progressively increased expression of the {alpha} subunit of the interleukin 2 receptor and proliferation commensurate with the number of multiple additions delivered, suggesting that prolonged protein kinase C activity is required for T-cell activation.

  11. Deficient regulatory T cell activity and low frequency of IL-17-producing T cells correlate with the extent of cardiomyopathy in human Chagas' disease.

    Paulo Marcos Matta Guedes

    Full Text Available BACKGROUND: Myocardium damage during Chagas' disease results from the immunological imbalance between pro- and production of anti-inflammatory cytokines and has been explained based on the Th1-Th2 dichotomy and regulatory T cell activity. Recently, we demonstrated that IL-17 produced during experimental T. cruzi infection regulates Th1 cells differentiation and parasite induced myocarditis. Here, we investigated the role of IL-17 and regulatory T cell during human Chagas' disease. METHODOLOGY/PRINCIPAL FINDINGS: First, we observed CD4(+IL-17(+ T cells in culture of peripheral blood mononuclear cells (PBMC from Chagas' disease patients and we evaluated Th1, Th2, Th17 cytokine profile production in the PBMC cells from Chagas' disease patients (cardiomyopathy-free, and with mild, moderate or severe cardiomyopathy cultured with T. cruzi antigen. Cultures of PBMC from patients with moderate and severe cardiomyopathy produced high levels of TNF-α, IFN-γ and low levels of IL-10, when compared to mild cardiomyopathy or cardiomyopathy-free patients. Flow cytometry analysis showed higher CD4(+IL-17(+ cells in PBMC cultured from patients without or with mild cardiomyopathy, in comparison to patients with moderate or severe cardiomyopathy. We then analyzed the presence and function of regulatory T cells in all patients. All groups of Chagas' disease patients presented the same frequency of CD4(+CD25(+ regulatory T cells. However, CD4(+CD25(+ T cells from patients with mild cardiomyopathy or cardiomyopathy-free showed higher suppressive activity than those with moderate and severe cardiomyopathy. IFN-γ levels during chronic Chagas' disease are inversely correlated to the LVEF (P = 0.007, r = -0.614, while regulatory T cell activity is directly correlated with LVEF (P = 0.022, r = 0.500. CONCLUSION/SIGNIFICANCE: These results indicate that reduced production of the cytokines IL-10 and IL-17 in association with high levels of IFN-γ and TNF

  12. Actin-dependent activation of serum response factor in T cells by the viral oncoprotein tip

    Katsch Kristin

    2012-03-01

    Full Text Available Abstract Serum response factor (SRF acts as a multifunctional transcription factor regulated by mutually exclusive interactions with ternary complex factors (TCFs or myocardin-related transcription factors (MRTFs. Binding of Rho- and actin-regulated MRTF:SRF complexes to target gene promoters requires an SRF-binding site only, whereas MAPK-regulated TCF:SRF complexes in addition rely on flanking sequences present in the serum response element (SRE. Here, we report on the activation of an SRE luciferase reporter by Tip, the viral oncoprotein essentially contributing to human T-cell transformation by Herpesvirus saimiri. SRE activation in Tip-expressing Jurkat T cells could not be attributed to triggering of the MAPK pathway. Therefore, we further analyzed the contribution of MRTF complexes. Indeed, Tip also activated a reporter construct responsive to MRTF:SRF. Activation of this reporter was abrogated by overexpression of a dominant negative mutant of the MRTF-family member MAL. Moreover, enrichment of monomeric actin suppressed the Tip-induced reporter activity. Further upstream, the Rho-family GTPase Rac, was found to be required for MRTF:SRF reporter activation by Tip. Initiation of this pathway was strictly dependent on Tip's ability to interact with Lck and on the activity of this Src-family kinase. Independent of Tip, T-cell stimulation orchestrates Src-family kinase, MAPK and actin pathways to induce SRF. These findings establish actin-regulated transcription in human T cells and suggest its role in viral oncogenesis.

  13. Circulating gamma delta T cells are activated and depleted during progression of high-grade gliomas: Implications for gamma delta T cell therapy of GBM

    Glioblastoma multiforme (GBM) remains frustratingly impervious to any existing therapy. We have previously shown that GBM is sensitive to recognition and lysis by ex vivo activated gamma delta T cells, a minor subset of lymphocytes that innately recognize autologous stress-associated target antigens...

  14. Response of Human T Cells to Tetanus Neurotoxin HCC Sub-Domain: T Cell Cytokine Production and Activation Marker Induced by HCC

    Maryam Ghafari-Khamene

    2015-10-01

    Full Text Available Tetanus is caused by the tetanus neurotoxin (TeNT, a 150 kDa single polypeptide molecule which is cleaved into active two-chain molecules composed of a 50 kDa N-terminal light (L and a 100 kDa C-terminal heavy (H chains. Fragment C is further subdivided into two subdomains: the proximal HCN  subdomain and the extreme carboxy subdomain, HCC. HCC is considered as an immunodominant part of TeNT and is responsible for TeNT binding activity to neurons.In the present study, we investigated the ability of recombinant HCC(r HCC to induce Tcell activation. Our results showed that recombinant HCC has a stimulatory effect on IFN-γ secretion by T cells after 48h co-incubation in the presence of anti-TLR-2 Ab. Also, Hcc can induce the expression of CD69 on T cells.Our finding indicated that stimulatory effects of HCC on T cells are TLR-2 independentand anti-TLR-2 inhibitory antibody fails to neutralize HCC stimulatory effects on T cells.Furthermore, HCC  is critical for immunogenic activity of TeNT and is able to induce Tcells through TLR-2 independent pathway.

  15. Murine T cell activation is regulated by surfen (bis-2-methyl-4-amino-quinolyl-6-carbamide)

    Warford, Jordan, E-mail: jordan.warford@dal.ca [Department of Pathology, Dalhousie University, Tupper Building, 5850 College Street, Halifax, Nova Scotia B3H 4R2 (Canada); Doucette, Carolyn D., E-mail: carolyn.doucette@dal.ca [Department of Microbiology and Immunology, Dalhousie University, Tupper Building, 5850 College Street, Halifax, Nova Scotia B3H 4R2 (Canada); Hoskin, David W., E-mail: d.w.hoskin@dal.ca [Department of Pathology, Dalhousie University, Tupper Building, 5850 College Street, Halifax, Nova Scotia B3H 4R2 (Canada); Department of Microbiology and Immunology, Dalhousie University, Tupper Building, 5850 College Street, Halifax, Nova Scotia B3H 4R2 (Canada); Easton, Alexander S., E-mail: alexander.easton@dal.ca [Department of Pathology, Dalhousie University, Tupper Building, 5850 College Street, Halifax, Nova Scotia B3H 4R2 (Canada); Department of Microbiology and Immunology, Dalhousie University, Tupper Building, 5850 College Street, Halifax, Nova Scotia B3H 4R2 (Canada); Department of Surgery (Neurosurgery), Dalhousie University, Tupper Building, 5850 College Street, Halifax, Nova Scotia B3H 4R2 (Canada)

    2014-01-10

    Highlights: •Surfen is the first inhibitor of glycosaminoglycan function to be studied in murine T cells. •Surfen reduces T cell proliferation stimulated in vitro and in vivo. •Surfen reduces CD25 expression in T cells activated in vivo but not in vitro. •Surfen increases T cell proliferation when T cell receptor activation is bypassed. •Surfen’s effects are blocked by co-administration of heparin sulfate. -- Abstract: Surfen (bis-2-methyl-4-amino-quinolyl-6-carbamide) binds to glycosaminoglycans (GAGs) and has been shown to influence their function, and the function of proteoglycans (complexes of GAGs linked to a core protein). T cells synthesize, secrete and express GAGs and proteoglycans which are involved in several aspects of T cell function. However, there are as yet no studies on the effect of GAG-binding agents such as surfen on T cell function. In this study, surfen was found to influence murine T cell activation. Doses between 2.5 and 20 μM produced a graduated reduction in the proliferation of T cells activated with anti-CD3/CD28 antibody-coated T cell expander beads. Surfen (20 mg/kg) was also administered to mice treated with anti-CD3 antibody to activate T cells in vivo. Lymphocytes from surfen-treated mice also showed reduced proliferation and lymph node cell counts were reduced. Surfen reduced labeling with a cell viability marker (7-ADD) but to a much lower extent than its effect on proliferation. Surfen also reduced CD25 (the α-subunit of the interleukin (IL)-2 receptor) expression with no effect on CD69 expression in T cells treated in vivo but not in vitro. When receptor activation was bypassed by treating T cells in vitro with phorbyl myristate acetate (10 ng/ml) and ionomycin (100 ng/ml), surfen treatment either increased proliferation (10 μM) or had no effect (2.5, 5 and 20 μM). In vitro treatment of T cells with surfen had no effect on IL-2 or interferon-γ synthesis and did not alter proliferation of the IL-2 dependent cell

  16. Impaired proteasome function activates GATA3 in T-cells and upregulates CTLA-4: relevance for Sezary syndrome

    Gibson, Heather M; Mishra, Anjali; Chan, Derek V.; Hake, Timothy S.; Porcu, Pierluigi; Wong, Henry K.

    2012-01-01

    Highly regulated expression of the negative co-stimulatory molecule CTLA-4 on T-cells modulates T-cell activation and proliferation. CTLA-4 is preferentially expressed in Th2 T-cells, whose differentiation depends on the transcriptional regulator GATA3. Sezary syndrome (SS) is a T-cell malignancy characterized by Th2 cytokine skewing, impaired T-cell responses, and over-expression of GATA3 and CTLA-4. GATA3 is regulated by phosphorylation and ubiquitination. In SS cells, we detected increased...

  17. Computational investigations into the orgins of 'short term' biochemical memory in T cell activation

    Locasale, Jason W

    2007-01-01

    Recent studies have reported that T cells can integrate signals between interrupted encounters with Antigen Presenting Cells (APCs) in such a way that the process of signal integration exhibits a form of memory. Here, we carry out a computational study using a simple mathematical model of T cell activation to investigate the ramifications of interrupted T cell-APC contacts on signal integration. We consider several mechanisms of how signal integration at these time scales may be achieved and conclude that feedback control of immediate early gene products (IEGs) appears to be a highly plausible mechanism that allows for effective signal integration and cytokine production from multiple exposures to APCs. Analysis of these computer simulations provides an experimental roadmap involving several testable predictions.

  18. 5-Azacytidine Promotes an Inhibitory T-Cell Phenotype and Impairs Immune Mediated Antileukemic Activity

    Thomas Stübig; Anita Badbaran; Tim Luetkens; York Hildebrandt; Djordje Atanackovic; Binder, Thomas M. C.; Boris Fehse; Nicolaus Kröger

    2014-01-01

    Demethylating agent, 5-Azacytidine (5-Aza), has been shown to be active in treatment of myeloid malignancies. 5-Aza enhances anticancer immunity, by increasing expression of tumor-associated antigens. However, the impact of 5-Aza immune responses remains poorly understood. Here, T-cell mediated tumor immunity effects of 5-Aza, are investigated in vitro and in vivo. T-cells from healthy donors were treated with 5-Aza and analyzed by qRT-PCR and flow cytometry for changes in gene expression and...

  19. The calcineurin-NFAT pathway controls activity-dependent circadian gene expression in slow skeletal muscle

    Dyar, Kenneth A.; Stefano Ciciliot; Guidantonio Malagoli Tagliazucchi; Giorgia Pallafacchina; Jana Tothova; Carla Argentini; Lisa Agatea; Reimar Abraham; Miika Ahdesmäki; Mattia Forcato; Silvio Bicciato; Stefano Schiaffino; Bert Blaauw

    2015-01-01

    Objective: Physical activity and circadian rhythms are well-established determinants of human health and disease, but the relationship between muscle activity and the circadian regulation of muscle genes is a relatively new area of research. It is unknown whether muscle activity and muscle clock rhythms are coupled together, nor whether activity rhythms can drive circadian gene expression in skeletal muscle. Methods: We compared the circadian transcriptomes of two mouse hindlimb muscles wi...

  20. Polymerization-dependent activation of porcine γδ T-cells by proanthocyanidins

    Williams, Andrew Richard; Fryganas, Christos; Reichwald, Kirsten;

    2016-01-01

    Plant-derived proanthocyanidins (PAC) have been promoted as a natural method of improving health and immune function in livestock. It has previously been shown that PAC are effective agonists for activating ruminant γδ T-cells in vitro, however effects on other livestock species are not yet clear....... Moreover, the fine structural characteristics of the PAC which contribute to this stimulatory effect have not been elucidated. Here, we demonstrate activation of porcine γδ T-cells by PAC via up-regulation of CD25 (IL-2Rα) and show that 1) activation is dependent on degree of polymerization (DP), with PAC...... this effect of PAC is restricted to the γδ T-cell population within porcine peripheral mononuclear cells as significant CD25 up-regulation was not observed in non γδ T-cells, and no activation (via CD80/86 up-regulation) was evident in monocytes. Our results show that dietary PAC may contribute to...

  1. Transcriptional regulation by Poly(ADP-ribose polymerase-1 during T cell activation

    Parrilla Pascual

    2008-04-01

    Full Text Available Abstract Background Accumulating evidence suggests an important role for the enzyme poly(ADP-ribose polymerase-1 (PARP-1 as an integral part of the gene expression regulatory machinery during development and in response to specific cellular signals. PARP-1 might modulate gene expression through its catalytic activity leading to poly(ADP-ribosylation of nuclear proteins or by its physical association with relevant proteins. Recently, we have shown that PARP-1 is activated during T cell activation. However, the proposed role of PARP-1 in reprogramming T cell gene expression upon activation remains largely unexplored. Results In the present study we use oligonucleotide microarray analysis to gain more insight into the role played by PARP-1 during the gene expression reprogramming that takes place in T cells upon activation with anti-CD3 stimulation alone, or in combination with anti-CD28 co-stimulation. We have identified several groups of genes with expression modulated by PARP-1. The expression of 129 early-response genes to anti-CD3 seems to be regulated by PARP-1 either in a positive (45 genes or in a negative manner (84 genes. Likewise, in the presence of co-stimulation (anti-CD3 + anti-CD28 stimulation, the expression of 203 genes is also regulated by PARP-1 either up (173 genes or down (30 genes. Interestingly, PARP-1 deficiency significantly alters expression of genes associated with the immune response such as chemokines and genes involved in the Th1/Th2 balance. Conclusion This study provides new insights into changes in gene expression mediated by PARP-1 upon T cell activation. Pathway analysis of PARP-1 as a nuclear signalling molecule in T cells would be of relevance for the future development of new therapeutic approaches targeting PARP-1 in the acquired immune response.

  2. T-cell activation is an immune correlate of risk in BCG vaccinated infants.

    Fletcher, Helen A; Snowden, Margaret A; Landry, Bernard; Rida, Wasima; Satti, Iman; Harris, Stephanie A; Matsumiya, Magali; Tanner, Rachel; O'Shea, Matthew K; Dheenadhayalan, Veerabadran; Bogardus, Leah; Stockdale, Lisa; Marsay, Leanne; Chomka, Agnieszka; Harrington-Kandt, Rachel; Manjaly-Thomas, Zita-Rose; Naranbhai, Vivek; Stylianou, Elena; Darboe, Fatoumatta; Penn-Nicholson, Adam; Nemes, Elisa; Hatheril, Mark; Hussey, Gregory; Mahomed, Hassan; Tameris, Michele; McClain, J Bruce; Evans, Thomas G; Hanekom, Willem A; Scriba, Thomas J; McShane, Helen

    2016-01-01

    Vaccines to protect against tuberculosis (TB) are urgently needed. We performed a case-control analysis to identify immune correlates of TB disease risk in Bacille Calmette-Guerin (BCG) immunized infants from the MVA85A efficacy trial. Among 53 TB case infants and 205 matched controls, the frequency of activated HLA-DR(+) CD4(+) T cells associates with increased TB disease risk (OR=1.828, 95% CI=1.25-2.68, P=0.002, FDR=0.04, conditional logistic regression). In an independent study of Mycobacterium tuberculosis-infected adolescents, activated HLA-DR(+) CD4(+) T cells also associate with increased TB disease risk (OR=1.387, 95% CI=1.068-1.801, P=0.014, conditional logistic regression). In infants, BCG-specific T cells secreting IFN-γ associate with reduced risk of TB (OR=0.502, 95% CI=0.29-0.86, P=0.013, FDR=0.14). The causes and impact of T-cell activation on disease risk should be considered when designing and testing TB vaccine candidates for these populations. PMID:27068708

  3. Immunosuppressive activity of pogostone on T cells: Blocking proliferation via S phase arrest.

    Su, Ji-Yan; Luo, Xia; Zhang, Xiao-Jun; Deng, Xiang-Liang; Su, Zi-Ren; Zhou, Lian; Li, Shan-Shan; Dai, Zhenhua; Xu, Yang; Lai, Xiao-Ping

    2015-06-01

    Pogostone (PO) is one of the major chemical constituents of the essential oil of Pogostemon cablin (Blanco) Benth. In the present study, the effect of PO on T cell responsiveness was investigated to explore its potential in immunosuppression by a Concanavalin A (ConA)-stimulation model using splenocytes isolated from C57BL/6 mice. Cytotoxicity by PO on normal splenocytes was evaluated by MTS assays. Characteristics of apoptosis, proliferation, and cell cycle were analyzed by flow cytometry. Related expressions of cyclins and cyclin-dependent kinases (CDKs) were also determined by flow cytometry. Inflammatory cytokine profiling was performed emplying cytometric beads assays (CBA). Moreover, the T cell-mediated delayed Type hepersensity (DTH) model was applied to evaluate the immunosuppressive activity of PO. Neither viability reduction in normal splenocytes nor apoptosis in ConA-stimulated splenocytes was observed under PO treatments. Meanwhile, PO remarkably reduced the total population of ConA-stimulated T cell, blocked T cell proliferation induced by Con A, and inhibited the production of IFN-γ and IL-10. This blockade of stimulated T cell proliferation by PO was likely attributed to down-regulation of cyclin E, cyclin B and CDK1 and the subsequent S-phase arrest. Additionally, PO could inhibit the DTH reaction by alleviating ear swelling and inflammatory infiltrations in the DNCB-challenged ear. Taken together, PO exhibited an immunosuppressive property by directly blocking T cell proliferation as well as altering inflammatory cytokine profile, suggesting that PO may have clinical implications for treating autoimmune diseases and other immune-based disorders. PMID:25912345

  4. A Functionalized Sphingolipid Analogue for Studying Redistribution during Activation in Living T Cells.

    Collenburg, Lena; Walter, Tim; Burgert, Anne; Müller, Nora; Seibel, Jürgen; Japtok, Lukasz; Kleuser, Burkhard; Sauer, Markus; Schneider-Schaulies, Sibylle

    2016-05-01

    Sphingolipids are major components of the plasma membrane. In particular, ceramide serves as an essential building hub for complex sphingolipids, but also as an organizer of membrane domains segregating receptors and signalosomes. Sphingomyelin breakdown as a result of sphingomyelinase activation after ligation of a variety of receptors is the predominant source of ceramides released at the plasma membrane. This especially applies to T lymphocytes where formation of ceramide-enriched membrane microdomains modulates TCR signaling. Because ceramide release and redistribution occur very rapidly in response to receptor ligation, novel tools to further study these processes in living T cells are urgently needed. To meet this demand, we synthesized nontoxic, azido-functionalized ceramides allowing for bio-orthogonal click-reactions to fluorescently label incorporated ceramides, and thus investigate formation of ceramide-enriched domains. Azido-functionalized C6-ceramides were incorporated into and localized within plasma membrane microdomains and proximal vesicles in T cells. They segregated into clusters after TCR, and especially CD28 ligation, indicating efficient sorting into plasma membrane domains associated with T cell activation; this was abolished upon sphingomyelinase inhibition. Importantly, T cell activation was not abrogated upon incorporation of the compound, which was efficiently excluded from the immune synapse center as has previously been seen in Ab-based studies using fixed cells. Therefore, the functionalized ceramides are novel, highly potent tools to study the subcellular redistribution of ceramides in the course of T cell activation. Moreover, they will certainly also be generally applicable to studies addressing rapid stimulation-mediated ceramide release in living cells. PMID:27036914

  5. Aurora A drives early signalling and vesicle dynamics during T-cell activation

    Blas-Rus, Noelia; Bustos-Morán, Eugenio; Pérez de Castro, Ignacio; de Cárcer, Guillermo; Borroto, Aldo; Camafeita, Emilio; Jorge, Inmaculada; Vázquez, Jesús; Alarcón, Balbino; Malumbres, Marcos; Martín-Cófreces, Noa B.; Sánchez-Madrid, Francisco

    2016-01-01

    Aurora A is a serine/threonine kinase that contributes to the progression of mitosis by inducing microtubule nucleation. Here we have identified an unexpected role for Aurora A kinase in antigen-driven T-cell activation. We find that Aurora A is phosphorylated at the immunological synapse (IS) during TCR-driven cell contact. Inhibition of Aurora A with pharmacological agents or genetic deletion in human or mouse T cells severely disrupts the dynamics of microtubules and CD3ζ-bearing vesicles at the IS. The absence of Aurora A activity also impairs the activation of early signalling molecules downstream of the TCR and the expression of IL-2, CD25 and CD69. Aurora A inhibition causes delocalized clustering of Lck at the IS and decreases phosphorylation levels of tyrosine kinase Lck, thus indicating Aurora A is required for maintaining Lck active. These findings implicate Aurora A in the propagation of the TCR activation signal. PMID:27091106

  6. Glucose Uptake Is Limiting in T Cell Activation and Requires CD28-Mediated Akt-Dependent and Independent Pathways1

    Jacobs, Sarah R.; Herman, Catherine E.; MacIver, Nancie J.; Wofford, Jessica A.; Wieman, Heather L.; Hammen, Jeremy J.; Rathmell, Jeffrey C.

    2008-01-01

    T cell activation potently stimulates cellular metabolism to support the elevated energetic and biosynthetic demands of growth, proliferation, and effector function. We show that glucose uptake is limiting in T cell activation and that CD28 costimulation is required to allow maximal glucose uptake following TCR stimulation by up-regulating expression and promoting the cell surface trafficking of the glucose transporter Glut1. Regulation of T cell glucose uptake and Glut1 was critical, as low ...

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

    Crucian, Brian E.; Sams, Clarence F.

    2001-01-01

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

  8. Dysregulation of complement system and CD4+ T cell activation pathways implicated in allergic response.

    Alexessander Couto Alves

    Full Text Available Allergy is a complex disease that is likely to involve dysregulated CD4+ T cell activation. Here we propose a novel methodology to gain insight into how coordinated behaviour emerges between disease-dysregulated pathways in response to pathophysiological stimuli. Using peripheral blood mononuclear cells of allergic rhinitis patients and controls cultured with and without pollen allergens, we integrate CD4+ T cell gene expression from microarray data and genetic markers of allergic sensitisation from GWAS data at the pathway level using enrichment analysis; implicating the complement system in both cellular and systemic response to pollen allergens. We delineate a novel disease network linking T cell activation to the complement system that is significantly enriched for genes exhibiting correlated gene expression and protein-protein interactions, suggesting a tight biological coordination that is dysregulated in the disease state in response to pollen allergen but not to diluent. This novel disease network has high predictive power for the gene and protein expression of the Th2 cytokine profile (IL-4, IL-5, IL-10, IL-13 and of the Th2 master regulator (GATA3, suggesting its involvement in the early stages of CD4+ T cell differentiation. Dissection of the complement system gene expression identifies 7 genes specifically associated with atopic response to pollen, including C1QR1, CFD, CFP, ITGB2, ITGAX and confirms the role of C3AR1 and C5AR1. Two of these genes (ITGB2 and C3AR1 are also implicated in the network linking complement system to T cell activation, which comprises 6 differentially expressed genes. C3AR1 is also significantly associated with allergic sensitisation in GWAS data.

  9. NK-like homeodomain proteins activate NOTCH3-signaling in leukemic T-cells

    Homeodomain proteins control fundamental cellular processes in development and in cancer if deregulated. Three members of the NK-like subfamily of homeobox genes (NKLs), TLX1, TLX3 and NKX2-5, are implicated in T-cell acute lymphoblastic leukemia (T-ALL). They are activated by particular chromosomal aberrations. However, their precise function in leukemogenesis is still unclear. Here we screened further NKLs in 24 T-ALL cell lines and identified the common expression of MSX2. The subsequent aim of this study was to analyze the role of MSX2 in T-cell differentiation which may be disturbed by oncogenic NKLs. Specific gene activity was examined by quantitative real-time PCR, and globally by expression profiling. Proteins were analyzed by western blot, immuno-cytology and immuno-precipitation. For overexpression studies cell lines were transduced by lentiviruses. Quantification of MSX2 mRNA in primary hematopoietic cells demonstrated higher levels in CD34+ stem cells as compared to peripheral blood cells and mature CD3+ T-cells. Furthermore, analysis of MSX2 expression levels in T-cell lines after treatment with core thymic factors confirmed their involvement in regulation. These results indicated that MSX2 represents an hematopoietic NKL family member which is downregulated during T-cell development and may functionally substituted by oncogenic NKLs. For functional analysis JURKAT cells were lentivirally transduced, overexpressing either MSX2 or oncogenic TLX1 and NKX2-5, respectively. These cells displayed transcriptional activation of NOTCH3-signaling, including NOTCH3 and HEY1 as analyzed by gene expression profiling and quantitative RT-PCR, and consistently attenuated sensitivity to gamma-secretase inhibitor as analyzed by MTT-assays. Furthermore, in addition to MSX2, both TLX1 and NKX2-5 proteins interacted with NOTCH-pathway repressors, SPEN/MINT/SHARP and TLE1/GRG1, representing a potential mechanism for (de)regulation. Finally, elevated expression of NOTCH3

  10. A virtual lymph node model to dissect the requirements for T-cell activation by synapses and kinapses

    Moreau, Hélène D; Bogle, Gib; Bousso, Philippe

    2016-01-01

    The initiation of T-cell responses in lymph nodes requires T cells to integrate signals delivered by dendritic cells (DCs) during long-lasting contacts (synapses) or more transient interactions (kinapses). However, it remains extremely challenging to understand how a specific sequence of contacts established by T cells ultimately dictates T-cell fate. Here, we have coupled a computational model of T-cell migration and interactions with DCs with a real-time, flow cytometry-like representation of T-cell activation. In this model, low-affinity peptides trigger T-cell proliferation through kinapses but we show that this process is only effective under conditions of high DC densities and prolonged antigen availability. By contrast, high-affinity peptides favor synapse formation and a vigorous proliferation under a wide range of antigen presentation conditions. In line with the predictions, decreasing the DC density in vivo selectively abolished proliferation induced by the low-affinity peptide. Finally, our results suggest that T cells possess a biochemical memory of previous stimulations of at least 1–2 days. We propose that the stability of T-cell–DC interactions, apart from their signaling potency, profoundly influences the robustness of T-cell activation. By offering the ability to control parameters that are difficult to manipulate experimentally, the virtual lymph node model provides new possibilities to tackle the fundamental mechanisms that regulate T-cell responses elicited by infections or vaccines. PMID:27089942

  11. GATA3 induces human T-cell commitment by restraining Notch activity and repressing NK-cell fate.

    Van de Walle, Inge; Dolens, Anne-Catherine; Durinck, Kaat; De Mulder, Katrien; Van Loocke, Wouter; Damle, Sagar; Waegemans, Els; De Medts, Jelle; Velghe, Imke; De Smedt, Magda; Vandekerckhove, Bart; Kerre, Tessa; Plum, Jean; Leclercq, Georges; Rothenberg, Ellen V; Van Vlierberghe, Pieter; Speleman, Frank; Taghon, Tom

    2016-01-01

    The gradual reprogramming of haematopoietic precursors into the T-cell fate is characterized by at least two sequential developmental stages. Following Notch1-dependent T-cell lineage specification during which the first T-cell lineage genes are expressed and myeloid and dendritic cell potential is lost, T-cell specific transcription factors subsequently induce T-cell commitment by repressing residual natural killer (NK)-cell potential. How these processes are regulated in human is poorly understood, especially since efficient T-cell lineage commitment requires a reduction in Notch signalling activity following T-cell specification. Here, we show that GATA3, in contrast to TCF1, controls human T-cell lineage commitment through direct regulation of three distinct processes: repression of NK-cell fate, upregulation of T-cell lineage genes to promote further differentiation and restraint of Notch activity. Repression of the Notch1 target gene DTX1 hereby is essential to prevent NK-cell differentiation. Thus, GATA3-mediated positive and negative feedback mechanisms control human T-cell lineage commitment. PMID:27048872

  12. The role of class I histocompatibility antigens in the regulation of T-cell activation.

    Dasgupta, J D; Cemach, K; Dubey, D P; Yunis, E J; Amos, D. B.

    1987-01-01

    Class I major histocompatibility antigens in humans (HLA antigens) were found to participate in the regulation of T-cell activation and proliferation induced by phytohemagglutinin. W6/32, a monomorphic antibody directed against class I HLA-A,B,C antigens, significantly inhibited the phytohemagglutinin-induced cell proliferation of peripheral blood lymphocytes. Almost complete suppression of cell activation was achieved on a subfraction of peripheral blood lymphocytes enriched in Mo1+ monocyte...

  13. Friend of GATA-1 Represses GATA-3–dependent Activity in CD4+ T Cells

    Zhou, Meixia; Ouyang, Wenjun; Gong, Qian; Katz, Samuel G.; White, J. Michael; Orkin, Stuart H.; Murphy, Kenneth M.

    2001-01-01

    The development of naive CD4+ T cells into a T helper (Th) 2 subset capable of producing interleukin (IL)-4, IL-5, and IL-13 involves a signal transducer and activator of transcription (Stat)6-dependent induction of GATA-3 expression, followed by Stat6-independent GATA-3 autoactivation. The friend of GATA (FOG)-1 protein regulates GATA transcription factor activity in several stages of hematopoietic development including erythrocyte and megakaryocyte differentiation, but whether FOG-1 regulat...

  14. Bacterial capsular polysaccharide prevents the onset of asthma through T-cell activation.

    Johnson, Jenny L; Jones, Mark B; Cobb, Brian A

    2015-04-01

    Over the last four decades, increases in the incidence of immune-mediated diseases in the Western world have been linked to changes in microbial exposure. It is becoming increasingly clear that the normal microbiota in the gut can profoundly alter susceptibility to a wide range of diseases, such as asthma, in which immune homeostasis is disrupted, yet the mechanisms governing this microbial influence remains poorly defined. In this study, we show that gastrointestinal exposure to PSA, a capsular polysaccharide derived from the commensal bacterium Bacteroides fragilis, significantly limits susceptibility to the induction of experimental asthma. We report that direct treatment of mice with PSA generates protection from asthma, and this effect can be given to a naïve recipient by adoptive transfer of CD4(+) T cells from PSA-exposed mice. Remarkably, we found that these PSA-induced T cells are not canonical FoxP3(+) regulatory T cells, but that they potently inhibit both Th1 and Th2 models of asthma in an IL-10-dependent fashion. These findings reveal that bacterial polysaccharides link the microbiota with the peripheral immune system by activating CD4(+)Foxp3(-) T cells upon exposure in the gut, and they facilitate resistance to unnecessary inflammatory responses via the production of IL-10. PMID:25347992

  15. CD11b expression as a marker to distinguish between recently activated effector CD8(+) T cells and memory cells

    Christensen, Jeanette Erbo; Ørding Andreasen, Susanne; Christensen, Jan Pravsgaard;

    2001-01-01

    subset. Polyclonal virus-specific effector and memory CD8(+) T cells from lymphocytic choriomeningitis- and vesicular stomatitis virus-infected mice were visualized through staining for intracellular IFN-gamma or binding of MHC-peptide tetramers, and Mac-1 expression was evaluated. Naive T cells and most......CD8(+) T cells in different activation states have been difficult to identify phenotypically. In this study we have investigated whether Mac-1 (CD11b) expression can be used as a criterion to distinguish between recently activated effector cells and memory cells belonging to the CD8(+) T cell...... virus-specific memory CD8(+) T cells express little or no Mac-1 independent of the virus model employed. In contrast, the majority of CD8(+) T cells present during acute infection express a significant level of Mac-1 and, similarly, Mac-1 expression is found on secondary effectors generated in response...

  16. Selection and characterization of T-cell variants lacking molecules involved in T-cell activation (T3 T-cell receptor, T44, and T11): analysis of the functional relationship among different pathways of activation

    Moretta, A.; Poggi, A.; Olive, D.; Bottino, C.; Fortis, C.; Pantaleo, G.; Moretta, L.

    1987-03-01

    A clone of the interleukin 2-producing Jurkat leukemia cell line termed JA3 (surface phenotype, T3/sup +/, Ti/sup +/, T44/sup +/, T11/sup +/, T40/sup +/) has been used to induce and select cell variants lacking surface molecules involved in T-cell activation. Following 200 rad of ..gamma..-radiation (1 rad = 0.01 Gy), cells were treated with monoclonal antibodies (mAbs) directed to T3, Ti, T44, or T11 antigen and complement. After growth of the residual cells in culture, negative cells were cloned under limiting conditions. Depending on the specificity of the mAb used for the immunoselection, three groups of variants were obtained. (i) The use of mAbs directed to T3 or Ti resulted in cell variants that expressed the T3/sup -/ Ti/sup -/T44/sup +/ Leu1/sup +/ T11/sup +/ T40/sup +/ 4F2/sup +/ HLA class I/sup +/ surface phenotype. (ii) Immunoselection with anti-T44 mAb resulted in 2 variants that shared the T3/sup -/ Ti/sup -/ T44/sup -/ Leu1/sup -/ T11/sup -/ T40/sup -/ 4F2/sup -/ HLA class I/sup +/ phenotype. (iii) Cell treatment with anti-T11 mAb resulted in 15 variants characterized by the lack of T11 antigen expression and of all the other T-cell-specific surface antigens. Therefore, it appears that the different sets of JA3 cell variants, like T cells at discrete stages of intrathymic differentiation, may follow a coordinated expression of surface differentiation antigens. Analysis of the functional responsiveness of the three distinct groups of JA3 cell variants to different stimuli showed that all produced interleukin 2 in response to A23187 calcium ionophore plus phorbol 12-myristate 13-acetate.

  17. FOXP3+Helios+ Regulatory T Cells, Immune Activation, and Advancing Disease in HIV-Infected Children.

    Khaitan, Alka; Kravietz, Adam; Mwamzuka, Mussa; Marshed, Fatma; Ilmet, Tiina; Said, Swalehe; Ahmed, Aabid; Borkowsky, William; Unutmaz, Derya

    2016-08-15

    Regulatory T cells (Tregs) are functionally suppressive CD4 T cells, critical for establishing peripheral tolerance and controlling inflammatory responses. Previous reports of Tregs during chronic HIV disease have conflicting results with higher or lower levels compared with controls. Identifying true Tregs with suppressive activity proves challenging during HIV infection, as traditional Treg markers, CD25 and FOXP3, may transiently upregulate expression as a result of immune activation (IA). Helios is an Ikaros family transcription factor that marks natural Tregs with suppressive activity and does not upregulate expression after activation. Coexpression of FOXP3 and Helios has been suggested as a highly specific marker of "bona fide" Tregs. We evaluated Treg subsets by FOXP3 coexpressed with either CD25 or Helios and their association with HIV disease progression in perinatally infected HIV-positive children. Identifying Tregs by FOXP3 coexpression with Helios rather than CD25 revealed markedly higher Treg frequencies, particularly in HIV+ children. Regardless of antiretroviral therapy, HIV-infected children had a selective expansion of memory FOXP3+Helios+ Tregs. The rise in memory Tregs correlated with declining HIV clinical status, indicated by falling CD4 percentages and CD4:CD8 ratios and increasing HIV plasma viremia and IA. In addition, untreated HIV+ children exhibited an imbalance between the levels of Tregs and activated T cells. Finally, memory Tregs expressed IA markers CD38 and Ki67 and exhaustion marker, PD-1, that tightly correlated with a similar phenotype in memory CD4 T cells. Overall, HIV-infected children had significant disruptions of memory Tregs that associated with advancing HIV disease. PMID:27003495

  18. Alterations of T cell activation signalling and cytokine production by postmenopausal estrogen levels

    Taylor Douglas D

    2009-03-01

    Full Text Available Abstract Background Immunosenescence is an age-associated disorder occurring primarily in T cell compartments, including altered subset composition, functions, and activation. In women, evidence implicates diminished estrogen in the postmenopausal period as a contributing factor to diminished T cell responsiveness. Since hypoestrogenism is present in postmenopausal women, our objective focused on whether T cell activation, defined as signalling molecule expressions and activation, and function, identified as IL-2 production, were affected by low estrogen. Methods Using Jurkat 6.1 T cells, consequences of 4 pg/ml (corresponding to postmenopausal levels or 40 pg/ml (premenopausal levels of estradiol (E2 were analyzed on signalling proteins, CD3-zeta, JAK2, and JAK3, determined by Western immunoblotting. These consequences were correlated with corresponding gene expressions, quantified by real time-polymerase chain reaction. Tyrosine phosphorylation of CD3-zeta was defined by immunoprecipitation and western immunoblotting following activation by T cell receptor (TcR cross-linking. CD3-zeta expression and modulation was also confirmed in T cells from pre- and postmenopausal women. To assess functional consequences, IL-2 production, induced by PMA and ionomycin, was determined using enzyme-linked immunosorbent spot assay (ELISpot. Results At 40 pg/ml E2, the level of signalling protein CD3-zeta was elevated 1.57-fold, compared with cells exposed to 4 pg/ml E2. The CD3-zeta proteins also exhibited altered levels of activation-induced phosphorylation in the presence of 40 pg/ml E2 versus 4 pg/ml: 23 kD phosphorylated form increased 2.64-fold and the 21 kD form was elevated 2.95-fold. Examination of kinases associated with activation signalling also demonstrated that, in the presence of 40 pg/ml E2, JAK2 protein expression was increased 1.64-fold (p 2 (2.39, 2.01, and 2.21 fold, respectively versus 4 pg/ml. These findings were confirmed in vivo, since T

  19. Affinity and dose of TCR engagement yield proportional enhancer and gene activity in CD4+ T cells

    Allison, Karmel A; Sajti, Eniko; Collier, Jana G; Gosselin, David; Troutman, Ty Dale; Stone, Erica L; Hedrick, Stephen M; Glass, Christopher K

    2016-01-01

    Affinity and dose of T cell receptor (TCR) interaction with antigens govern the magnitude of CD4+ T cell responses, but questions remain regarding the quantitative translation of TCR engagement into downstream signals. We find that while the response of mouse CD4+ T cells to antigenic stimulation is bimodal, activated cells exhibit analog responses proportional to signal strength. Gene expression output reflects TCR signal strength, providing a signature of T cell activation. Expression changes rely on a pre-established enhancer landscape and quantitative acetylation at AP-1 binding sites. Finally, we show that graded expression of activation genes depends on ERK pathway activation, suggesting that an ERK-AP-1 axis plays an important role in translating TCR signal strength into proportional activation of enhancers and genes essential for T cell function. DOI: http://dx.doi.org/10.7554/eLife.10134.001 PMID:27376549

  20. In-vitro T cell mediated function in patients with active rheumatoid arthritis.

    Slavin, S; Strober, S

    1981-01-01

    In-vitro synthesis of peripheral blood lymphocytes from patients with rheumatoid arthritis was measured after stimulation with phytohaemagglutinin (PHA) in a short-term, serum-free culture system. Diminished responses were found in 16 out of 17 consecutive patients with active disease. Normal PHA responsiveness was recovered by assaying Ficoll-Hypaque isolated E rosette forming cells in serum-free medium, indicating basically normal T cell function in RA. Preincubation of normal peripheral bl...

  1. Quantal concept of T-cell activation: adhesion domains as immunological synapses

    Sackmann, Erich

    2011-06-01

    Adhesion micro-domains (ADs) formed during encounters of lymphocytes with antigen-presenting cells (APC) mediate the genetic expression of quanta of cytokines interleukin-2 (IL-2). The IL-2-induced activation of IL-2 receptors promotes the stepwise progression of the T-cells through the cell cycle, hence their name, immunological synapses. The ADs form short-lived reaction centres controlling the recruitment of activators of the biochemical pathway (the kinases Lck and ZAP) while preventing the access of inhibitors (phosphatase CD45) through steric repulsion forces. CD45 acts as the generator of adhesion domains and, through its role as a spacer protein, also as the promoter of the reaction. In a second phase of T-cell-APC encounters, long-lived global reaction spaces (called supramolecular activation complexes (SMAC)) form by talin-mediated binding of the T-cell integrin (LFA-1) to the counter-receptor ICAM-1, resulting in the formation of ring-like tight adhesion zones (peripheral SMAC). The ADs move to the centre of the intercellular adhesion zone forming the central SMAC, which serve in the recycling of the AD. We propose that cell stimulation is triggered by integrating the effect evoked by the short-lived adhesion domains. Similar global reaction platforms are formed by killer cells to destruct APC. We present a testable mechanical model showing that global reaction spaces (SMAC or dome-like contacts between cytotoxic cells and APC) form by self-organization through delayed activation of the integrin-binding affinity and stabilization of the adhesion zones by F-actin recruitment. The mechanical stability and the polarization of the adhering T-cells are mediated by microtubule-actin cross-talk.

  2. Quantal concept of T-cell activation: adhesion domains as immunological synapses

    Sackmann, Erich, E-mail: sackmann@ph.tum.de [Physics Department E22, Technical University Munich, D-85748 Garching (Germany)

    2011-06-15

    Adhesion micro-domains (ADs) formed during encounters of lymphocytes with antigen-presenting cells (APC) mediate the genetic expression of quanta of cytokines interleukin-2 (IL-2). The IL-2-induced activation of IL-2 receptors promotes the stepwise progression of the T-cells through the cell cycle, hence their name, immunological synapses. The ADs form short-lived reaction centres controlling the recruitment of activators of the biochemical pathway (the kinases Lck and ZAP) while preventing the access of inhibitors (phosphatase CD45) through steric repulsion forces. CD45 acts as the generator of adhesion domains and, through its role as a spacer protein, also as the promoter of the reaction. In a second phase of T-cell-APC encounters, long-lived global reaction spaces (called supramolecular activation complexes (SMAC)) form by talin-mediated binding of the T-cell integrin (LFA-1) to the counter-receptor ICAM-1, resulting in the formation of ring-like tight adhesion zones (peripheral SMAC). The ADs move to the centre of the intercellular adhesion zone forming the central SMAC, which serve in the recycling of the AD. We propose that cell stimulation is triggered by integrating the effect evoked by the short-lived adhesion domains. Similar global reaction platforms are formed by killer cells to destruct APC. We present a testable mechanical model showing that global reaction spaces (SMAC or dome-like contacts between cytotoxic cells and APC) form by self-organization through delayed activation of the integrin-binding affinity and stabilization of the adhesion zones by F-actin recruitment. The mechanical stability and the polarization of the adhering T-cells are mediated by microtubule-actin cross-talk.

  3. Increased Mucosal CD4+ T Cell Activation in Rhesus Macaques following Vaccination with an Adenoviral Vector

    Bukh, Irene; Calcedo, Roberto; Roy, Soumitra; Carnathan, Diane G.; Grant, Rebecca; Qin, Qiuyue; Boyd, Surina; Ratcliffe, Sarah J.; Veeder, Christin L; Bellamy, Scarlett L.; Betts, Michael R.; James M Wilson

    2014-01-01

    The possibility that vaccination with adenovirus (AdV) vectors increased mucosal T cell activation remains a central hypothesis to explain the potential enhancement of HIV acquisition within the Step trial. Modeling this within rhesus macaques is complicated because human adenoviruses, including human adenovirus type 5 (HAdV-5), are not endogenous to macaques. Here, we tested whether vaccination with a rhesus macaque-derived adenoviral vector (simian adenovirus 7 [SAdV-7]) enhances mucosal T ...

  4. Spaceflight alters expression of microRNA during T-cell activation.

    Hughes-Fulford, Millie; Chang, Tammy T; Martinez, Emily M; Li, Chai-Fei

    2015-12-01

    Altered immune function has been demonstrated in astronauts during spaceflights dating back to Apollo and Skylab; this could be a major barrier to long-term space exploration. We tested the hypothesis that spaceflight causes changes in microRNA (miRNA) expression. Human leukocytes were stimulated with mitogens on board the International Space Station using an onboard normal gravity control. Bioinformatics showed that miR-21 was significantly up-regulated 2-fold during early T-cell activation in normal gravity, and gene expression was suppressed under microgravity. This was confirmed using quantitative real-time PCR (n = 4). This is the first report that spaceflight regulates miRNA expression. Global microarray analysis showed significant (P < 0.05) suppression of 85 genes under microgravity conditions compared to normal gravity samples. EGR3, FASLG, BTG2, SPRY2, and TAGAP are biologically confirmed targets and are co-up-regulated with miR-21. These genes share common promoter regions with pre-mir-21; as the miR-21 matures and accumulates, it most likely will inhibit translation of its target genes and limit the immune response. These data suggest that gravity regulates T-cell activation not only by transcription promotion but also by blocking translation via noncoding RNA mechanisms. Moreover, this study suggests that T-cell activation itself may induce a sequence of gene expressions that is self-limited by miR-21. PMID:26276131

  5. CD26 surface molecule involvement in T cell activation and lymphokine synthesis in rheumatoid and other inflammatory synovitis.

    Gerli, R; Muscat, C; Bertotto, A; Bistoni, O; Agea, E; Tognellini, R; Fiorucci, G; Cesarotti, M; Bombardieri, S

    1996-07-01

    T cell surface expression and the functional role of CD26 antigen (Ag), a surface ectoenzyme involved in T cell activation and migration across the extracellular matrix, were analyzed in the peripheral blood (PB) and synovial fluid (SF) from patients with inflammatory arthritides. CD26 membrane expression on T cells was detected by cytofluorometry using two different monoclonal antibodies, anti-Ta1 and anti-1F7, while cell proliferation and both IL-2 and IFN-gamma production were evaluated in anti-CD3- or anti-CD2-stimulated cell cultures after Ag surface modulation with anti-1F7. The results showed that Ta1 and 1F7 Ag expression were increased on T cells from PB of patients with active, but not inactive, rheumatoid arthritis (RA). Most SF T cells from RA or other inflammatory arthritides displayed the memory marker CD45R0 and the Ta1 Ag, but lacked the 1F7 molecule. In addition, in vitro 1F7 modulation, which enhanced RA PB T cell proliferation and both IL-2 and IFN-gamma synthesis, did not synergize with anti-CD3 or anti-CD2 in inducing IL-2-dependent activation of SF T cells, but reduced IFN-gamma production. A spontaneous reappearance of 1F7 Ag on the SF T cell surface was seen after 2-5 days in culture. Phorbol myristate acetate, able to accelerate its reexpression, also restored a normal response of SF T cells to anti-1F7 comitogenic effects. These data confirm a role of the CD26 surface molecule in regulating T cell activation and lymphokine synthesis. This observation may have important implications in the regulation of T cell activity at the joint level during chronic inflammatory processes. PMID:8674237

  6. Vitamin E reverses impaired linker for activation of T cells activation in T cells from aged C57BL/6 mice

    Supplemental vitamin E restores age-related defects in IL-2 production, T cell proliferation, and immune synapse formation. Here, we evaluated the effect of vitamin E on TCR-proximal signaling events. In aged murine CD4+ T cells stimulated via CD3 and CD28, tyrosine 191 of the adaptor protein LAT wa...

  7. Tumor Antigen Specific Activation of Primary Human T-Cells Expressing a Virally Encoded Chimeric T-Cell Receptor Specific for p185HER2

    杨建民; MichaelSFRIEDMAN; ChristopherMREYNOLDS; MarianneTHUBEN; LeeWILKE; JenniferFULLER; 李桥; ZeligESHHAR; JamesJMULE; KevimTMCDONAGH

    2004-01-01

    We have developed and tested chimeric T-cell receptors (TCR) specific for p185HER2. In these experiments,retroviral vectors expressing the N297 or N29ξ receptors were constructed in pRET6. Amphotropic viral producer cells were established in the GALV-based PG13 packaging cell line. Ficoll purified human peripheral blood lymphocytes (PBL) were vitally transduced using an optimized protocol incorporating activation with immobilized anti-CD3/anti-CD28 monoclonal antibodies, followed by viral infection in the presence of fibronectin fragment CH296. Transduced cells were co-cultured with human tumor cell lines that overexpress (SK-OV-3) or underexpress (MCF7) p185HER2 to assay for antigen specific immune responses. Both CD4+ and CD8+ T-cells transduced with the N297 or N29ξ chTCR demonstrated HER2-specific antigen responses, as determined by release of Th1 like cytokines, and cellular cytotoxicity assays. Our results support the feasibility of adoptive immunothempy with genetically modified T-cells expressing a chTCR specific for p185HER2.

  8. Temporal protein expression pattern in intracellular signalling cascade during T-cell activation: a computational study.

    Ganguli, Piyali; Chowdhury, Saikat; Bhowmick, Rupa; Sarkar, Ram Rup

    2015-10-01

    Various T-cell co-receptor molecules and calcium channel CRAC play a pivotal role in the maintenance of cell's functional responses by regulating the production of effector molecules (mostly cytokines) that aids in immune clearance and also maintaining the cell in a functionally active state. Any defect in these co-receptor signalling pathways may lead to an altered expression pattern of the effector molecules. To study the propagation of such defects with time and their effect on the intracellular protein expression patterns, a comprehensive and largest pathway map of T-cell activation network is reconstructed manually. The entire pathway reactions are then translated using logical equations and simulated using the published time series microarray expression data as inputs. After validating the model, the effect of in silico knock down of co-receptor molecules on the expression patterns of their downstream proteins is studied and simultaneously the changes in the phenotypic behaviours of the T-cell population are predicted, which shows significant variations among the proteins expression and the signalling routes through which the response is propagated in the cytoplasm. This integrative computational approach serves as a valuable technique to study the changes in protein expression patterns and helps to predict variations in the cellular behaviour. PMID:26564978

  9. TGF-β and IL-21 cooperatively stimulate activated CD8(+) T cells to differentiate into Tc17 cells.

    Chen, Hsin-Wei; Tsai, Jy-Ping; Yao, Tsung-You; Hsieh, Chia-Ling; Chen, I-Hua; Liu, Shin-Jen

    2016-06-01

    TGF-β together with IL-21 or IL-6 can drive the differentiation of naïve CD8(+) T cells into IL-17-producing CD8(+) T cells. These IL-17-producing CD8(+) T cells are termed Tc17 cells. Tc17 cells preserve plasticity under various conditions in vitro and in vivo. IFN-γ-producing CD8(+) T cells are termed Tc1 cells. However, Tc1 cells are considered relatively stable. In the present study, we show that the combination of TGF-β plus IL-21, but not IL-6, converts Tc1 cells into Tc17 cells; this conversion is associated with elevated RORα, RORγt, and Batf mRNA levels. These results indicate that Tc1 cells are skewed to the Tc17 cell phenotype under TGF-β plus IL-21-polarizing conditions. Furthermore, IL-6R is expressed on naïve, but not activated, CD8(+) T cells. In contrast, IL-21R is expressed on both naïve and activated CD8(+) T cells. Thus, differential expression profiles of IL-6R and IL-21R on naïve and activated CD8(+) T cells may be one mechanism by which TGF-β plus IL-21, but not IL-6, can drive activated CD8(+) T cells to differentiate into IL-17-producing cells. Taken together, these results provide a novel viewpoint for the plasticity of Tc1 cells. PMID:27085379

  10. Tcf1 and Lef1 transcription factors establish CD8(+) T cell identity through intrinsic HDAC activity.

    Xing, Shaojun; Li, Fengyin; Zeng, Zhouhao; Zhao, Yunjie; Yu, Shuyang; Shan, Qiang; Li, Yalan; Phillips, Farrah C; Maina, Peterson K; Qi, Hank H; Liu, Chengyu; Zhu, Jun; Pope, R Marshall; Musselman, Catherine A; Zeng, Chen; Peng, Weiqun; Xue, Hai-Hui

    2016-06-01

    The CD4(+) and CD8(+) T cell dichotomy is essential for effective cellular immunity. How individual T cell identity is established remains poorly understood. Here we show that the high-mobility group (HMG) transcription factors Tcf1 and Lef1 are essential for repressing CD4(+) lineage-associated genes including Cd4, Foxp3 and Rorc in CD8(+) T cells. Tcf1- and Lef1-deficient CD8(+) T cells exhibit histone hyperacetylation, which can be ascribed to intrinsic histone deacetylase (HDAC) activity in Tcf1 and Lef1. Mutation of five conserved amino acids in the Tcf1 HDAC domain diminishes HDAC activity and the ability to suppress CD4(+) lineage genes in CD8(+) T cells. These findings reveal that sequence-specific transcription factors can utilize intrinsic HDAC activity to guard cell identity by repressing lineage-inappropriate genes. PMID:27111144

  11. Human T cell leukemia virus-I (HTLV-I) Tax-mediated apoptosis in activated T cells requires an enhanced intracellular prooxidant state

    Los, Marek Jan; Khazaie, K; Schulze-Osthoff, Klaus; Baeuerle, P A; Schirrmacher, V.; Chlichlia, K.

    1998-01-01

    We have shown that an estradiol-dependent activation of human T cell leukemia virus-I Tax leads to the inhibition of cell proliferation and to the induction of apoptosis, The present study demonstrates that a hormone-dependent activation of Tax promotes an enhanced prooxidant state in stably transfected Jurkat cells as measured by changes in the intracellular levels of glutathione and H2O2; these changes are followed by apoptotic cell death. Additional stimulation of the CD3/TCR pathway enhan...

  12. Human Liver Stem Cells Suppress T-Cell Proliferation, NK Activity, and Dendritic Cell Differentiation

    Stefania Bruno

    2016-01-01

    Full Text Available Human liver stem cells (HLSCs are a mesenchymal stromal cell-like population resident in the adult liver. Preclinical studies indicate that HLSCs could be a good candidate for cell therapy. The aim of the present study was to evaluate the immunogenicity and the immunomodulatory properties of HLSCs on T-lymphocytes, natural killer cells (NKs, and dendritic cells (DCs in allogeneic experimental settings. We found that HLSCs inhibited T-cell proliferation by a mechanism independent of cell contact and dependent on the release of prostaglandin E2 (PGE2 and on indoleamine 2,3-dioxygenase activity. When compared with mesenchymal stromal cells (MSCs, HLSCs were more efficient in inhibiting T-cell proliferation. At variance with MSCs, HLSCs did not elicit NK degranulation. Moreover, HLSCs inhibited NK degranulation against K562, a NK-sensitive target, by a mechanism dependent on HLA-G release. When tested on DC generation from monocytes, HLSCs were found to impair DC differentiation and DCs ability to induce T-cell proliferation through PGE2. This study shows that HLSCs have immunomodulatory properties similar to MSCs, but, at variance with MSCs, they do not elicit a NK response.

  13. Leukemia-associated activating mutation of Flt3 expands dendritic cells and alters T cell responses.

    Lau, Colleen M; Nish, Simone A; Yogev, Nir; Waisman, Ari; Reiner, Steven L; Reizis, Boris

    2016-03-01

    A common genetic alteration in acute myeloid leukemia is the internal tandem duplication (ITD) in FLT3, the receptor for cytokine FLT3 ligand (FLT3L). Constitutively active FLT3-ITD promotes the expansion of transformed progenitors, but also has pleiotropic effects on hematopoiesis. We analyzed the effect of FLT3-ITD on dendritic cells (DCs), which express FLT3 and can be expanded by FLT3L administration. Pre-leukemic mice with the Flt3(ITD) knock-in allele manifested an expansion of classical DCs (cDCs) and plasmacytoid DCs. The expansion originated in DC progenitors, was cell intrinsic, and was further enhanced in Flt3(ITD/ITD) mice. The mutation caused the down-regulation of Flt3 on the surface of DCs and reduced their responsiveness to Flt3L. Both canonical Batf3-dependent CD8(+) cDCs and noncanonical CD8(+) cDCs were expanded and showed specific alterations in their expression profiles. Flt3(ITD) mice showed enhanced capacity to support T cell proliferation, including a cell-extrinsic expansion of regulatory T (T reg) cells. Accordingly, these mice restricted alloreactive T cell responses during graft-versus-host reaction, but failed to control autoimmunity without T reg cells. Thus, the FLT3-ITD mutation directly affects DC development, indirectly modulating T cell homeostasis and supporting T reg cell expansion. We hypothesize that this effect of FLT3-ITD might subvert immunosurveillance and promote leukemogenesis in a cell-extrinsic manner. PMID:26903243

  14. Kinetics of T cell-activation molecules in response to Mycobacterium tuberculosis antigens

    Antas Paulo RZ

    2002-01-01

    Full Text Available The phenotypic features acquired subsequent to antigen-specific stimulation in vitro were evaluated by means of the kinetic expressions of CD69 and CD25 activation molecules on T lymphocytes and assayed by flow cytometry in response to PPD, Ag85B, and ferritin in PPD-positive healthy control individuals. In response to PHA, CD69 staining on both CD4+ and CD8+ T cells became initially marked after 4 h, peaked at 24 h, and quickly decreased after 120 h. For CD25, a latter expression was detected around 8 h, having increased after 96 h. As expected, the response rate to the mycobacterial antigens was much lower than that to the mitogen. Positive staining was high after 96 h for CD25 and after 24 h for CD69. CD69 expression was significantly enhanced (p < 0.05 on CD8+ as compared to CD4+ T cells. High levels were also found between 96-120 h. Regarding Ag85B, CD25+ cells were mostly CD4+ instead of CD8+ T cells. Moreover, in response to ferritin, a lower CD25 expression was noted. The present data will allow further characterization of the immune response to new mycobacterial-specific antigens and their evaluation for possible inclusion in developing new diagnostic techniques for tuberculosis as well in a new vaccine to prevent the disease.

  15. Activated iNKT cells promote memory CD8+ T cell differentiation during viral infection.

    Emma C Reilly

    Full Text Available α-Galactosylceramide (α-GalCer is the prototypical lipid ligand for invariant NKT cells. Recent studies have proposed that α-GalCer is an effective adjuvant in vaccination against a range of immune challenges, however its mechanism of action has not been completely elucidated. A variety of delivery methods have been examined including pulsing dendritic cells with α-GalCer to optimize the potential of α-GalCer. These methods are currently being used in a variety of clinical trials in patients with advanced cancer but cannot be used in the context of vaccine development against pathogens due to their complexity. Using a simple delivery method, we evaluated α-GalCer adjuvant properties, using the mouse model for cytomegalovirus (MCMV. We measured several key parameters of the immune response to MCMV, including inflammation, effector, and central memory CD8(+ T cell responses. We found that α-GalCer injection at the time of the infection decreases viral titers, alters the kinetics of the inflammatory response, and promotes both increased frequencies and numbers of virus-specific memory CD8(+ T cells. Overall, our data suggest that iNKT cell activation by α-GalCer promotes the development of long-term protective immunity through increased fitness of central memory CD8(+ T cells, as a consequence of reduced inflammation.

  16. B cells pulsed with Helicobacter pylori antigen efficiently activate memory CD8+ T cells from H. pylori-infected individuals.

    Azem, Josef; Svennerholm, Ann-Mari; Lundin, B Samuel

    2006-01-01

    Helicobacter pylori infection causes chronic gastritis that may progress to peptic ulcers or gastric adenocarcinoma and thereby cause major world-wide health problems. Previous studies have shown that CD4+ T cells are important in the immune response to H. pylori in humans, but the role of CD8+ T cells is less clear. In order to study the CD8+ T cell response to H. pylori in greater detail, we have evaluated efficient conditions for activation of CD8+ T cells in vitro. We show that H. pylori-reactive CD8+ T cells can be activated most efficiently by B cells or dendritic cells pulsed with H. pylori antigens. We further show that the majority of CD8+ T cells in H. pylori-infected gastric mucosa are memory cells, and that memory CD8+ T cells sorted from peripheral blood of H. pylori-infected individuals respond 15-fold more to H. pylori urease compared to memory cells from uninfected subjects. We conclude that CD8+ T cells do participate in the immune response to H. pylori, and this may have implications for the development of more severe disease outcomes in H. pylori-infected subjects. PMID:16324887

  17. Bcl-2 Knockdown Accelerates T Cell Receptor-Triggered Activation-Induced Cell Death in Jurkat T Cells

    Lee, Yun-Jung; Won, Tae Joon; Hyung, Kyeong Eun; Lee, Mi Ji; Moon, Young-hye; Lee, Ik Hee; Go, Byung Sung; Hwang, Kwang Woo

    2014-01-01

    Cell death and survival are tightly controlled through the highly coordinated activation/inhibition of diverse signal transduction pathways to insure normal development and physiology. Imbalance between cell death and survival often leads to autoimmune diseases and cancer. Death receptors sense extracellular signals to induce caspase-mediated apoptosis. Acting upstream of CED-3 family proteases, such as caspase-3, Bcl-2 prevents apoptosis. Using short hairpin RNAs (shRNAs), we suppressed Bcl-...

  18. The Role of E3 Ubiquitin Ligase Cbl Proteins in Interleukin-2-Induced Jurkat T-Cell Activation

    Ming-Fang Zhao; Xiu-Juan Qu; Jing-Lei Qu; You-Hong Jiang; Ye Zhang; Ke-Zuo Hou; Hao Deng; Yun-Peng Liu

    2013-01-01

    Interleukin- (IL-) 2 is the major growth factor for T-cell activation and proliferation. IL-2 has multiple functions in the regulation of immunological processes. Although most studies focus on T-cell immunomodulation, T-cell activation by IL-2 is the foundation of priming the feedback loop. Here, we investigated the effect of MAPK/ERK and PI3K/Akt signaling pathways on IL-2-induced cell activation and the regulatory mechanisms of upstream ubiquitin ligase Cbl-b and c-Cbl. Morphological analy...

  19. Naive human T cells are activated and proliferate in response to the heme oxygenase-1 inhibitor tin mesoporphyrin.

    Burt, Trevor D; Seu, Lillian; Mold, Jeffrey E; Kappas, Attallah; McCune, Joseph M

    2010-11-01

    Heme oxygenase-1 (HO-1) and its catabolic by-products have potent anti-inflammatory activity in many models of disease. It is not known, however, if HO-1 also plays a role in the homeostatic control of T cell activation and proliferation. We demonstrate here that the HO-1 inhibitor tin mesoporphyrin (SnMP) induces activation, proliferation, and maturation of naive CD4(+) and CD8(+) T cells via interactions with CD14(+) monocytes in vitro. This response is dependent upon interactions of T cells with MHC class I and II on the surface of CD14(+) monocytes. Furthermore, CD4(+)CD25(+)FoxP3(+) regulatory T cells were able to suppress this proliferation, even though their suppressive activity was itself impaired by SnMP. Given the magnitude of the Ag-independent T cell response induced by SnMP, we speculate that HO-1 plays an important role in dampening nonspecific T cell activation. Based on these findings, we propose a potential role for HO-1 in the control of naive T cell homeostatic proliferation. PMID:20921523

  20. Functional activation of the T-cell antigen receptor induces tyrosine phosphorylation of phospholipase C-gamma 1.

    Weiss, A; Koretzky, G; Schatzman, R C; Kadlecek, T

    1991-01-01

    Stimulation of the T-cell antigen receptor (TCR), which itself is not a protein-tyrosine kinase (PTK), activates a PTK and phospholipase C (PLC). Using the human T-cell leukemic line Jurkat and normal peripheral blood lymphocytes, we demonstrate that stimulation of the TCR specifically induces the recovery of PLC activity in eluates from anti-phosphotyrosine immunoprecipitates. Stimulation of the human muscarinic receptor, subtype 1, when expressed in Jurkat activates PLC through a guanine nu...

  1. IL-12 and IL-4 activate a CD39-dependent intrinsic peripheral tolerance mechanism in CD8(+) T cells.

    Noble, Alistair; Mehta, Hema; Lovell, Andrew; Papaioannou, Eleftheria; Fairbanks, Lynette

    2016-06-01

    Immune responses to protein antigens involve CD4(+) and CD8(+) T cells, which follow distinct programs of differentiation. Naïve CD8 T cells rapidly develop cytotoxic T-cell (CTL) activity after T-cell receptor stimulation, and we have previously shown that this is accompanied by suppressive activity in the presence of specific cytokines, i.e. IL-12 and IL-4. Cytokine-induced CD8(+) regulatory T (Treg) cells are one of several Treg-cell phenotypes and are Foxp3(-) IL-10(+) with contact-dependent suppressive capacity. Here, we show they also express high level CD39, an ecto-nucleotidase that degrades extracellular ATP, and this contributes to their suppressive activity. CD39 expression was found to be upregulated on CD8(+) T cells during peripheral tolerance induction in vivo, accompanied by release of IL-12 and IL-10. CD39 was also upregulated during respiratory tolerance induction to inhaled allergen and on tumor-infiltrating CD8(+) T cells. Production of IL-10 and expression of CD39 by CD8(+) T cells was independently regulated, being respectively blocked by extracellular ATP and enhanced by an A2A adenosine receptor agonist. Our results suggest that any CTL can develop suppressive activity when exposed to specific cytokines in the absence of alarmins. Thus negative feedback controls CTL expansion under regulation from both nucleotide and cytokine environment within tissues. PMID:26990545

  2. Investigation of biochemical property changes in activation-induced CD 8 + T cell apoptosis using Raman spectroscopy

    Lee, Young Ju; Ahn, Hyung Joon; Lee, Gi-Ja; Jung, Gyeong Bok; Lee, Gihyun; Kim, Dohyun; Shin, Jae-Ho; Jin, Kyung-Hyun; Park, Hun-Kuk

    2015-07-01

    The study was to investigate the changes in biochemical properties of activated mature CD8+ T cells related to apoptosis at a molecular level. We confirmed the activation and apoptosis of CD8+ T cells by fluorescence-activated cell sorting and atomic force microscopy and then performed Raman spectral measurements on activated mature CD8+ T cells and cellular deoxyribose nucleic acid (DNA). In the activated mature CD8+ T cells, there were increases in protein spectra at 1002 and 1234 cm-1. In particular, to assess the apoptosis-related DNA spectral signatures, we investigated the spectra of the cellular DNA isolated from resting and activated mature CD8+ T cells. Raman spectra at 765 to 786 cm-1 and 1053 to 1087 cm-1 were decreased in activated mature DNA. In addition, we analyzed Raman spectrum using the multivariate statistical method including principal component analysis. Raman spectra of activated mature DNA are especially well-discriminated from those of resting DNA. Our findings regarding the biochemical and structural changes associated with apoptosis in activated mature T cells and cellular DNA according to Raman spectroscopy provide important insights into allospecific immune responses generated after organ transplantation, and may be useful for therapeutic manipulation of the immune response.

  3. Interference with Ca2+ release activated Ca2+ (CRAC) channel function delays T-cell arrest in vivo

    Waite, Janelle C.; Vardhana, Santosh; Shaw, Patrick J.; Jang, Jung-Eun; McCarl, Christie-Ann; Cameron, Thomas O; Feske, Stefan; Dustin, Michael L

    2013-01-01

    Entry of lymphocytes into secondary lymphoid organs (SLOs) involves intravascular arrest and intracellular calcium ion ([Ca2+]i) elevation. TCR activation triggers increased [Ca2+]i and can arrest T-cell motility in vitro. However the requirement for [Ca2+]i elevation in arresting T cells in vivo has not been tested. Here, we have manipulated the Ca2+ release-activated Ca2+ (CRAC) channel pathway required for [Ca2+]i elevation in T cells through genetic deletion of stromal interaction molecul...

  4. Cell surface polypeptides of murine T-cell clones expressing cytolytic or amplifier activity.

    Sarmiento, M.; Glasebrook, A L; Fitch, F. W.

    1980-01-01

    Murine cytolytic T-cell and amplifier T-cell clones derived from secondary unidirectional mixed leukocyte cultures were labeled with 125I by the lactoperoxidase method and their polypeptide profiles were analyzed by NaDodSO4/polyacrylamide gel electrophoresis. All cytolytic T-cell clones derived from the same mouse strain yeilded similar cell surface polypeptide profiles. However, profiles obtained with three amplifier T-cell clones were strikingly different from each other as well as from th...

  5. NF-kappa B activity in T cells stably expressing the Tax protein of human T cell lymphotropic virus type I

    Lacoste, J.; Cohen, L.; Hiscott, J. (Lady Davis Institute for Medical Research, Sir Mortimer B. Davis-Jewish General Hospital, Montreal, Quebec (Canada))

    1991-10-01

    The effect of constitutive Tax expression on the interaction of NF-{kappa} B with its recognition sequence and on NF-{kappa} B-dependent gene expression was examined in T lymphoid Jurkat cell lines (19D and 9J) stably transformed with a Tax expression vector. Tax expressing T cell lines contained a constitutive level of NF-{kappa} B binding activity, detectable by mobility shift assay and uv cross-linking using a palindromic NF-{kappa} B probe homologous to the interferon beta PRDII site. In Jurkat and NC2.10 induction with phorbol esters resulted in the appearance of new DNA binding proteins of 85, 75, and 54 kDa, whereas in Tax expressing cells the 85-kDa protein and a 92-kDa DNA binding protein were constitutively induced. Expression of Tax protein in 19D and 9J resulted in transcription of the endogenous NF-kappa B-dependent granulocyte-macrophage colony stimulating factor gene and increased basal level expression of transfected NF-kappa B-regulated promoters. Nonetheless transcription of both the endogenous and the transfected gene was inducible by PMA treatment. Tax expression in Jurkat T cells may alter the stoichiometry of NF-kappa B DNA binding proteins and thus change the expression of NF-kappa B-regulated promoters.

  6. Activation of regulatory T cells during inflammatory response is not an exclusive property of stem cells.

    Jan-Hendrik Gosemann

    Full Text Available BACKGROUND: Sepsis and systemic-inflammatory-response-syndrome (SIRS remain major causes for fatalities on intensive care units despite up-to-date therapy. It is well accepted that stem cells have immunomodulatory properties during inflammation and sepsis, including the activation of regulatory T cells and the attenuation of distant organ damage. Evidence from recent work suggests that these properties may not be exclusively attributed to stem cells. This study was designed to evaluate the immunomodulatory potency of cellular treatment during acute inflammation in a model of sublethal endotoxemia and to investigate the hypothesis that immunomodulations by cellular treatment during inflammatory response is not stem cell specific. METHODOLOGY/PRINCIPAL FINDINGS: Endotoxemia was induced via intra-peritoneal injection of lipopolysaccharide (LPS in wild type mice (C3H/HeN. Mice were treated with either vital or homogenized amniotic fluid stem cells (AFS and sacrificed for specimen collection 24 h after LPS injection. Endpoints were plasma cytokine levels (BD™ Cytometric Bead Arrays, T cell subpopulations (flow-cytometry and pulmonary neutrophil influx (immunohistochemistry. To define stem cell specific effects, treatment with either vital or homogenized human-embryonic-kidney-cells (HEK was investigated in a second subset of experiments. Mice treated with homogenized AFS cells showed significantly increased percentages of regulatory T cells and Interleukin-2 as well as decreased amounts of pulmonary neutrophils compared to saline-treated controls. These results could be reproduced in mice treated with vital HEK cells. No further differences were observed between plasma cytokine levels of endotoxemic mice. CONCLUSIONS/SIGNIFICANCE: The results revealed that both AFS and HEK cells modulate cellular immune response and distant organ damage during sublethal endotoxemia. The observed effects support the hypothesis, that immunomodulations are not

  7. Retinal pigment epithelial cells upregulate expression of complement factors after co-culture with activated T cells

    Juel, Helene Bæk; Kaestel, Charlotte; Folkersen, Lasse;

    2011-01-01

    In this study we examined the effect of T cell-derived cytokines on retinal pigment epithelial (RPE) cells with respect to expression of complement components. We used an in vitro co-culture system in which CD3/CD28-activated human T cells were separated from the human RPE cell line (ARPE-19) by a...... membrane. Differential gene expression in the RPE cells of complement factor genes was identified using gene arrays, and selected gene transcripts were validated by q-RT-PCR. Protein expression was determined by ELISA and immunoblotting. Co-culture with activated T cells increased RPE mRNA and/or protein...... expression of complement components C3, factors B, H, H-like 1, CD46, CD55, CD59, and clusterin, in a dose-dependent manner. Soluble factors derived from activated T cells are capable of increasing expression of complement components in RPE cells. This is important for the further understanding of...

  8. Impact of MAPK pathway activation in BRAFV600 melanoma on T cell and Dendritic Cell function

    Patrick Alexander Ott

    2013-10-01

    Full Text Available Constitutive upregulation of the MAPK pathway by a BRAFV600 mutation occurs in about half of melanomas. This leads to increased oncogenic properties such as tumor cell invasion, metastatic potential, and resistance to apoptosis. Blockade of the MAPK pathway with highly specific kinase inhibitors induces unprecedented tumor response rates in patients with advanced BRAFV600 mutant melanoma. Immune checkpoint blockade with monoclonal antibodies targeting CTLA-4 and PD-1/PD-L1 has also demonstrated striking anti-tumor activity in patients with advanced melanoma. Tumor responses are likely limited by multiple additional layers of immune suppression in the tumor microenvironment. There is emerging preclinical and clinical evidence suggesting that MAPK inhibition has a beneficial effect on the immunosuppressive tumor microenvironment, providing a strong rationale for combined immunotherapy and MAPK pathway inhibition in melanoma. The T cell response has been the main focus in the studies reported to date. Since dendritic cells (DCs are important in the induction of tumor-specific T cell responses, the impact of MAPK pathway activation in melanoma on DC function is critical for the melanoma directed immune response. BRAFV600E melanoma cells modulate DC through the MAPK pathway because its blockade in melanoma cells can reverse suppression of DC function. As both MEK/BRAF inhibition and immune checkpoint blockade have recently taken center stage in the treatment of melanoma, a deeper understanding of how MAPK pathway inhibition affects the tumor immune response is needed.

  9. Activation-induced and damage-induced cell death in aging human T cells.

    Sikora, Ewa

    2015-11-01

    In multicellular organisms the proper system functionality is ensured by the balance between cell division, differentiation, senescence and death. This balance is changed during aging. Immunosenescence plays a crucial role in aging and leads to the shrinkage of T cell repertoire and the propensity to apoptosis. The elimination of expanded T cells at the end of immune response is crucial to maintain homeostasis and avoid any uncontrolled inflammation. Resting mature T lymphocytes, when activated via their antigen-specific receptor (TCR) and CD28 co-receptor, start to proliferate and then undergo the so called activation induced cell death (AICD), which mechanistically is triggered by the death receptor and leads to apoptosis. T lymphocytes, like other cells, are also exposed to damage, which can trigger the so called damage-induced cell death (DICD). It was hypothesized that oxidative stress and chronic antigenic load increasing with age reduced lymphocyte susceptibility to DICD and enhanced a proinflamatory status leading to increased AICD. However, data collected so far are inconsistent and does not support this assumption. Systematic and comprehensive studies are still needed for conclusive elucidation of the role of AICD and DICD in human immunosenescence, including the role of autophagy and necroptosis in the processes. PMID:25843236

  10. Loss of TAL-1 protein activity induces premature apoptosis of Jurkat leukemic T cells upon medium depletion.

    Leroy-Viard, K; Vinit, M A; Lecointe, N; JOUAULT, H.; Hibner, U; Roméo, P H; Mathieu-Mahul, D

    1995-01-01

    Transcriptional activation of the tal-1 gene occurs in -30% of patients with T cell Acute Lymphoblastic Leukemia and is therefore likely to be involved in human T cell leukemogenesis. However, the TAL-1 protein functional properties involved in this process have not been assessed so far. We have derived a clonal subline of the Jurkat T cell line which produced solely a mutant truncated form of TAL-1 protein. Sequencing of genomic DNA and cDNAs showed that the only transcribed tal-1 allele of ...

  11. Activation of p38α in T cells regulates the intestinal host defense against attaching and effacing bacterial infections

    Shim, Eun-Jin; Bang, Bo Ram; Kang, Seung-Goo; Ma, Jianhui; Otsuka, Motoyuki; Kang, Jiman; Stahl, Martin; Han, Jiahuai; Xiao, Changchun; Vallance, Bruce A.; Kang, Young Jun

    2013-01-01

    Intestinal infections by attaching and effacing (A/E) bacterial pathogens cause severe colitis and bloody diarrhea. Although p38α in intestine epithelial cells (IEC) plays an important role in promoting protection against A/E bacteria by regulating T cell recruitment, its impact on immune responses remains unclear. In this study, we show that activation of p38α in T cells is critical for the clearance of the A/E pathogen Citrobacter rodentium. Mice deficient of p38α in T cells, but not in mac...

  12. The early activation marker CD69 regulates the expression of chemokines and CD4 T cell accumulation in intestine.

    Katarina Radulovic

    Full Text Available Migration of naïve and activated lymphocytes is regulated by the expression of various molecules such as chemokine receptors and ligands. CD69, the early activation marker of C-type lectin domain family, is also shown to regulate the lymphocyte migration by affecting their egress from the thymus and secondary lymphoid organs. Here, we aimed to investigate the role of CD69 in accumulation of CD4 T cells in intestine using murine models of inflammatory bowel disease. We found that genetic deletion of CD69 in mice increases the expression of the chemokines CCL-1, CXCL-10 and CCL-19 in CD4(+ T cells and/or CD4(- cells. Efficient in vitro migration of CD69-deficient CD4 T cells toward the chemokine stimuli was the result of increased expression and/or affinity of chemokine receptors. In vivo CD69(-/- CD4 T cells accumulate in the intestine in higher numbers than B6 CD4 T cells as observed in competitive homing assay, dextran sodium sulphate (DSS-induced colitis and antigen-specific transfer colitis. In DSS colitis CD69(-/- CD4 T cell accumulation in colonic lamina propria (cLP was associated with increased expression of CCL-1, CXCL-10 and CCL-19 genes. Furthermore, treatment of DSS-administrated CD69(-/- mice with the mixture of CCL-1, CXCL-10 and CCL-19 neutralizing Abs significantly decreased the histopathological signs of colitis. Transfer of OT-II×CD69(-/- CD45RB(high CD4 T cells into RAG(-/- hosts induced CD4 T cell accumulation in cLP. This study showed CD69 as negative regulator of inflammatory responses in intestine as it decreases the expression of chemotactic receptors and ligands and reduces the accumulation of CD4 T cells in cLP during colitis.

  13. Constitutive STAT3 activation in intestinal T cells from patients with Crohn's disease

    Lovato, Paola; Brender, Christine; Agnholt, Jørgen;

    2003-01-01

    Via cytoplasmic signal transduction pathways, cytokines induce a variety of biological responses and modulate the outcome of inflammatory diseases and malignancies. Crohn's disease is a chronic inflammatory bowel disease of unknown etiology. Perturbation of the intestinal cytokine homeostasis is...... believed to play a pivotal role, but the pathogenesis of Crohn's disease is not fully understood. Here, we study intestinal T cells from Crohn's disease and healthy volunteers. We show that STAT3 and STAT4 are constitutively activated in Crohn's patients but not in healthy volunteers. The activation is...... specific, because other STAT proteins are not constitutively activated. Furthermore, the STAT3 regulated protein, SOCS3, is also constitutively expressed in Crohn's patients but not in healthy volunteers. Taken together, these data provide evidence of abnormal STAT/SOCS signaling in Crohn's disease. This...

  14. Inhibition of NAMPT pathway by FK866 activates the function of p53 in HEK293T cells

    Highlights: ► In 293T cells, p53 is considered to be inactive due to its interaction with the large T-antigen. ► Acetylation of p53 at lysine 382 is important for its functional activation. ► First evidence to document the presence of a functional p53 in 293T cells. ► Inhibition of NAMPT/SIRT pathway by FK866 in 293T cells increases the functional activity of p53. ► This activation of p53 involves reversible acetylation of p53 at lysine 382. -- Abstract: Inactivation of p53 protein by endogenous and exogenous carcinogens is involved in the pathogenesis of different human malignancies. In cancer associated with SV-40 DNA tumor virus, p53 is considered to be non-functional mainly due to its interaction with the large T-antigen. Using the 293T cell line (HEK293 cells transformed with large T antigen) as a model, we provide evidence that p53 is one of the critical downstream targets involved in FK866-mediated killing of 293T cells. A reduced rate of apoptosis and an increased number of cells in S-phase was accompanied after knockdown of p53 in these cells. Inhibition of NAMPT by FK866, or inhibition of SIRT by nicotinamide decreased proliferation and triggered death of 293T cells involving the p53 acetylation pathway. Additionally, knockdown of p53 attenuated the effect of FK866 on cell proliferation, apoptosis, and cell cycle arrest. The data presented here shed light on two important facts: (1) that p53 in 293T cells is active in the presence of FK866, an inhibitor of NAMPT pathway; (2) the apoptosis induced by FK866 in 293T cells is associated with increased acetylation of p53 at Lys382, which is required for the functional activity of p53.

  15. Activated human γδ T cells as stimulators of specific CD8+ T cell responses to subdominant Epstein Barr virus (EBV) epitopes: Potential for immunotherapy of cancer

    Landmeier, Silke; Altvater, Bianca; Pscherer, Sibylle; Juergens, Heribert; Varnholt, Lena; Hansmeier, Anna; Bollard, Catherine M.; Moosmann, Andreas; Bisping, Guido; Rossig, Claudia

    2009-01-01

    The efficacy of current cancer vaccines is limited by the functional heterogeneity and poor availability and expansion of professional antigen-presenting cells (APCs). Besides their potent innate effector properties, γδ T cells have been suggested to be involved in the initiation and maintenance of adaptive immune responses. Here, we investigated the capacity of human γδ T cells to induce expansion of virus-specific T cells to Epstein Barr virus (EBV) antigens. Aminobisphosphonate-stimulated ...

  16. Regulatory T cells suppress systemic and mucosal immune activation to control intestinal inflammation.

    Izcue, Ana; Coombes, Janine L; Powrie, Fiona

    2006-08-01

    The gastrointestinal (GI) tract is the main interface where the body encounters exogenous antigens. It is crucial that the local response here is tightly regulated to avoid an immune reaction against dietary antigens and commensal flora while still mounting an efficient defense against pathogens. Faults in establishing intestinal tolerance can lead to disease, inducing local and often also systemic inflammation. Studies in human as well as in animal models suggest a role for regulatory T cells (Tregs) in maintaining intestinal homeostasis. Transfer of Tregs can not only prevent the development of colitis in animal models but also cure established disease, acting both systemically and at the site of inflammation. In this review, we discuss the major regulatory pathways, including transforming growth factor-beta (TGF-beta), interleukin-10 (IL-10), and cytotoxic T-lymphocyte antigen-4 (CTLA-4), and their role in Treg-mediated control of systemic and mucosal responses. In addition, we give an overview of the known mechanisms of lymphocyte migration to the intestine and discuss how CD103 expression can influence the balance between regulatory and effector T cells. Further understanding of the factors that control the activity of Tregs in different immune compartments may facilitate the design of strategies to target regulation in a tissue-specific way. PMID:16903919

  17. Id1 expression promotes peripheral CD4+ T cell proliferation and survival upon TCR activation without co-stimulation

    Highlights: •Id1 expression enables naïve T cell proliferation without anti-CD28 co-stimulation. •Id1 expression facilitates T cells survival when stimulated with anti-CD3. •Elevation of IL-2 production by Id1 contributes increased proliferation and survival. •Id1 potentiates NF-κB activation by anti-CD3 stimulation. -- Abstract: Although the role of E proteins in the thymocyte development is well documented, much less is known about their function in peripheral T cells. Here we demonstrated that CD4 promoter-driven transgenic expression of Id1, a naturally occurring dominant-negative inhibitor of E proteins, can substitute for the co-stimulatory signal delivered by CD28 to facilitate the proliferation and survival of naïve CD4+ cells upon anti-CD3 stimulation. We next discovered that IL-2 production and NF-κB activity after anti-CD3 stimulation were significantly elevated in Id1-expressing cells, which may be, at least in part, responsible for the augmentation of their proliferation and survival. Taken together, results from this study suggest an important role of E and Id proteins in peripheral T cell activation. The ability of Id proteins to by-pass co-stimulatory signals to enable T cell activation has significant implications in regulating T cell immunity

  18. Shock Waves Increase T-cell Proliferation or IL-2 Expression by Activating p38 MAP Kinase

    Tie-Cheng YU; Yi LIU; Yan TAN; Yanfang JIANG; Xueqing ZHENG; Xinxiang XU

    2004-01-01

    Shock waves were elicited by transient pressure disturbances, which could be used to treat musculoskeletal disorders. In present studies, we i. nvestigated whether the low-density shock waves (LDSWs), which are able to damage plasma membrane without impairing the vimentin or other organelles, might augment T-cell proliferation as well as IL-2 expression, and if mitogen activated protein kinase p38 (p38 MAPK)might be an underlying mechanism through which the LDSWs enhanced T-cell function. We found that the LDSWs increased activation of p38 MAPK in Jurkat T cells. The LDSWs alone didn't result in the T-cell proliferation and IL-2 expression. However, in combination with other stimuli, LDSWs could augment the T-cell proliferation and IL-2 expression. Inhibition of p38 MAPK using SB203580 reduced the stimulatory effects of the LDSWs, which indicated that the LDSWs enhanced IL-2 expression through a mechanism that involved p38 MAPK activation. We concluded that the p38 MAPK activation played a key role in the regulation of T cell function by the LDSWs.

  19. CD8+ Foxp3+ T cells share developmental and phenotypic features with classical CD4+ Foxp3+ regulatory T cells but lack potent suppressive activity.

    Mayer, Christian T.; Floess, Stefan; Baru, Abdul Mannan; Lahl, Katharina; Huehn, Jochen; Sparwasser, Tim

    2011-01-01

    "Suppressor T cells" were historically defined within the CD8(+) T-cell compartment and recent studies have highlighted several naturally occurring CD8(+) Foxp3(-) Treg populations. However, the relevance of CD8(+) Foxp3(+) T cells, which represent a minor population in both thymi and secondary lymphoid organs of nonmanipulated mice, remains unclear. We here demonstrate that de novo Foxp3 induction in peripheral CD8(+) Foxp3(-) T cells is counter-regulated by DC-mediated co-stimulation via CD...

  20. Human T cells express CD25 and Foxp3 upon activation and exhibit effector/memory phenotypes without any regulatory/suppressor function

    Godder Kamar

    2009-10-01

    Full Text Available Abstract Background Foxp3 has been suggested to be a standard marker for murine Tregs whereas its role as marker for human Tregs is controversial. While some reports have shown that human Foxp3+ T cells had no regulatory function others have shown their role in the inhibition of T cell proliferation. Methods T cell activation was performed by means of brayostatin-1/ionomycin (B/I, mixed lymphocyte reaction (MLR, and CD3/CD28 activation. T cell proliferation was performed using BrdU and CFSE staining. Flow cytometry was performed to determine Foxp3 expression, cell proliferation, viabilities and phenotype analyses of T cells. Results Both CD4+ and CD8+ T cells expressed Foxp3 upon activation in vitro. Expression of Foxp3 remained more stable in CD4+CD25+ T cells compared to that in CD8+CD25+ T cells. The CD4+CD25+Foxp3+ T cells expressed CD44 and CD62L, showing their effector and memory phenotypes. Both FoxP3- responder T cells and CD4+FoxP3+ T cells underwent proliferation upon CD3/CD28 activation. Conclusion Expression of Foxp3 does not necessarily convey regulatory function in human CD4+CD25+ T cells. Increased FoxP3 on CD44+ effector and CD44+CD62L+ memory T cells upon stimulation suggest the activation-induced regulation of FoxP3 expression.

  1. Acute plasma biomarkers of T cell activation set-point levels and of disease progression in HIV-1 infection.

    Anne-Sophie Liovat

    Full Text Available T cell activation levels, viral load and CD4(+ T cell counts at early stages of HIV-1 infection are predictive of the rate of progression towards AIDS. We evaluated whether the inflammatory profile during primary HIV-1 infection is predictive of the virological and immunological set-points and of disease progression. We quantified 28 plasma proteins during acute and post-acute HIV-1 infection in individuals with known disease progression profiles. Forty-six untreated patients, enrolled during primary HIV-1 infection, were categorized into rapid progressors, progressors and slow progressors according to their spontaneous progression profile over 42 months of follow-up. Already during primary infection, rapid progressors showed a higher number of increased plasma proteins than progressors or slow progressors. The plasma levels of TGF-β1 and IL-18 in primary HIV-1 infection were both positively associated with T cell activation level at set-point (6 months after acute infection and together able to predict 74% of the T cell activation variation at set-point. Plasma IP-10 was positively and negatively associated with, respectively, T cell activation and CD4(+ T cell counts at set-point and capable to predict 30% of the CD4(+ T cell count variation at set-point. Moreover, plasma IP-10 levels during primary infection were predictive of rapid progression. In primary infection, IP-10 was an even better predictor of rapid disease progression than viremia or CD4(+ T cell levels at this time point. The superior predictive capacity of IP-10 was confirmed in an independent group of 88 HIV-1 infected individuals. Altogether, this study shows that the inflammatory profile in primary HIV-1 infection is associated with T cell activation levels and CD4(+ T cell counts at set-point. Plasma IP-10 levels were of strong predictive value for rapid disease progression. The data suggest IP-10 being an earlier marker of disease progression than CD4(+ T cell counts or

  2. Regulatory T Cell Responses to High-Dose Methylprednisolone in Active Systemic Lupus Erythematosus.

    Alexis Mathian

    Full Text Available A slight increase in the proportion of circulating regulatory T (Treg cells has been reported in systemic lupus erythematosus (SLE patients taking oral prednisone. The effects of intravenous (IV high dose methylprednisolone (MP on Tregs have not yet been described, especially in active SLE.We prospectively analyzed the proportion of circulating CD4+ Treg cell subsets defined as follows: (1 naïve Treg (nTreg FoxP3lowCD45RA+ cells; (2 effector Treg (eTreg FoxP3highCD45RA- cells; and (3 non-suppressive FoxP3lowCD45RA- cells (non-regulatory Foxp3low T cells. Peripheral blood mononuclear cells of patients with active SLE were analyzed before the first infusion of IV high dose MP (day 0 and the following days (day 1, day 2, ±day 3 and ±day 8. The activity of SLE was assessed by the SLEDAI score.Seventeen patients were included. Following MP infusions, the median (range percentage of eTregs significantly increased from 1.62% (0.53-8.43 at day 0 to 2.80% (0.83-14.60 at day 1 (p = 0.003 versus day 0, 4.64% (0.50-12.40 at day 2 (p = 0.06 versus day 1 and 7.50% (1.02-20.70 at day 3 (p = 0.008 versus day 2, and declined to baseline values at day 8. Expanding eTreg cells were actively proliferating, as they expressed Ki-67. The frequency of non-regulatory FoxP3low T cells decreased from 6.39% (3.20-17.70 at day 0 to 4.74% (1.03-9.72 at day 2 (p = 0.005; nTreg frequency did not change. All patients clinically improved immediately after MP pulses. The absence of flare after one year of follow up was associated with a higher frequency of eTregs at day 2.IV high dose MP induces a rapid, dramatic and transient increase in circulating regulatory T cells. This increase may participate in the preventive effect of MP on subsequent flares in SLE.

  3. β-Catenin Inhibits T Cell Activation by Selective Interference with Linker for Activation of T Cells–Phospholipase C-γ1 Phosphorylation

    Driessens, Gregory; Zheng, Yan; Locke, Frederick; Cannon, Judy L.; Gounari, Fotini; Gajewski, Thomas F.

    2016-01-01

    Despite the defined function of the β-catenin pathway in thymocytes, its functional role in peripheral T cells is poorly understood. We report that in a mouse model, β-catenin protein is constitutively degraded in peripheral T cells. Introduction of stabilized β-catenin into primary T cells inhibited proliferation and cytokine secretion after TCR stimulation and blunted effector cell differentiation. Functional and biochemical studies revealed that β-catenin selectively inhibited linker for activation of T cells phosphorylation on tyrosine 136, which was associated with defective phospholipase C-γ1 phosphorylation and calcium signaling but normal ERK activation. Our findings indicate that β-catenin negatively regulates T cell activation by a previously undescribed mechanism and suggest that conditions under which β-catenin might be inducibly stabilized in vivo would be inhibitory for T cell-based immunity. PMID:21149602

  4. Cish actively silences TCR signaling in CD8+ T cells to maintain tumor tolerance

    Douglas C Palmer; Guittard, Geoffrey C.; Franco, Zulmarie; Crompton, Joseph G.; Eil, Robert L; Patel, Shashank J.; Ji, Yun; van Panhuys, Nicholas; Klebanoff, Christopher A.; Sukumar, Madhusudhanan; Clever, David; Chichura, Anna; Roychoudhuri, Rahul; Varma, Rajat; Wang, Ena

    2015-01-01

    Improving the functional avidity of effector T cells is critical in overcoming inhibitory factors within the tumor microenvironment and eliciting tumor regression. We have found that Cish, a member of the suppressor of cytokine signaling (SOCS) family, is induced by TCR stimulation in CD8+ T cells and inhibits their functional avidity against tumors. Genetic deletion of Cish in CD8+ T cells enhances their expansion, functional avidity, and cytokine polyfunctionality, resulting in pronounced a...

  5. Activation of resting T cells: distinct roles of intact accessory cells, phorbol myristate acetate and interleukin 1

    Davis, L.; Lipsky, P.E.

    1986-03-05

    The accessory cell (AC) signals involved in the activation of resting guinea pig T lymphocytes stimulated with mitogen (PHA), or the calcium ionophore, ionomycin (Ion) were examined. Activation of T cells was assessed by cell cycle analysis after acridine orange staining and /sup 3/H-thymidine incorporation. PHA-stimulated T cells depleted of all AC were unable to respond in the presence of phorbol myristate acetate (PMA), and/or interleukin 1 (IL 1). With suboptimal numbers of AC, PMA greatly augmented the number of T cells activated by PHA to enter and progress through the cell cycle, but only when present during the first few hours of culture. By contrast, IL 1 had little effect on the number of cells entering the cell cycle, although it enhanced S phase entry of the activated cells. IL 1 augmented DNA synthesis when added initially or later in culture. In contrast to the effects noted with PHA, PMA promoted activation and DNA synthesis of the majority of Ion stimulated cells in the complete absence of AC. IL 1 alone could not support Ion induced T cell activation although it enhanced T cell DNA synthesis in cultures stimulated by PMA and Ion. These studies indicate that intact AC, IL 1 and PMA-like signals play distinct roles in the progression of T cells through the initial cell cycle. Stimulation by Ion requires only PMA whereas PHA responses require intact AC and can be amplified by PMA. The major effect of IL 1 is to promote S phase entry of activated T cells.

  6. Regulation of ITAM adaptor molecules and their receptors by inhibition of calcineurin-NFAT signalling during late stage osteoclast differentiation

    Highlights: ► Calcineurin/NFAT inhibitors FK506 and VIVIT treated human PBMC derived osteoclasts in vitro. ► Differential regulation of ITAM receptors and adaptor molecules by calcineurin/NFAT inhibitors. ► FK506 and VIVIT suppress ITAM factors during late phase osteoclast differentiation. -- Abstract: Osteoclasts are specialised bone resorptive cells responsible for both physiological and pathological bone loss. Osteoclast differentiation and activity is dependent upon receptor activator NF-kappa-B ligand (RANKL) interacting with its receptor RANK to induce the transcription factor, nuclear factor of activated T-cells, cytoplasmic, calcineurin-dependent 1 (NFATc1). The immunoreceptor tyrosine-based activation motif (ITAM)-dependent pathway has been identified as a co-stimulatory pathway in osteoclasts. Osteoclast-associated receptor (OSCAR) and triggering receptor expressed in myeloid cells (TREM2) are essential receptors that pair with adaptor molecules Fc receptor common gamma chain (FcRγ) and DNAX-activating protein 12 kDa (DAP12) respectively to induce calcium signalling. Treatment with calcineurin-NFAT inhibitors, Tacrolimus (FK506) and the 11R-VIVIT (VIVIT) peptide, reduces NFATc1 expression consistent with a reduction in osteoclast differentiation and activity. This study aimed to investigate the effects of inhibiting calcineurin-NFAT signalling on the expression of ITAM factors and late stage osteoclast genes including cathepsin K (CathK), Beta 3 integrin (β3) and Annexin VIII (AnnVIII). Human peripheral blood mononuclear cells (PBMCs) were differentiated with RANKL and macrophage-colony stimulating factor (M-CSF) over 10 days in the presence or absence of FK506 or VIVIT. Osteoclast formation (as assessed by tartrate resistant acid phosphatase (TRAP)) and activity (assessed by dentine pit resorption) were significantly reduced with treatment. Quantitative real-time polymerase chain reaction (qRT-PCR) analysis demonstrated that FK506 treatment

  7. Regulation of ITAM adaptor molecules and their receptors by inhibition of calcineurin-NFAT signalling during late stage osteoclast differentiation

    Zawawi, M.S.F. [Universiti Sains Malaysia (USM) (Malaysia); Discipline of Anatomy and Pathology, School of Medical Sciences, University of Adelaide, Adelaide, SA 5005 (Australia); Dharmapatni, A.A.S.S.K.; Cantley, M.D. [Discipline of Anatomy and Pathology, School of Medical Sciences, University of Adelaide, Adelaide, SA 5005 (Australia); McHugh, K.P. [University of Florida, College of Dentistry, Fl (United States); Haynes, D.R. [Discipline of Anatomy and Pathology, School of Medical Sciences, University of Adelaide, Adelaide, SA 5005 (Australia); Crotti, T.N., E-mail: tania.crotti@adelaide.edu.au [Discipline of Anatomy and Pathology, School of Medical Sciences, University of Adelaide, Adelaide, SA 5005 (Australia)

    2012-10-19

    Highlights: Black-Right-Pointing-Pointer Calcineurin/NFAT inhibitors FK506 and VIVIT treated human PBMC derived osteoclasts in vitro. Black-Right-Pointing-Pointer Differential regulation of ITAM receptors and adaptor molecules by calcineurin/NFAT inhibitors. Black-Right-Pointing-Pointer FK506 and VIVIT suppress ITAM factors during late phase osteoclast differentiation. -- Abstract: Osteoclasts are specialised bone resorptive cells responsible for both physiological and pathological bone loss. Osteoclast differentiation and activity is dependent upon receptor activator NF-kappa-B ligand (RANKL) interacting with its receptor RANK to induce the transcription factor, nuclear factor of activated T-cells, cytoplasmic, calcineurin-dependent 1 (NFATc1). The immunoreceptor tyrosine-based activation motif (ITAM)-dependent pathway has been identified as a co-stimulatory pathway in osteoclasts. Osteoclast-associated receptor (OSCAR) and triggering receptor expressed in myeloid cells (TREM2) are essential receptors that pair with adaptor molecules Fc receptor common gamma chain (FcR{gamma}) and DNAX-activating protein 12 kDa (DAP12) respectively to induce calcium signalling. Treatment with calcineurin-NFAT inhibitors, Tacrolimus (FK506) and the 11R-VIVIT (VIVIT) peptide, reduces NFATc1 expression consistent with a reduction in osteoclast differentiation and activity. This study aimed to investigate the effects of inhibiting calcineurin-NFAT signalling on the expression of ITAM factors and late stage osteoclast genes including cathepsin K (CathK), Beta 3 integrin ({beta}3) and Annexin VIII (AnnVIII). Human peripheral blood mononuclear cells (PBMCs) were differentiated with RANKL and macrophage-colony stimulating factor (M-CSF) over 10 days in the presence or absence of FK506 or VIVIT. Osteoclast formation (as assessed by tartrate resistant acid phosphatase (TRAP)) and activity (assessed by dentine pit resorption) were significantly reduced with treatment. Quantitative real

  8. Multifunctional CD4 T Cell Responses in Patients with Active Tuberculosis

    Qiu, Zhengang; Zhang, Mingxia; Zhu, Yuzhen; Zheng, Feiqun; Lu, Puxuan; Liu, Haiying; Michael W Graner; Zhou, Boping; Chen, Xinchun

    2012-01-01

    The roles of multifunctional CD4 T cells in human tuberculosis are not well defined. In this study, we found that patients with tuberculosis had decreased PMA/ionomycin stimulated multifunctional CD4 T cells, and increased Mycobacterium tuberculosis antigen-specific multifunctional CD4 T cells, when compared to individuals with latent tuberculosis infection and healthy controls. PMA/ionomycin stimulated IFN-γ+IL-2+TNF-α+ CD4 T cell responses were decreased in patients with smear-positive tube...

  9. GD2-specific CAR T Cells Undergo Potent Activation and Deletion Following Antigen Encounter but can be Protected From Activation-induced Cell Death by PD-1 Blockade.

    Gargett, Tessa; Yu, Wenbo; Dotti, Gianpietro; Yvon, Eric S; Christo, Susan N; Hayball, John D; Lewis, Ian D; Brenner, Malcolm K; Brown, Michael P

    2016-06-01

    Chimeric antigen receptor (CAR) T cells have shown great promise in the treatment of hematologic malignancies but more variable results in the treatment of solid tumors and the persistence and expansion of CAR T cells within patients has been identified as a key correlate of antitumor efficacy. Lack of immunological "space", functional exhaustion, and deletion have all been proposed as mechanisms that hamper CAR T-cell persistence. Here we describe the events following activation of third-generation CAR T cells specific for GD2. CAR T cells had highly potent immediate effector functions without evidence of functional exhaustion in vitro, although reduced cytokine production reversible by PD-1 blockade was observed after longer-term culture. Significant activation-induced cell death (AICD) of CAR T cells was observed after repeated antigen stimulation, and PD-1 blockade enhanced both CAR T-cell survival and promoted killing of PD-L1(+) tumor cell lines. Finally, we assessed CAR T-cell persistence in patients enrolled in the CARPETS phase 1 clinical trial of GD2-specific CAR T cells in the treatment of metastatic melanoma. Together, these data suggest that deletion also occurs in vivo and that PD-1-targeted combination therapy approaches may be useful to augment CAR T-cell efficacy and persistence in patients. PMID:27019998

  10. Extracellular signal-regulated kinase 1/2 signalling in SLE T cells is influenced by oestrogen and disease activity.

    Gorjestani, S; Rider, V; Kimler, B F; Greenwell, C; Abdou, N I

    2008-06-01

    Systemic lupus erythematosus (SLE) is an autoimmune disease that occurs primarily in women of reproductive age. The disease is characterized by exaggerated T-cell activity and abnormal T-cell signalling. The mitogen-activated protein kinase (MAPK) pathway is involved in the maintenance of T-cell tolerance that fails in patients with SLE. Oestrogen is a female sex hormone that binds to nuclear receptors and alters the rate of gene transcription. Oestrogen can also act through the plasma membrane and rapidly stimulate second messengers including calcium flux and kinase activation. In this study, we investigated whether oestrogen influences the activation of MAPK signalling through the phosphorylation of extracellular signal-regulated kinase 1/2 (ERK1/2) in activated SLE T cells. SLE and control T cells were cultured in serum-free medium without and with oestradiol (10(-7) M) for 18 h. The T cells were activated with phorbol 12 myristate 13-acetate and ionomycin for various time points (0-60 min), and the amount of phosphorylated ERK1/2 was measured by immunoblotting. There were no differences in ERK1/2 phosphorylation between SLE and control T cells at 5 and 15 min after the activation stimulus. However, comparison between the amount of phosphorylated ERK1/2 in SLE T cells from the same patients cultured without and with oestradiol showed a significant oestrogen-dependent suppression (P=0.48) of ERK1/2 in patients with inactive/mild systemic lupus erythematosus disease activity index (SLEDAI) (0-2) compared with patients with moderate (4-6) or active (8-12) SLEDAI scores. These results suggest that the suppression of MAPK through ERK1/2 phosphorylation is sensitive to oestradiol in patients with inactive or mild disease, but the sensitivity is not maintained when disease activity increases. Furthermore, studies are now necessary to understand the mechanisms by which oestrogen influences MAPK activation in SLE T cells. PMID:18539708

  11. Alpha 4 integrin directs virus-activated CD8+ T cells to sites of infection

    Christensen, Jan Pravsgaard; Andersson, E C; Scheynius, A; Marker, O; Thomsen, Allan Randrup

    1995-01-01

    response is induced, which is associated with marked CD8+ cell-mediated inflammation. Two expressions of LCMV-induced inflammation were studied: meningitis induced by intracerebral infection and adoptive transfer of virus-specific delayed-type hypersensitivity. Our previous studies have shown that LCMV...... infection results in the appearance of activated CD8+ cells with an increased expression of VLA-4. In this study we have compared various T cell high and low responder situations, and these experiments revealed that acute inflammation correlates directly with VLA-4 expression on splenic CD8+ cells. This...... ability to transfer virus-specific, delayed-type hypersensitivity when the donor cells were given i.v., but not when the cells were injected directly into the test site. Co-transfer of CD8-depleted cells with anti-VLA-4-blocked cells did not reveal any cooperation. Taken together, these results indicate...

  12. Curcumin induces apoptotic cell death of activated human CD4+ T cells via increasing endoplasmic reticulum stress and mitochondrial dysfunction.

    Zheng, Min; Zhang, Qinggao; Joe, Yeonsoo; Lee, Bong Hee; Ryu, Do Gon; Kwon, Kang Beom; Ryter, Stefan W; Chung, Hun Taeg

    2013-03-01

    Curcumin, a natural polyphenolic antioxidant compound, exerts well-known anti-inflammatory and immunomodulatory effects, the latter which can influence the activation of immune cells including T cells. Furthermore, curcumin can inhibit the expression of pro-inflammatory cytokines and chemokines, through suppression of the NF-κB signaling pathway. The beneficial effects of curcumin in diseases such as arthritis, allergy, asthma, atherosclerosis, diabetes and cancer may be due to its immunomodulatory properties. We studied the potential of curcumin to modulate CD4+ T cells-mediated autoimmune disease, by examining the effects of this compound on human CD4+ lymphocyte activation. Stimulation of human T cells with PHA or CD3/CD28 induced IL-2 mRNA expression and activated the endoplasmic reticulum (ER) stress response. The treatment of T cells with curcumin induced the unfolded protein response (UPR) signaling pathway, initiated by the phosphorylation of PERK and IRE1. Furthermore, curcumin increased the expression of the ER stress associated transcriptional factors XBP-1, cleaved p50ATF6α and C/EBP homologous protein (CHOP) in human CD4+ and Jurkat T cells. In PHA-activated T cells, curcumin further enhanced PHA-induced CHOP expression and reduced the expression of the anti-apoptotic protein Bcl-2. Finally, curcumin treatment induced apoptotic cell death in activated T cells via eliciting an excessive ER stress response, which was reversed by the ER-stress inhibitor 4-phenylbutyric acid or transfection with CHOP-specific siRNA. These results suggest that curcumin can impact both ER stress and mitochondria functional pathways, and thereby could be used as a promising therapy in the context of Th1-mediated autoimmune diseases. PMID:23415873

  13. A Subset of Protective γ9δ2 T Cells Is Activated by Novel Mycobacterial Glycolipid Components.

    Xia, Mei; Hesser, Danny C; De, Prithwiraj; Sakala, Isaac G; Spencer, Charles T; Kirkwood, Jay S; Abate, Getahun; Chatterjee, Delphi; Dobos, Karen M; Hoft, Daniel F

    2016-09-01

    γ9δ2 T cells provide a natural bridge between innate and adaptive immunity, rapidly and potently respond to pathogen infection in mucosal tissues, and are prominently induced by both tuberculosis (TB) infection and bacillus Calmette Guérin (BCG) vaccination. Mycobacterium-expanded γ9δ2 T cells represent only a subset of the phosphoantigen {isopentenyl pyrophosphate [IPP] and (E)-4-hydroxy-3-methyl-but-2-enylpyrophosphate [HMBPP]}-responsive γ9δ2 T cells, expressing an oligoclonal set of T cell receptor (TCR) sequences which more efficiently recognize and inhibit intracellular Mycobacterium tuberculosis infection. Based on this premise, we have been searching for M. tuberculosis antigens specifically capable of inducing a unique subset of mycobacterium-protective γ9δ2 T cells. Our screening strategy includes the identification of M. tuberculosis fractions that expand γ9δ2 T cells with biological functions capable of inhibiting intracellular mycobacterial replication. Chemical treatments of M. tuberculosis whole-cell lysates (MtbWL) ruled out protein, nucleic acid, and nonpolar lipids as the M. tuberculosis antigens inducing protective γ9δ2 T cells. Mild acid hydrolysis, which transforms complex carbohydrate to monomeric residues, abrogated the specific activity of M. tuberculosis whole-cell lysates, suggesting that a polysaccharide was required for biological activity. Extraction of MtbWL with chloroform-methanol-water (10:10:3) resulted in a polar lipid fraction with highly enriched specific activity; this activity was further enriched by silica gel chromatography. A combination of mass spectrometry and nuclear magnetic resonance analysis of bioactive fractions indicated that 6-O-methylglucose-containing lipopolysaccharides (mGLP) are predominant components present in this active fraction. These results have important implications for the development of new immunotherapeutic approaches for prevention and treatment of TB. PMID:27297390

  14. Zizyphus lotus L. (Desf. modulates antioxidant activity and human T-cell proliferation

    Belarbi Meriem

    2010-09-01

    Full Text Available Abstract Background Zizyphus lotus L. (Desf. also known as Jujube, is a deciduous shrub which belongs to Rhamnaceae family. This plant is used in Algerian traditional medicine for its anti-diabetic, sedative, analgesic, anti-inflammatory and hypoglycaemic activities. In the present study, we determined the concentrations of different vitamins (vitamin A, C and E and fatty acids in root, stem, leaves, fruit pulp and seed of Zizyphus lotus L. (Desf. and assessed the effects of their aqueous extracts on antioxidant status and human T-cell proliferation. Methods Aqueous filtrates from different parts, i.e, root, leaf, stem, fruit pulp and seed, of Zizyphus lotus L. (Desf. were prepared. Vitamin C levels were determined by precipitating with 10% trichloroacetic acid and vitamin A and E were assessed by HPLC. Lipid composition of these extracts was determined by gas-liquid chromatography. Anti-oxidant capacity was evaluated by using anti-radical resistance kit [Kit Radicaux Libres (KRL@; Kirial International SA, Couternon, France]. T-cell blastogenesis was assessed by the incorporation of 3H-thymidine. IL-2 gene expression was evaluated by RT-qPCR. Results Our results show that fruit pulp contained higher vitamin A and C contents than other parts of the plant. Furthermore, the fruit pulp was the richest source of linoleic acid (18:2n-6, a precursor of n-6 fatty acids. Fruit seeds possessed higher vitamin C levels than leaves, roots and stem. The leaves were the richest source of vitamin E and linolenic acid (18:3n-3, a precursor of n-3 fatty acids. The antioxidant capacity of the different extracts, measured by KRL@ test, was as follows: pulp Zizyphus lotus L. (Desf. exerted immunosuppressive effects. Conclusion Seed extracts exerted the most potent immunosuppressive effects on T cell proliferation and IL-2 mRNA expression. The results of the present study are discussed in the light of their use to modulate the immune-mediated diseases.

  15. Normal T-cell telomerase activity and upregulation in human immunodeficiency virus-1 infection

    Wolthers, KC; Otto, SA; Wisman, GBA; Fleury, S; Reiss, P; ten Kate, RW; van der Zee, AGJ; Miedema, F

    1999-01-01

    In human immunodeficiency virus (HIV)-1 infection, decrease of telomere length is mainly found in CD8(+) T cells and not in CD4(+) T cells. Telomerase, a ribonucleoprotein enzyme that can synthesize telomeric sequence onto chromosomal ends, can compensate for telomere loss. Here, we investigated if

  16. Systemic T-cell activation in acute clinically isolated optic neuritis

    Roed, Hanne; Frederiksen, Jette; Langkilde, Annika Reynberg; Sørensen, Torben Lykke; Lauritzen, Martin; Sellebjerg, Finn

    We examined untreated 60 patients with acute monosymptomatic optic neuritis (ON). Patients examined early after onset showed increased expression of HLA-DR and CD45R0 on CD4 and CD8 T cells. Expression of HLA-DR on CD4 T cells was higher in patients without IgG oligoclonal bands. Expression of HL...

  17. NKG2D-Dependent Activation of Dendritic Epidermal T cells in Contact Hypersensitivity

    Nielsen, Morten Milek; Dyring-Andersen, Beatrice; Schmidt, Jonas Damgård;

    2015-01-01

    The interaction between keratinocytes (KC) and skin-resident immune cells plays an important role in induction of contact hypersensitivity (CHS). A specific subset of γδ T cells termed dendritic epidermal T cells (DETC) are located in mouse epidermis, and we have recently shown that DETC become a...

  18. Irradiated autologous T cells restore the in vitro responsiveness of PWM-activated peripheral blood lymphocytes from splenectomized individuals

    The in vitro immunoglobulin (Ig) secrection of pokeweed mitogen (PWM)-activated peripheral blood lymphocytes (PBL) from individuals splenectomized post-trauma was monitored with a protein A plaque-forming cell (PFC) assay. Cultures of unfractionated as well as reconstituted cultures of isolated erythrocyte rosette-forming (E-RFC)-positive (T lymphocytes) and E-RFC-negative (B lymphocytes) cells were established. Using unfractionated cells, the response was substantially reduced or absent, whereas cultures of autologous untreated B and 2000 rads irradited T cells restored the response to normal levels. Normal T cells were not able to stimulate patients' B cells to Ig-secretion and patients' untreated T cells did not induce plaque formation in normal B cells, whereas irradiated patients' T cells induced development of approximately 50% of the response induced by normal irradiated T cells. These results indicate that the immunological defect in splenectomized individuals is not merely restricted to a high level of radiosensitive T cell suppression but also involves an impaired B cell function and T/B cell cooperation. (author)

  19. Determination of the Absolute Number of Cytokine mRNA Molecules within Individual Activated Human T Cells

    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.

  20. Inhibitory receptor expression depends more dominantly on differentiation and activation than exhaustion of human CD8 T cells

    Amandine eLegat

    2013-12-01

    Full Text Available Under conditions of chronic antigen stimulation, such as persistent viral infection and cancer, CD8 T cells may diminish effector function, which has been termed exhaustion. Expression of inhibitory Receptors (iRs is often regarded as a hallmark of exhaustion. Here we studied the expression of eight different iRs by CD8 T cells of healthy humans, including CTLA-4, PD1, TIM3, LAG3, 2B4, BTLA, CD160 and KLRG-1. We show that many iRs are expressed upon activation, and with progressive differentiation to effector cells, even in absence of long-term (chronic antigenic stimulation. In particular, we evaluated the direct relationship between iR expression and functionality in CD8 T cells by using anti-CD3 and anti-CD28 stimulation to stimulate all cells and differentiation subsets. We observed a striking upregulation of certain iRs following the cytokine production wave, in agreement with the notion that iRs function as a negative feedback mechanism. Intriguingly, we found no major impairment of cytokine production in cells positive for a broad array of iRs, as previously shown for PD1 in healthy donors. Rather, the expression of the various iRs strongly correlated with T cell differentiation or activation states, or both. Furthermore, we analyzed CD8 T cells from lymph nodes (LNs of melanoma patients. Interestingly, we found altered iR expression and lower cytokine production by T cells from metastatic LNs, but also from non-metastatic LNs, likely due to mechanisms which are not related to exhaustion. Together, our data shows that expression of iRs per se does not mark dysfunctional cells, but is rather tightly linked to activation and differentiation. This study highlights the importance of considering the status of activation and differentiation for the study and the clinical monitoring of CD8 T cells.

  1. Increased expression of T-cell KV1.3 and KCa3.1 channels in the inflamed intestinal wall from patients with active ulcerative colitis

    Hansen, Lars Koch; Larsen, Dorte; Sadda, Veeranjaneyulu; Nielsen, Gorm; Klinge, Lone; Schaffalitzky de Muckadell, Ove B.; Knudsen, Torben; Kjeldsen, Jens; Köhler, Ralf

    INTRODUCTION: T-cell KV1.3 and KCa3.1 channels have been proposed to be important effector proteins during T-cell activation and also in autoimmune disease by controlling T-cell motility, cytokine production, and proliferation. The role of KV1.3 channels in ulcerative colitis (UC) has not been...

  2. Superantigen and HLA-DR ligation induce phospholipase-C gamma 1 activation in class II+ T cells

    Kanner, S B; Odum, Niels; Grosmaire, L;

    1992-01-01

    Bacterial enterotoxin superantigens bind directly to HLA class II molecules (HLA-DR) expressed on both APC and activated human T cells, and simultaneously bind to certain V beta chains of the TCR. In this report, we compared early T cell signaling events in human alloantigen-stimulated T cells when...... activated by HLA-DR ligation through antibody cross-linking or by direct enterotoxin superantigen binding. Both types of stimuli induced tyrosine phosphorylation of phosphatidylinositol-specific phospholipase C gamma 1 (PLC gamma 1) and an increase in intracellular calcium concentration; however......, superantigen-induced signaling was stronger than class II ligation alone. Antibody-mediated ligation of HLA-DR with CD3 resulted in augmented PLC gamma 1 activation and increased calcium mobilization, consistent with a mechanism of superantigen activity through a combination of class II and CD3/Ti signals. In...

  3. Regulation of Ras exchange factors and cellular localization of Ras activation by lipid messengers in T cells

    Jesse E. Jun

    2013-09-01

    Full Text Available The Ras-MAPK signaling pathway is highly conserved throughout evolution and is activated downstream of a wide range of receptor stimuli. Ras guanine nucleotide exchange factors (RasGEFs catalyze GTP loading of Ras and play a pivotal role in regulating receptor-ligand induced Ras activity. In T cells, three families of functionally important RasGEFs are expressed: RasGRF, RasGRP, and SOS-family GEFs.Early on it was recognized that Ras activation is critical for T cell development and that the RasGEFs play an important role herein. More recent work has revealed that nuances in Ras activation appear to significantly impact T cell development and selection. These nuances include distinct biochemical patterns of analog versus digital Ras activation, differences in cellular localization of Ras activation, and intricate interplays between the RasGEFs during distinct T cell developmental stages as revealed by various new mouse models. In many instances, the exact nature of these nuances in Ras activation or how these may result from fine-tuning of the RasGEFs is not understood.One large group of biomolecules critically involved in the control of Ras-GEFs´functions are lipid second messengers. Multiple, yet distinct lipid products are generated following T cell receptor (TCR stimulation and bind to different domains in the RasGRP and SOS RasGEFs to facilitate the activation of the membrane-anchored Ras GTPases. In this review we highlight how different lipid-based elements are generated by various enzymes downstream of the TCR and other receptors and how these dynamic and interrelated lipid products may fine-tune Ras activation by RasGEFs in developing T cells.

  4. Differential activation of proliferation and cytotoxicity in human T-cell lymphotropic virus type I Tax-specific CD8 T cells by an altered peptide ligand.

    Höllsberg, P; Weber, W E; Dangond, F; Batra, V; Sette, A.; Hafler, D A

    1995-01-01

    Human T-cell leukemia virus type I (HTLV-I) gives rise to a neurologic disease known as HTLV-I-associated myelopathy/tropical spastic paraparesis (HAM/TSP). Although the pathogenesis of the disease is unknown, the presence of a remarkably high frequency of Tax-specific, cytotoxic CD8 T cells may suggest a role of these cells in the development of HAM/TSP. Antigen-mediated signaling in a CD8 T-cell clone specific for the Tax(11-19) peptide of HTLV-I was studied using analog peptides substitute...

  5. T cell immune response is correlated with fibrosis and inflammatory activity in hepatitis B cirrhotics

    Jie-Ting Tang; Jing-Yuan Fang; Wei-Qi Gu; En-Lin Li

    2006-01-01

    AIM: To explore the relationship among interferon-γ (IFN-γ) activity, fibrogenesis, T cell immune responses and hepatic inflammatory activity.METHODS: Peripheral blood samples from a total of 43 hepatitis B cirrhotic patients (LC) and 19 healthy controls (NC) were collected to measure their serum levels of IFN-γ, interleukin-2 (IL-2), soluble interleukin-2 receptor (sIL-2R), interleukin-10 (IL-10) and three serological markers of fibrosis including hyaluronic acid (HA), procollagen type Ⅲ peptide (PⅢP), and type Ⅳ collagen were measured using a double antibody sandwich ELISA. Also,serum total bilirubin (TB) and alanine aminotransferase (ALT) were measured by routine measures.RESULTS: The concentrations of serological markers of fibrosis in patients with active cirrhosis (ALC) were significantly higher than those in stationary liver cirrhosis (SLC) or NC groups. The levels of serological markers in HBeAg-positive patients were significantly higher than those in HBeAg-negative patients. In SLC and ALC patients, a negative linear correlation was found between IFN-γ levels and the serological markers of fibrosis. IFN-γ and IL-2 levels in the ALC group were significantly higher than those in the SLC and NC groups, but the statistical difference was not significant between the latter two. In contrast, IL-10 levels in the SLC group were significantly higher than that in the NC group, but no significant difference was found between SLC and ALC groups. The sIL-2R level was elevated gradually in all these groups,and the differences were significant. Positive linear correlations were seen between IFN-γ activity and ALT levels (r = 0.339, P < 0.05), and IL-2 activity and TB levels (r = 0.517, P < 0.05). sIL-2R expression was positively correlated with both ALT and TB levels (r = 0.324, 0.455,P < 0.05), whereas there was no statistically significant correlation between IL-10 expression and serum ALT and TB levels (r = -0.102, -0.093, P > 0.05). Finally

  6. Regulation of cathepsin G reduces the activation of proinsulin-reactive T cells from type 1 diabetes patients.

    Fang Zou

    Full Text Available Autoantigenic peptides resulting from self-proteins such as proinsulin are important players in the development of type 1 diabetes mellitus (T1D. Self-proteins can be processed by cathepsins (Cats within endocytic compartments and loaded to major histocompatibility complex (MHC class II molecules for CD4(+ T cell inspection. However, the processing and presentation of proinsulin by antigen-presenting cells (APC in humans is only partially understood. Here we demonstrate that the processing of proinsulin by B cell or myeloid dendritic cell (mDC1-derived lysosomal cathepsins resulted in several proinsulin-derived intermediates. These intermediates were similar to those obtained using purified CatG and, to a lesser extent, CatD, S, and V in vitro. Some of these intermediates polarized T cell activation in peripheral blood mononuclear cells (PBMC from T1D patients indicative for naturally processed T cell epitopes. Furthermore, CatG activity was found to be elevated in PBMC from T1D patients and abrogation of CatG activity resulted in functional inhibition of proinsulin-reactive T cells. Our data suggested the notion that CatG plays a critical role in proinsulin processing and is important in the activation process of diabetogenic T cells.

  7. Telomerase activity is increased and telomere length shortened in T cells from blood of patients with atopic dermatitis and psoriasis

    Wu, Kehuai; Higashi, N; Hansen, E R;

    2000-01-01

    We studied telomerase activity and telomere length in PBMC and purified CD4(+) and CD8(+) T cells from blood obtained from a total of 32 patients with atopic dermatitis, 16 patients with psoriasis, and 30 normal controls. The telomerase activity was significantly increased in PBMC from the patients......(+) T cell subsets from normal donors. In conclusion, the increased telomerase activity and shortened telomere length indicates that T lymphocytes in atopic dermatitis and psoriasis are chronically stimulated and have an increased cellular turnover in vivo....

  8. 4-1BB signaling activates the t cell factor 1 effector/β-catenin pathway with delayed kinetics via ERK signaling and delayed PI3K/AKT activation to promote the proliferation of CD8+ T Cells.

    Do Y Lee

    Full Text Available 4-1BB (CD137, an inducible costimulatory molecule, strongly enhances the proliferation and effector function of CD8(+ T cells. Since the serine/threonine kinase, glycogen synthase kinase-3 (GSK-3, is involved in a variety of signaling pathways of cellular proliferation, migration, immune responses, and apoptosis, we examined whether 4-1BB signaling activates GSK-3/β-catenin signaling and downstream transcription factors to enhance the proliferation of CD8(+ T cells. 4-1BB signaling induces rapid activation of ERK and IκB degradation, and shows delayed activation of AKT at 24 h post 4-1BB stimulation on anti-CD3 activated T cells. ERK and AKT signals were required for sustained β-catenin levels by inactivating GSK-3, which was also observed with delayed kinetics after 4-1BB stimulation. As a transcriptional partner of β-catenin, 4-1BB signaling decreased levels of FOXO1 and increased levels of stimulatory TCF1 in CD8(+ T cells at 2-3 days but not at early time points after 4-1BB engagement. The enhanced proliferation of CD8(+ T cells due to 4-1BB signaling was completely abolished by treatment with the TCF1/β-catenin inhibitor quercetin. These results show that 4-1BB signaling enhances the proliferation of activated CD8(+ T cells by activating the TCF1/β-catenin axis via the PI3K/AKT/ERK pathway. As effects of 4-1BB on AKT, FOXO1, β-catenin and GSK-3β showed delayed kinetics it is likely that an intervening molecule induced by 4-1BB and ERK signaling in activated T cells is responsible for these effects. These effects were observed on CD8(+ but not on CD4(+ T cells. Moreover, 4-1BB appeared to be unique among several TNFRs tested in inducing increase in stimulatory over inhibitory TCF-1.

  9. Binding of recombinant T cell receptor ligands (RTL) to antigen presenting cells prevents upregulation of CD11b and inhibits T cell activation and transfer of experimental autoimmune encephalomyelitis

    Sinha, Sushmita; Miller, Lisa; Subramanian, Sandhya; McCarty, Owen; Proctor, Thomas; Meza-Romero, Roberto; Burrows, Gregory G.; Vandenbark, Arthur A.; Offner, Halina

    2010-01-01

    Recombinant T cell ligands (RTLs) ameliorate experimental autoimmune encephalomyelitis (EAE) in antigen specific manner. We evaluated effects of RTL401 (I-As α1β1 + PLP-139-151) on splenocytes from mice with EAE to study RTL- T cell-tolerance-inducing mechanisms. RTLs bound to B, macrophages and DCs, through RTL-MHC-α1β1 moiety. RTL binding reduced CD11b expression on splenic macrophages/DC, and RTL401-conditioned macrophages/DC, not B cells, inhibited T cell activation. Reduced ability of RT...

  10. Activation of endogenous c-fos proto-oncogene expression by human T-cell leukemia virus type I-encoded p40tax protein in the human T-cell line, Jurkat.

    Nagata, K.(Faculty of Pure and Applied Sciences, University of Tsukuba, Tsukuba, Japan); Ohtani, K; Nakamura, M.; Sugamura, K

    1989-01-01

    We examined the ability of the trans-acting factor p40tax of human T-cell leukemia virus type I (HTLV-I), which is thought to be a crucial molecule in T-cell transformation by HTLV-I, to activate expression of a set of endogenous cellular genes related to T-cell proliferation. For this purpose we established a subclone (JPX-9) of Jurkat cells that was stably transfected with an expression plasmid containing the p40tax gene, whose expression is definitely dependent on heavy-metal ions. Express...