Science.gov

Sample records for promote t-cell binding

  1. IL-15 prolongs CD154 expression on human CD4 T cells via STAT5 binding to the CD154 transcriptional promoter

    PubMed Central

    Lowe, RM; Genin, A; Orgun, N; Cron, RQ

    2014-01-01

    Activation induced CD154 expression on CD4 T cells is prolonged in systemic lupus erythematosus but the mechanism(s) for its dysregulation are unknown. The studies reported herein demonstrate that IL-15 is capable of prolonging CD154 expression on PHA activated CD4 T cells. Since IL-15 signals through STAT5, predicted STAT5 binding sites in the human CD154 transcriptional promoter were identified, and STAT5 binding to the proximal CD154 promoter in vitro and in vivo following primary CD4 T cell activation was demonstrated. Moreover, overexpression of wild-type(WT) STAT5 in primary human CD4 T cells augmented CD154 transcription, whereas overexpression of a dominant negative (DN) STAT5 protein inhibited CD154 transcription. Mutation of the most proximal STAT5 binding site in the CD154 promoter resulted in diminished DNA binding and reduced CD154 transcriptional activity. Interestingly, STAT5-specific siRNA inhibited CD154 surface expression at 48 but not 24 hours after T cell activation. Thus, these findings provide some of the first evidence to support a possible mechanistic link to explain how the overexpression of IL-15 observed in lupus patients may be involved in the prolonged expression of CD154 that has also been observed on lupus CD4 T cells. PMID:24500400

  2. Anti-TNF drives regulatory T cell expansion by paradoxically promoting membrane TNF-TNF-RII binding in rheumatoid arthritis.

    PubMed

    Nguyen, Dao Xuan; Ehrenstein, Michael R

    2016-06-27

    The interplay between inflammatory and regulatory pathways orchestrates an effective immune response that provides protection from pathogens while limiting injury to host tissue. Tumor necrosis factor (TNF) is a pivotal inflammatory cytokine, but there is conflicting evidence as to whether it boosts or inhibits regulatory T cells (T reg cells). In this study, we show that the therapeutic anti-TNF antibody adalimumab, but not the soluble TNF receptor etanercept, paradoxically promoted the interaction between monocytes and T reg cells isolated from patients with rheumatoid arthritis (RA). Adalimumab bound to monocyte membrane TNF from RA patients and unexpectedly enhanced its expression and its binding to TNF-RII expressed on T reg cells. As a consequence, adalimumab expanded functional Foxp3(+) T reg cells equipped to suppress Th17 cells through an IL-2/STAT5-dependent mechanism. Our data not only highlight the beneficial effect of membrane TNF on T reg cell numbers during chronic inflammation, but in addition reveal how a therapeutic antibody that is thought to act by simply blocking its target can enhance the regulatory properties of this proinflammatory cytokine. PMID:27270893

  3. Tumor-promoting function and prognostic significance of the RNA-binding protein T-cell intracellular antigen-1 in esophageal squamous cell carcinoma

    PubMed Central

    Fujita, Yuji; Naruto, Takuya; Kohmoto, Tomohiro; Miyakami, Yuko; Watanabe, Miki; Kudo, Yasusei; Fujiwara, Hitoshi; Ichikawa, Daisuke; Otsuji, Eigo; Imoto, Issei

    2016-01-01

    T-cell intracellular antigen-1 (TIA1) is an RNA-binding protein involved in many regulatory aspects of mRNA metabolism. Here, we report previously unknown tumor-promoting activity of TIA1, which seems to be associated with its isoform-specific molecular distribution and regulation of a set of cancer-related transcripts, in esophageal squamous cell carcinoma (ESCC). Immunohistochemical overexpression of TIA1 ectopically localized in the cytoplasm of tumor cells was an independent prognosticator for worse overall survival in a cohort of 143 ESCC patients. Knockdown of TIA1 inhibited proliferation of ESCC cells. By exogenously introducing each of two major isoforms, TIA1a and TIA1b, only TIA1a, which was localized to both the nucleus and cytoplasm, promoted anchorage-dependent and anchorage-independent ESCC cell proliferation. Ribonucleoprotein immunoprecipitation, followed by microarray analysis or massive-parallel sequencing, identified a set of TIA1-binding mRNAs, including SKP2 and CCNA2. TIA1 increased SKP2 and CCNA2 protein levels through the suppression of mRNA decay and translational induction, respectively. Our findings uncover a novel oncogenic function of TIA1 in esophageal tumorigenesis, and implicate its use as a marker for prognostic evaluation and as a therapeutic target in ESCC. PMID:26958940

  4. β-Catenin Upregulates the Constitutive and Virus-Induced Transcriptional Capacity of the Interferon Beta Promoter through T-Cell Factor Binding Sites

    PubMed Central

    Marcato, Vasco; Luron, Lionel; Laqueuvre, Lucie M.; Simon, Dominique; Mansuroglu, Zeyni; Flamand, Marie; Panthier, Jean-Jacques; Souès, Sylvie; Massaad, Charbel

    2015-01-01

    Rapid upregulation of interferon beta (IFN-β) expression following virus infection is essential to set up an efficient innate antiviral response. Biological roles related to the antiviral and immune response have also been associated with the constitutive production of IFN-β in naive cells. However, the mechanisms capable of modulating constitutive IFN-β expression in the absence of infection remain largely unknown. In this work, we demonstrate that inhibition of the kinase glycogen synthase kinase 3 (GSK-3) leads to the upregulation of the constitutive level of IFN-β expression in noninfected cells, provided that GSK-3 inhibition is correlated with the binding of β-catenin to the IFN-β promoter. Under these conditions, IFN-β expression occurred through the T-cell factor (TCF) binding sites present on the IFN-β promoter independently of interferon regulatory factor 3 (IRF3). Enhancement of the constitutive level of IFN-β per se was able to confer an efficient antiviral state to naive cells and acted in synergy with virus infection to stimulate virus-induced IFN-β expression. Further emphasizing the role of β-catenin in the innate antiviral response, we show here that highly pathogenic Rift Valley fever virus (RVFV) targets the Wnt/β-catenin pathway and the formation of active TCF/β-catenin complexes at the transcriptional and protein level in RVFV-infected cells and mice. PMID:26459757

  5. PI3Kδ promotes CD4+ T-cell interactions with antigen-presenting cells by increasing LFA-1 binding to ICAM-1

    PubMed Central

    Garçon, Fabien; Okkenhaug, Klaus

    2016-01-01

    Activation of T lymphocytes by peptide/major histocompatibility complex on antigen-presenting cells (APCs) involves dynamic contacts between the two cells, during which T cells undergo marked morphological changes. These interactions are facilitated by integrins. Activation of the T cells increases the binding of the integrin lymphocyte function-associated antigen 1 (LFA-1) expressed by T cells to intercellular adhesion molecule (ICAM)-1 and ICAM-2 expressed by APCs. The signalling pathways that control integrin affinities are incompletely defined. The phosphoinositide 3-kinases (PI3Ks) generate second-messenger signalling molecules that control cell growth, proliferation, differentiation and trafficking. Here we show that in T cells, PI3Kδ attenuates the activation of Rac1, but sustains the activation of Rap1. Consequently, PI3Kδ increases LFA-1-dependent adhesion to form stable conjugates with APCs. Increased Rap1 activity and LFA-1 adhesion were only in part mediated by the downstream kinase Akt, suggesting the involvement of additional phosphatidylinositol(3,4,5)P3-binding proteins. These results establish a link between PI3K activity, cytoskeletal changes and integrin binding and help explain the impaired T-cell-dependent immune responses in PI3Kδ-deficient mice. PMID:26740009

  6. PI3Kδ promotes CD4(+) T-cell interactions with antigen-presenting cells by increasing LFA-1 binding to ICAM-1.

    PubMed

    Garçon, Fabien; Okkenhaug, Klaus

    2016-05-01

    Activation of T lymphocytes by peptide/major histocompatibility complex on antigen-presenting cells (APCs) involves dynamic contacts between the two cells, during which T cells undergo marked morphological changes. These interactions are facilitated by integrins. Activation of the T cells increases the binding of the integrin lymphocyte function-associated antigen 1 (LFA-1) expressed by T cells to intercellular adhesion molecule (ICAM)-1 and ICAM-2 expressed by APCs. The signalling pathways that control integrin affinities are incompletely defined. The phosphoinositide 3-kinases (PI3Ks) generate second-messenger signalling molecules that control cell growth, proliferation, differentiation and trafficking. Here we show that in T cells, PI3Kδ attenuates the activation of Rac1, but sustains the activation of Rap1. Consequently, PI3Kδ increases LFA-1-dependent adhesion to form stable conjugates with APCs. Increased Rap1 activity and LFA-1 adhesion were only in part mediated by the downstream kinase Akt, suggesting the involvement of additional phosphatidylinositol(3,4,5)P3-binding proteins. These results establish a link between PI3K activity, cytoskeletal changes and integrin binding and help explain the impaired T-cell-dependent immune responses in PI3Kδ-deficient mice. PMID:26740009

  7. Nck Binds to the T Cell Antigen Receptor Using Its SH3.1 and SH2 Domains in a Cooperative Manner, Promoting TCR Functioning.

    PubMed

    Paensuwan, Pussadee; Hartl, Frederike A; Yousefi, O Sascha; Ngoenkam, Jatuporn; Wipa, Piyamaporn; Beck-Garcia, Esmeralda; Dopfer, Elaine P; Khamsri, Boonruang; Sanguansermsri, Donruedee; Minguet, Susana; Schamel, Wolfgang W; Pongcharoen, Sutatip

    2016-01-01

    Ligand binding to the TCR causes a conformational change at the CD3 subunits to expose the CD3ε cytoplasmic proline-rich sequence (PRS). It was suggested that the PRS is important for TCR signaling and T cell activation. It has been shown that the purified, recombinant SH3.1 domain of the adaptor molecule noncatalytic region of tyrosine kinase (Nck) can bind to the exposed PRS of CD3ε, but the molecular mechanism of how full-length Nck binds to the TCR in cells has not been investigated so far. Using the in situ proximity ligation assay and copurifications, we show that the binding of Nck to the TCR requires partial phosphorylation of CD3ε, as it is based on two cooperating interactions. First, the SH3.1(Nck) domain has to bind to the nonphosphorylated and exposed PRS, that is, the first ITAM tyrosine has to be in the unphosphorylated state. Second, the SH2(Nck) domain has to bind to the second ITAM tyrosine in the phosphorylated state. Likewise, mutations of the SH3.1 and SH2 domains in Nck1 resulted in the loss of Nck1 binding to the TCR. Furthermore, expression of an SH3.1-mutated Nck impaired TCR signaling and T cell activation. Our data suggest that the exact pattern of CD3ε phosphorylation is critical for TCR functioning. PMID:26590318

  8. The transcription factor NFAT promotes exhaustion of activated CD8⁺ T cells.

    PubMed

    Martinez, Gustavo J; Pereira, Renata M; Äijö, Tarmo; Kim, Edward Y; Marangoni, Francesco; Pipkin, Matthew E; Togher, Susan; Heissmeyer, Vigo; Zhang, Yi Chen; Crotty, Shane; Lamperti, Edward D; Ansel, K Mark; Mempel, Thorsten R; Lähdesmäki, Harri; Hogan, Patrick G; Rao, Anjana

    2015-02-17

    During persistent antigen stimulation, CD8(+) T cells show a gradual decrease in effector function, referred to as exhaustion, which impairs responses in the setting of tumors and infections. Here we demonstrate that the transcription factor NFAT controls the program of T cell exhaustion. When expressed in cells, an engineered form of NFAT1 unable to interact with AP-1 transcription factors diminished T cell receptor (TCR) signaling, increased the expression of inhibitory cell surface receptors, and interfered with the ability of CD8(+) T cells to protect against Listeria infection and attenuate tumor growth in vivo. We defined the genomic regions occupied by endogenous and engineered NFAT1 in primary CD8(+) T cells and showed that genes directly induced by the engineered NFAT1 overlapped with genes expressed in exhausted CD8(+) T cells in vivo. Our data show that NFAT promotes T cell anergy and exhaustion by binding at sites that do not require cooperation with AP-1. PMID:25680272

  9. Ethanol promotes T cell apoptosis through the mitochondrial pathway

    PubMed Central

    Kapasi, Aditi A; Patel, Geeta; Goenka, Anuj; Nahar, Nilay; Modi, Neeraj; Bhaskaran, Madhu; Reddy, Krishna; Franki, Nicholas; Patel, Jaimita; Singhal, Pravin C

    2003-01-01

    Clinical reports suggest that acute ethanol intoxication is often associated with lymphopenia. Previously, ethanol was reported to invoke thymocyte apoptosis. We studied the effect of ethanol on T cell apoptosis. In addition, we evaluated the molecular mechanism of ethanol-induced T cell apoptosis. Human T cells harvested from healthy subjects after an alcohol drinking binge showed enhanced T cell apoptosis (before, 0·4 ± 0·2% versus after, 19·6 ± 2·5% apoptotic lymphocytes/field; P < 0·001). In in vitro studies, ethanol in a concentration of 50 mm and higher enhanced the apoptosis of Jurkat cells. DNA isolated from ethanol-treated Jurkat cells displayed integer multiples of 180 base pairs. Ethanol decreased Jurkat cell expression of Bcl-2, whereas ethanol increased Jurkat cell expression of Bax. Jurkat cells treated with ethanol also showed translocation of cytochrome C into cytosol. Moreover, a caspase-9 inhibitor partially inhibited ethanol-induced Jurkat cell apoptosis. In in vivo studies, after binge drinking, T cell expression of Bcl-2 also decreased. In addition, binge drinking induced the cleavage of caspase-3, suggesting activation of caspase-3 in T cells. These results suggest that ethanol promotes T cell apoptosis through the activation of intrinsic or mitochondrial pathway. PMID:12603597

  10. Dengue virus specific dual HLA binding T cell epitopes induce CD8+ T cell responses in seropositive individuals.

    PubMed

    Comber, Joseph D; Karabudak, Aykan; Huang, Xiaofang; Piazza, Paolo A; Marques, Ernesto T A; Philip, Ramila

    2014-01-01

    Dengue virus infects an estimated 300 million people each year and even more are at risk of becoming infected as the virus continues to spread into new areas. Despite the increase in viral prevalence, no anti-viral medications or vaccines are approved for treating or preventing infection. CD8+ T cell responses play a major role in viral clearance. Therefore, effective vaccines that induce a broad, multi-functional T cell response with substantial cross-reactivity between all virus serotypes can have major impacts on reducing infection rates and infection related complications. Here, we took an immunoproteomic approach to identify novel MHC class I restricted T cell epitopes presented by dengue virus infected cells, representing the natural and authentic targets of the T cell response. Using this approach we identified 4 novel MHC-I restricted epitopes: 2 with the binding motif for HLA-A24 molecules and 2 with both HLA-A2 and HLA-A24 binding motifs. These peptides were able to activate CD8+ T cell responses in both healthy, seronegative individuals and in seropositive individuals who have previously been infected with dengue virus. Importantly, the dual binding epitopes activated pre-existing T cell precursors in PBMCs obtained from both HLA-A2+ and HLA-A24+ seropositive individuals. Together, the data indicate that these epitopes are immunologically relevant T cell activating peptides presented on infected cells during a natural infection and therefore may serve as candidate antigens for the development of effective multi-serotype specific dengue virus vaccines. PMID:25668665

  11. Dengue virus specific dual HLA binding T cell epitopes induce CD8+ T cell responses in seropositive individuals

    PubMed Central

    Comber, Joseph D; Karabudak, Aykan; Huang, Xiaofang; Piazza, Paolo A; Marques, Ernesto T A; Philip, Ramila

    2015-01-01

    Dengue virus infects an estimated 300 million people each year and even more are at risk of becoming infected as the virus continues to spread into new areas. Despite the increase in viral prevalence, no anti-viral medications or vaccines are approved for treating or preventing infection. CD8+ T cell responses play a major role in viral clearance. Therefore, effective vaccines that induce a broad, multi-functional T cell response with substantial cross-reactivity between all virus serotypes can have major impacts on reducing infection rates and infection related complications. Here, we took an immunoproteomic approach to identify novel MHC class I restricted T cell epitopes presented by dengue virus infected cells, representing the natural and authentic targets of the T cell response. Using this approach we identified 4 novel MHC-I restricted epitopes: 2 with the binding motif for HLA-A24 molecules and 2 with both HLA-A2 and HLA-A24 binding motifs. These peptides were able to activate CD8+ T cell responses in both healthy, seronegative individuals and in seropositive individuals who have previously been infected with dengue virus. Importantly, the dual binding epitopes activated pre-existing T cell precursors in PBMCs obtained from both HLA-A2+ and HLA-A24+ seropositive individuals. Together, the data indicate that these epitopes are immunologically relevant T cell activating peptides presented on infected cells during a natural infection and therefore may serve as candidate antigens for the development of effective multi-serotype specific dengue virus vaccines. PMID:25668665

  12. The adenovirus e3 promoter is sensitive to activation signals in human T cells.

    PubMed

    Mahr, Jeffrey A; Boss, Jeremy M; Gooding, Linda R

    2003-01-01

    The group C adenoviruses typically cause acute respiratory disease in young children. In addition, a persistent phase of infection has been observed in which virus may be shed for years without producing overt pathology. Our laboratory recently reported that group C adenovirus DNA can be found in tonsil and adenoid T lymphocytes from the majority of pediatric donors (C. T. Garnett, D. Erdman, W. Xu, and L. R. Gooding, J. Virol. 76:10608-10616, 2002). This finding suggests that immune evasion strategies of human adenoviruses may be directed, in part, toward protection of persistently or latently infected T lymphocytes. Many of the adenoviral gene products implicated in prevention of immune destruction of virus-infected cells are encoded within the E3 transcription unit. In this study, the E3 promoter was evaluated for sensitivity to T-cell activation signals by using a promoter reporter plasmid. Indeed, this promoter is extremely sensitive to T-cell activation, with phorbol myristate acetate (PMA) plus ionomycin increasing E3-directed transcription 100-fold. By comparison, in the same cells E1A expression leads to a 5.5-fold increase in transcription from the E3 promoter. In contrast to induction by E1A, activation by PMA plus ionomycin requires the two E3 NF-kappaB binding sites. Interestingly, expression of E1A inhibits induction of the E3 promoter in response to T-cell activation while increasing E3 promoter activity in unactivated cells. Collectively, these data suggest that the E3 promoter may have evolved the capacity to respond to T-cell activation in the absence of E1A expression and may act to upregulate antiapoptotic gene expression in order to promote survival of persistently infected T lymphocytes. PMID:12502827

  13. The DNA-binding inhibitor Id3 regulates IL-9 production in CD4(+) T cells.

    PubMed

    Nakatsukasa, Hiroko; Zhang, Dunfang; Maruyama, Takashi; Chen, Hua; Cui, Kairong; Ishikawa, Masaki; Deng, Lisa; Zanvit, Peter; Tu, Eric; Jin, Wenwen; Abbatiello, Brittany; Goldberg, Nathan; Chen, Qianming; Sun, Lingyun; Zhao, Keji; Chen, WanJun

    2015-10-01

    The molecular mechanisms by which signaling via transforming growth factor-β (TGF-β) and interleukin 4 (IL-4) control the differentiation of CD4(+) IL-9-producing helper T cells (TH9 cells) remain incompletely understood. We found here that the DNA-binding inhibitor Id3 regulated TH9 differentiation, as deletion of Id3 increased IL-9 production from CD4(+) T cells. Mechanistically, TGF-β1 and IL-4 downregulated Id3 expression, and this process required the kinase TAK1. A reduction in Id3 expression enhanced binding of the transcription factors E2A and GATA-3 to the Il9 promoter region, which promoted Il9 transcription. Notably, Id3-mediated control of TH9 differentiation regulated anti-tumor immunity in an experimental melanoma-bearing model in vivo and also in human CD4(+) T cells in vitro. Thus, our study reveals a previously unrecognized TAK1-Id3-E2A-GATA-3 pathway that regulates TH9 differentiation. PMID:26322481

  14. Promoting transplantation tolerance; adoptive regulatory T cell therapy

    PubMed Central

    Safinia, N; Leech, J; Hernandez-Fuentes, M; Lechler, R; Lombardi, G

    2013-01-01

    Transplantation is a successful treatment for end-stage organ failure. Despite improvements in short-term outcome, long-term survival remains suboptimal because of the morbidity and mortality associated with long-term use of immunosuppression. There is, therefore, a pressing need to devise protocols that induce tolerance in order to minimize or completely withdraw immunosuppression in transplant recipients. In this review we will discuss how regulatory T cells (Tregs) came to be recognized as an attractive way to promote transplantation tolerance. We will summarize the preclinical data, supporting the importance of these cells in the induction and maintenance of immune tolerance and that provide the rationale for the isolation and expansion of these cells for cellular therapy. We will also describe the data from the first clinical trials, using Tregs to inhibit graft-versus-host disease (GVHD) after haematopoietic stem cell transplantation and will address both the challenges and opportunities in human Treg cell therapy. PMID:23574313

  15. EMSA Eritin Drives Expansion of Regulatory T Cells and Promotes T Cells Differentiation in Irradiated Mice.

    PubMed

    Ibrahim, Mansur; Widjajanto, Edi; Widodo, M Aris; Sumitro, Sutiman B

    2016-07-01

    Sublethal irradiation therapy in cancer treatment causes generalized immunosuppression, which results in a range of DNA damage. We examined the significance of a polyherbal medicine called "EMSA Eritin" on immunological responses in sublethally irradiated mice focusing on the involvement of Treg, naïve T cell, and also the development and differentiation of T cells in thymus. Normal BALB/c mice were sublethally irradiated with dose of 600 rad. The irradiated mice were then orally administered by EMSA Eritin once a day at different doses: 1.04, 3.12, 9.37 mg/g body weight. The treatment was performed for 14 days. On day 15, immunological responses were observed by analyzing the status of Treg and differentiation of T cells in thymus. The administration of EMSA Eritin to irradiated mice resulted in a significant increase of pre T cells, Treg cells, and naïve T cells, which in general could maintain and normalize healthy condition in mice. PMID:26170134

  16. Nuclear matrix binding protein SMAR1 regulates T-cell differentiation and allergic airway disease.

    PubMed

    Chemmannur, S V; Badhwar, A J; Mirlekar, B; Malonia, S K; Gupta, M; Wadhwa, N; Bopanna, R; Mabalirajan, U; Majumdar, S; Ghosh, B; Chattopadhyay, S

    2015-11-01

    Asthma is a complex airway allergic disease involving the interplay of various cell types, cytokines, and transcriptional factors. Though many factors contribute to disease etiology, the molecular control of disease phenotype and responsiveness is not well understood. Here we report an essential role of the matrix attachment region (MAR)-binding protein SMAR1 in regulating immune response during allergic airway disease. Conditional knockout of SMAR1 in T cells rendered the mice resistant to eosinophilic airway inflammation against ovalbumin (OVA) allergen with low immunoglobulin E (IgE) and interleukin-5 (IL-5) levels. Moreover, a lower IgE/IgG2a ratio and higher interferon-γ (IFN-γ) response suggested aberrant skewing of T-cell differentiation toward type 1 helper T cell (Th1) response. We show that SMAR1 functions as a negative regulator of Th1 and Th17 differentiation by interacting with two potential and similar MAR regions present on the promoters of T-bet and IL-17. Thus, we present SMAR1 as a regulator of T-cell differentiation that favors the establishment of Th2 cells by modulating Th1 and Th17 responses. PMID:25736456

  17. End-binding protein 1 controls signal propagation from the T cell receptor

    PubMed Central

    Martín-Cófreces, Noa B; Baixauli, Francesc; López, María J; Gil, Diana; Monjas, Alicia; Alarcón, Balbino; Sánchez-Madrid, Francisco

    2012-01-01

    The role of microtubules (MTs) in the control and dynamics of the immune synapse (IS) remains unresolved. Here, we show that T cell activation requires the growth of MTs mediated by the plus-end specific protein end-binding 1 (EB1). A direct interaction of the T cell receptor (TCR) complex with EB1 provides the molecular basis for EB1 activity promoting TCR encounter with signalling vesicles at the IS. EB1 knockdown alters TCR dynamics at the IS and prevents propagation of the TCR activation signal to LAT, thus inhibiting activation of PLCγ1 and its localization to the IS. These results identify a role for EB1 interaction with the TCR in controlling TCR sorting and its connection with the LAT/PLCγ1 signalosome. PMID:22922463

  18. Binding of WIP to Actin Is Essential for T Cell Actin Cytoskeleton Integrity and Tissue Homing

    PubMed Central

    Massaad, Michel J.; Oyoshi, Michiko K.; Kane, Jennifer; Koduru, Suresh; Alcaide, Pilar; Nakamura, Fumihiko; Ramesh, Narayanaswamy; Luscinskas, Francis W.; Hartwig, John

    2014-01-01

    The Wiskott-Aldrich syndrome protein (WASp) is important for actin polymerization in T cells and for their migration. WASp-interacting protein (WIP) binds to and stabilizes WASp and also interacts with actin. Cytoskeletal and functional defects are more severe in WIP−/− T cells, which lack WASp, than in WASp−/− T cells, suggesting that WIP interaction with actin may be important for T cell cytoskeletal integrity and function. We constructed mice that lack the actin-binding domain of WIP (WIPΔABD mice). WIPΔABD associated normally with WASp but not F-actin. T cells from WIPΔABD mice had normal WASp levels but decreased cellular F-actin content, a disorganized actin cytoskeleton, impaired chemotaxis, and defective homing to lymph nodes. WIPΔABD mice exhibited a T cell intrinsic defect in contact hypersensitivity and impaired responses to cutaneous challenge with protein antigen. Adoptively transferred antigen-specific CD4+ T cells from WIPΔABD mice had decreased homing to antigen-challenged skin of wild-type recipients. These findings show that WIP binding to actin, independently of its binding to WASp, is critical for the integrity of the actin cytoskeleton in T cells and for their migration into tissues. Disruption of WIP binding to actin could be of therapeutic value in T cell-driven inflammatory diseases. PMID:25246631

  19. Spatial congregation of STAT binding directs selective nuclear architecture during T-cell functional differentiation

    PubMed Central

    Hakim, Ofir; Sung, Myong-Hee; Nakayamada, Shingo; Voss, Ty C.; Baek, Songjoon; Hager, Gordon L.

    2013-01-01

    Higher-order genome organization shows tissue-specific patterns. However, functional relevance and the mechanisms shaping the genome architecture are poorly understood. Here we report a profound shift from promiscuous to highly selective genome organization that accompanies the effector lineage choice of differentiating T cells. As multipotent naive cells receive antigenic signals and commit to a T helper (Th) pathway, the genome-wide contacts of a lineage-specific cytokine locus are preferentially enriched for functionally relevant genes. Despite the establishment of divergent interactomes and global reprogramming of transcription in Th1 versus Th2, the overall expression status of the contact genes is surprisingly similar between the two lineages. Importantly, during differentiation, the genomic contacts are retained and strengthened precisely at DNA binding sites of the specific lineage-determining STAT transcription factor. In cells from the specific STAT knock-out mouse, the signature cytokine locus is unable to shed the promiscuous contacts established in the naive T cells, indicating the importance of genomic STAT binding. Altogether, the global aggregation of STAT binding loci from genic and nongenic regions highlights a new role for differentiation-promoting transcription factors in direct specification of higher-order nuclear architecture through interacting with regulatory regions. Such subnuclear environments have significant implications for efficient functioning of the mature effector lymphocytes. PMID:23212947

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

    PubMed

    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

  1. Promoting metastasis: neutrophils and T cells join forces.

    PubMed

    Fridlender, Zvi G; Albelda, Steven M; Granot, Zvi

    2015-07-01

    The role neutrophils play in cancer is a matter of debate as both pro- and anti-tumor functions have been documented. In a recent publication in Nature, Coffelt et al. identify a new mechanism where neutrophils and T cells cooperate to generate metastasis-supporting immune suppression. PMID:26138787

  2. Unusual features of Self-Peptide/MHC Binding by Autoimmune T Cell Receptors

    SciTech Connect

    Nicholson,M.; Hahn, M.; Wucherpfennig, K.

    2005-01-01

    Structural studies on T cell receptors (TCRs) specific for foreign antigens demonstrated a remarkably similar topology characterized by a central, diagonal TCR binding mode that maximizes interactions with the MHC bound peptide. However, three recent structures involving autoimmune TCRs demonstrated unusual interactions with self-peptide/MHC complexes. Two TCRs from multiple sclerosis patients bind with unconventional topologies, and both TCRs are shifted toward the peptide N terminus and the MHC class II {beta} chain helix. A TCR from the experimental autoimmune encephalomyelitis (EAE) model binds in a conventional orientation, but the structure is unusual because the self-peptide only partially fills the binding site. For all three TCRs, interaction with the MHC bound self-peptide is suboptimal, and only two or three TCR loops contact the peptide. Optimal TCR binding modes confer a competitive advantage for antimicrobial T cells during an infection, whereas altered binding properties may permit survival of a subset of autoreactive T cells during thymic selection.

  3. Predictions of T-cell receptor- and major histocompatibility complex-binding sites on staphylococcal enterotoxin C1.

    PubMed Central

    Hoffmann, M L; Jablonski, L M; Crum, K K; Hackett, S P; Chi, Y I; Stauffacher, C V; Stevens, D L; Bohach, G A

    1994-01-01

    We have focused on regions of staphylococcal enterotoxin C1 (SEC1) causing immunomodulation. N-terminal deletion mutants lacking residues 6 through 13 induced T-cell proliferation similar to that induced by native toxin. However, mutants with residues deleted between positions 19 and 33, although nonmitogenic themselves, were able to inhibit both SEC1-induced T-cell proliferation and binding of the native toxin to major histocompatibility complex (MHC) class II. Presumably, these deletions define a part of SEC1 that interacts with the T-cell receptor. Three synthetic peptides containing residues located in a region analogous to the alpha 5 groove of SEC3 had residual mitogenic activity or blocked T-cell proliferation induced by SEC1 and appear to recognize the same site as SEC1 on a receptor for the toxin, presumably MHC class II. We conclude that isolated portions of the SEC1 molecule can retain residual mitogenic activity but that the entire protein is needed to achieve maximal superantigenic stimulation. Our results, together with the results of other investigators, support a model in which SEC1 binds to an alpha helix of MHC class II through a central groove in the toxin and thereby promotes or stabilizes the interaction between antigen-presenting cells and T cells. Images PMID:8039910

  4. Immunopathology of experimental Chagas' disease: binding of T cells to Trypanosoma cruzi-infected heart tissue.

    PubMed Central

    Mortatti, R C; Maia, L C; de Oliveira, A V; Munk, M E

    1990-01-01

    The immunopathology of Chagas' disease was studied in the experimental model of chronic infection in C57BL/10JT or mice. Sublethal infection with Trypanosoma cruzi, Y strain, induced specific antibodies and a delayed hypersensitivity response to parasite antigens. Mice developed chronic chagasic myocarditis but not skeletal muscle myositis. Binding of T cells to infected heart tissue was investigated during short-term cocultivation of lymphocytes with heart cryostat sections. T cells from infected mice and from normal controls bound equally to myocardium and liver sections from both infected and normal mice. A search in depth was attempted with cells heavily tagged with 99mTc. Labeled T cells from chagasic mice bound to both normal and infected myocardium slices. 99mTc-labeled T cells from controls gave the same binding values. Glass-adherent spleen cells behaved identically to T cells. Prior treatment of the tissue with serum from chronically infected mice did not increase the number of binding cells. Peritoneal macrophages tagged with 99mTc-sulfur colloid also bound to infected myocardium slices. The binding of macrophages was not changed by pretreatment of infected tissue with anti-T, cruzi antibodies. In short, this work did not detect any population of T cells or macrophages which could bind specifically to infected heart tissue to initiate an autoreactive process. Images PMID:2228230

  5. MeCP2 enforces Foxp3 expression to promote regulatory T cells' resilience to inflammation.

    PubMed

    Li, Chaoran; Jiang, Shan; Liu, Si-Qi; Lykken, Erik; Zhao, Lin-tao; Sevilla, Jose; Zhu, Bo; Li, Qi-Jing

    2014-07-01

    Forkhead box P3(+) (Foxp3(+)) regulatory T cells (Tregs) are crucial for peripheral tolerance. During inflammation, steady Foxp3 expression in Tregs is essential for maintaining their lineage identity and suppressive function. However, the molecular machinery governing Tregs' resilience to inflammation-induced Foxp3 destabilization remains elusive. Here, we demonstrate that methyl-CpG binding protein 2 (MeCP2), an eminent epigenetic regulator known primarily as the etiological factor of Rett syndrome, is critical to sustain Foxp3 expression in Tregs during inflammation. In response to inflammatory stimuli, MeCP2 is specifically recruited to the Conserved Non-Coding sequence 2 region of the foxp3 locus, where it collaborates with cAMP responsive element binding protein 1 to promote local histone H3 acetylation, thereby counteracting inflammation-induced epigenetic silencing of foxp3. Consequently, Treg-specific deletion of MeCP2 causes spontaneous immune activation in mice and failure in protection against autoimmunity. Furthermore, we demonstrate that Foxp3 expression in MeCP2-deficient Tregs diminishes with time, resulting in their failure to suppress effector T-cell-mediated colitis. Thus, MeCP2 serves as a critical safeguard that confers Tregs with resilience against inflammation. PMID:24958888

  6. Differential interaction of nuclear factors with the leukocyte-specific pp52 promoter in B and T cells.

    PubMed

    Omori, S A; Smale, S; O'Shea-Greenfield, A; Wall, R

    1997-08-15

    The leukocyte-specific, cytoskeleton-binding pp52 (LSP-1, WP-34) protein is widely expressed in multiple leukocyte lineages, including B and T lymphocytes, granulocytes, and macrophages. We previously detected a tissue-specific promoter preceding the exon encoding the N terminus of the pp52 leukocyte protein. Here we describe the functional characterization of this promoter and identification of the factors in B and T cells that regulate its activity. The pp52 promoter contains an initiator specifying the unique 5' terminus of pp52 mRNA, tandem pairs of Ets and SP1 motifs, and a lone C/EBP motif. All these motifs are essential and collectively control transcriptional activity. DNA binding studies and Ab supershift assays revealed that different combinations of factors interact with these motifs in B cells vs T cells. The Ets motifs are preferentially bound by PU-1 in B cell extracts from all stages of development, whereas a different Ets family member reacts with these motifs in T cell extracts. The C/EBP motif is bound by Ig/EBP-1 in pre-B cell and T cell extracts, but is replaced by nuclear factor-IL-6beta or a nuclear factor-IL-6beta-Ig/EBP-1 heterodimer in plasmacytoma cell extracts. Despite its reported role as a negative regulator of transcription, Ig/EBP-1 appears to exert a stimulatory effect on this promoter. These findings reveal the features controlling the pp52 promoter in B and T cells and provide the foundation for determining the regulation of this promoter in other leukocyte lineages. PMID:9257843

  7. Functional analysis of the murine T-cell receptor beta enhancer and characteristics of its DNA-binding proteins.

    PubMed Central

    Takeda, J; Cheng, A; Mauxion, F; Nelson, C A; Newberry, R D; Sha, W C; Sen, R; Loh, D Y

    1990-01-01

    The minimal T-cell receptor (TCR) beta-chain (TCR beta) enhancer has been identified by transfection into lymphoid cells. The minimal enhancer was active in T cells and in some B-lineage cells. When a larger fragment containing the minimal enhancer was used, its activity was apparent only in T cells. Studies with phytohemagglutinin and 4 beta-phorbol-12,13-dibutyrate revealed that the enhancer activity was increased by these agents. By a combination of DNase I footprinting, gel mobility shift assay, and methylation interference analysis, seven different motifs were identified within the minimal enhancer. Furthermore, competition experiments showed that some of these elements bound identical or similar factors that are known to bind to the TCR V beta promoter decamer or to the immunoglobulin enhancer kappa E2 or muEBP-E motif. These shared motifs may be important in the differential gene activity among the different lymphoid subsets. Images PMID:2144608

  8. A Unique DNA Binding Domain Converts T-Cell Factors into Strong Wnt Effectors▿

    PubMed Central

    Atcha, Fawzia A.; Syed, Adeela; Wu, Beibei; Hoverter, Nate P.; Yokoyama, Noriko N.; Ting, Ju-Hui T.; Munguia, Jesus E.; Mangalam, Harry J.; Marsh, J. Lawrence; Waterman, Marian L.

    2007-01-01

    Wnt regulation of gene expression requires binding of LEF/T-cell factor (LEF/TCF) transcription factors to Wnt response elements (WREs) and recruitment of the activator β-catenin. There are significant differences in the abilities of LEF/TCF family members to regulate Wnt target genes. For example, alternatively spliced isoforms of TCF-1 and TCF-4 with a C-terminal “E” tail are uniquely potent in their activation of LEF1 and CDX1. Here we report that the mechanism responsible for this unique activity is an auxiliary 30-amino-acid DNA interaction motif referred to here as the “cysteine clamp” (or C-clamp). The C-clamp contains invariant cysteine, aromatic, and basic residues, and surface plasmon resonance (SPR) studies with recombinant C-clamp protein showed that it binds double-stranded DNA but not single-stranded DNA or RNA (equilibrium dissociation constant = 16 nM). CASTing (Cyclic Amplification and Selection of Targets) experiments were used to test whether this motif influences WRE recognition. Full-length LEF-1, TCF-1E, and TCF-1E with a mutated C-clamp all bind nearly identical WREs (TYYCTTTGATSTT), showing that the C-clamp does not alter WRE specificity. However, a GC element downstream of the WRE (RCCG) is enriched in wild-type TCF-1E binding sites but not in mutant TCF-1E binding sites. We conclude that the C-clamp is a sequence-specific DNA binding motif. C-clamp mutations destroy the ability of β-catenin to regulate the LEF1 promoter, and they severely impair the ability of TCF-1 to regulate growth in colon cancer cells. Thus, E-tail isoforms of TCFs utilize two DNA binding activities to access a subset of Wnt targets important for cell growth. PMID:17893322

  9. Itk Signals Promote Neuroinflammation by Regulating CD4+ T-Cell Activation and Trafficking

    PubMed Central

    Kannan, Arun K.; Kim, Do-Geun

    2015-01-01

    Here we demonstrate that interleukin-2-inducible T-cell kinase (Itk) signaling in cluster of differentiation 4-positive (CD4+) T cells promotes experimental autoimmune encephalomyelitis (EAE), the animal model of multiple sclerosis (MS). We show that Itk−/− mice exhibit reduced disease severity, and transfer of Itk−/− CD4+ T cells into T cell-deficient recipients results in lower disease severity. We observed a significant reduction of CD4+ T cells in the CNS of Itk−/− mice or recipients of Itk−/− CD4+ T cells during EAE, which is consistent with attenuated disease. Itk−/− CD4+ T cells exhibit defective response to myelin antigen stimulation attributable to displacement of filamentous actin from the CD4+ coreceptor. This results in inadequate transmigration of Itk−/− CD4+ T cells into the CNS and across brain endothelial barriers in vitro. Finally, Itk−/− CD4+ T cells show significant reduction in production of T-helper 1 (Th1) and Th17 cytokines and exhibit skewed T effector/T regulatory cell ratios. These results indicate that signaling by Itk promotes autoimmunity and CNS inflammation, suggesting that it may be a viable target for treatment of MS. PMID:25568116

  10. Mechanisms of SHP-1 P2 promoter regulation in hematopoietic cells and its silencing in HTLV-1-transformed T cells.

    PubMed

    Nakase, Koichi; Cheng, Jihua; Zhu, Quan; Marasco, Wayne A

    2009-01-01

    The Src homology-2-containing protein-tyrosine phosphatase 1 (SHP-1), is a negative regulator of cell signaling. It is also considered a tumor suppressor gene because of its ability to antagonize the action of tyrosine kinases. Although SHP-1 is expressed strongly in hematopoietic cells, decreased expression has been observed in various hematological malignancies, which suggests a central involvement of SHP-1 in leukemogenesis. We have shown previously that human T cell lymphotropic virus type-1 (HTLV-1) Tax-induced promoter silencing (TIPS) is an early event causing down-regulation of SHP-1 expression, which is dependent on NF-kappaB. In this study, DNase I footprinting and EMSA also revealed binding of transcription factors, specificity protein 1 (Sp1) and octamer-binding transcription factor 1 (Oct-1) to the P2 promoter, and site-directed mutagenesis confirmed that these factors contribute to the basal P2 promoter activity. Chromatin immunoprecipitation (CHIP) assays showed that Sp1, Oct-1, NF-kappaB, CREB-1, and RNA polymerase II interacted with the core SHP-1 P2 promoter in CD4+ T cells and Jurkat cells but not in HTLV-1-transformed MT-2 and HUT102 cells when HTLV-1 Tax is present. Furthermore, bisulfite sequencing of the SHP-1 P2 core region revealed heavy CpG methylation in HTLV-1-transformed cells compared with freshly isolated CD4+ T cells and HTLV-1-noninfected T cell lines. A significant inverse correlation between degree of CpG methylation and expression of SHP-1 mRNA or protein was observed. Taken together, our data support the notion that in HTLV-1-transformed CD4+ T cells, TIPS causes dissociation of transcription factors from the core SHP-1 P2 promoter, which in turn leads to subsequent DNA methylation, an important early step for leukemogenesis. PMID:18948549

  11. R-Ras Regulates Murine T Cell Migration and Intercellular Adhesion Molecule-1 Binding.

    PubMed

    Yan, Xiaocai; Yan, Mingfei; Guo, Yihe; Singh, Gobind; Chen, Yuhong; Yu, Mei; Wang, Demin; Hillery, Cheryl A; Chan, Andrew M

    2015-01-01

    The trafficking of T-lymphocytes to peripheral draining lymph nodes is crucial for mounting an adaptive immune response. The role of chemokines in the activation of integrins via Ras-related small GTPases has been well established. R-Ras is a member of the Ras-subfamily of small guanosine-5'-triphosphate-binding proteins and its role in T cell trafficking has been investigated in R-Ras null mice (Rras-/-). An examination of the lymphoid organs of Rras-/- mice revealed a 40% reduction in the cellularity of the peripheral lymph nodes. Morphologically, the high endothelial venules of Rras-/- mice were more disorganized and less mature than those of wild-type mice. Furthermore, CD4+ and CD8+ T cells from Rras-/- mice had approximately 42% lower surface expression of L-selectin/CD62L. These aberrant peripheral lymph node phenotypes were associated with proliferative and trafficking defects in Rras-/- T cells. Furthermore, R-Ras could be activated by the chemokine, CCL21. Indeed, Rras-/- T cells had approximately 14.5% attenuation in binding to intercellular adhesion molecule 1 upon CCL21 stimulation. Finally, in a graft-versus host disease model, recipient mice that were transfused with Rras-/- T cells showed a significant reduction in disease severity when compared with mice transplanted with wild-type T cells. These findings implicate a role for R-Ras in T cell trafficking in the high endothelial venules during an effective immune response. PMID:26710069

  12. Human T-cell leukemia virus type-1 antisense-encoded gene, Hbz, promotes T-lymphocyte proliferation.

    PubMed

    Arnold, Joshua; Zimmerman, Bevin; Li, Min; Lairmore, Michael D; Green, Patrick L

    2008-11-01

    Human T-cell leukemia virus type 1 (HTLV-1) basic leucine zipper factor (HBZ) is dispensable for HTLV-1-mediated cellular transformation in cell culture, but is required for efficient viral infectivity and persistence in rabbits. In most adult T-cell leukemia (ATL) cells, Tax oncoprotein expression is typically low or undetectable, whereas Hbz gene expression is maintained, suggesting that Hbz expression may support infected cell survival and, ultimately, leukemogenesis. Emerging data indicate that HBZ protein can interact with cAMP response element binding protein (CREB) and Jun family members, altering transcription factor binding and transactivation of both viral and cellular promoters. Herein, lentiviral vectors that express Hbz-specific short hairpin (sh)-RNA effectively decreased both Hbz mRNA and HBZ protein expression in transduced HTLV-1-transformed SLB-1 T cells. Hbz knockdown correlated with a significant decrease in T-cell proliferation in culture. Both SLB-1 and SLB-1-Hbz knockdown cells engrafted into inoculated NOD/SCID(gammachain-/-) mice to form solid tumors that also infiltrated multiple tissues. However, tumor formation and organ infiltration were significantly decreased in animals challenged with SLB-1-Hbz knockdown cells. Our data indicate that Hbz expression enhances the proliferative capacity of HTLV-1-infected T cells, playing a critical role in cell survival and ultimately HTLV-1 tumorigenesis in the infected host. PMID:18689544

  13. Activated T cell exosomes promote tumor invasion via Fas signaling pathway.

    PubMed

    Cai, Zhijian; Yang, Fei; Yu, Lei; Yu, Zhou; Jiang, Lingling; Wang, Qingqing; Yang, Yunshan; Wang, Lie; Cao, Xuetao; Wang, Jianli

    2012-06-15

    Activated T cells release bioactive Fas ligand (FasL) in exosomes, which subsequently induce self-apoptosis of T cells. However, their potential effects on cell apoptosis in tumors are still unknown. In this study, we purified exosomes expressing FasL from activated CD8(+) T cell from OT-I mice and found that activated T cell exosomes had little effect on apoptosis and proliferation of tumor cells but promoted the invasion of B16 and 3LL cancer cells in vitro via the Fas/FasL pathway. Activated T cell exosomes increased the amount of cellular FLICE inhibitory proteins and subsequently activated the ERK and NF-κB pathways, which subsequently increased MMP9 expression in the B16 murine melanoma cells. In a tumor-invasive model in vivo, we observed that the activated T cell exosomes promoted the migration of B16 tumor cells to lung. Interestingly, pretreatment with FasL mAb significantly reduced the migration of B16 tumor cells to lung. Furthermore, CD8 and FasL double-positive exosomes from tumor mice, but not normal mice, also increased the expression of MMP9 and promoted the invasive ability of B16 murine melanoma and 3LL lung cancer cells. In conclusion, our results indicate that activated T cell exosomes promote melanoma and lung cancer cell metastasis by increasing the expression of MMP9 via Fas signaling, revealing a new mechanism of tumor immune escape. PMID:22573809

  14. LPA Promotes T Cell Recruitment through Synthesis of CXCL13

    PubMed Central

    Hui, Weili; Zhao, Chenqi; Bourgoin, Sylvain G.

    2015-01-01

    Lysophosphatidic acid (LPA) is a bioactive phospholipid playing an important role in various inflammatory diseases by inducing expression and secretion of many inflammatory cytokines/chemokines. Here we report in a murine air pouch model of inflammation that LPA induced CXCL13 secretion in a time-dependent manner and with exacerbation of the response when LPA was administered after a pretreatment with TNF-α, a key inflammatory cytokine. LPA mediates recruitment of leukocytes, including that of CD3+ cells into unprimed and TNF-α-primed air pouches. CXCL13 neutralization using a blocking antibody injected into air pouches prior to administration of LPA into TNF-α-primed air pouches decreased CD3+ cell influx. Our data highlight that LPA-mediated CXCL13 secretion plays a role in T cell recruitment and participates in regulation of the inflammatory response. PMID:26339130

  15. Signal strength regulates antigen-mediated T-cell deceleration by distinct mechanisms to promote local exploration or arrest

    PubMed Central

    Moreau, Hélène D.; Lemaître, Fabrice; Garrod, Kym R.; Garcia, Zacarias; Lennon-Duménil, Ana-Maria; Bousso, Philippe

    2015-01-01

    T lymphocytes are highly motile cells that decelerate upon antigen recognition. These cells can either completely stop or maintain a low level of motility, forming contacts referred to as synapses or kinapses, respectively. Whether similar or distinct molecular mechanisms regulate T-cell deceleration during synapses or kinapses is unclear. Here, we used microfabricated channels and intravital imaging to observe and manipulate T-cell kinapses and synapses. We report that high-affinity antigen induced a pronounced deceleration selectively dependent on Ca2+ signals and actin-related protein 2/3 complex (Arp2/3) activity. In contrast, low-affinity antigens induced a switch of migration mode that promotes T-cell exploratory behavior, characterized by partial deceleration and frequent direction changes. This switch depended on T-cell receptor binding but was largely independent of downstream signaling. We propose that distinct mechanisms of T-cell deceleration can be triggered during antigenic recognition to favor local exploration and signal integration upon suboptimal stimulus and complete arrest on the best antigen-presenting cells. PMID:26371316

  16. CD31 is required on CD4+ T cells to promote T cell survival during Salmonella infection

    PubMed Central

    Ross, Ewan A; Coughlan, Ruth E; Flores-Langarica, Adriana; Bobat, Saeeda; Marshall, Jennifer L; Hussain, Khiyam; Charlesworth, James; Abhyankar, Nikita; Hitchcock, Jessica; Gil, Cristina; López-Macías, Constantino; Henderson, Ian R; Khan, Mahmood; Watson, Steve P; MacLennan, Ian C M; Buckley, Christopher D; Cunningham, Adam F

    2011-01-01

    Haematopoietic cells constitutively express CD31/PECAM1 a signalling, adhesion receptor associated with controlling responses to inflammatory stimuli. Although expressed on CD4+ T cells, its function on these cells is unclear. To address this we have used a model of systemic Salmonella infection that induces high levels of T cell activation and depends upon CD4+ T cells for resolution. Infection of CD31-deficient (CD31KO) mice demonstrates that these mice fail to control infection effectively. During infection, CD31KO mice have diminished numbers of total CD4+ T cells and IFN-γ-secreting Th1 cells. This is despite a higher proportion of CD31KO CD4+ T cells exhibiting an activated phenotype, and an undiminished capacity to prime normally and polarize to Th1. Reduced numbers of T cells reflected the increased propensity of naive and activated CD31KO T cells to undergo apoptosis after infection compared to wild-type (WT) T cells. Using adoptive transfer experiments we show that loss of CD31 on CD4+ T cells alone is sufficient to account for the defective CD31KO T cell accumulation. These data are consistent with CD31 helping to control T cell activation as in its absence T cells have a greater propensity to become activated, resulting in increased susceptibility to become apoptotic. The impact of CD31 loss on T cell homeostasis becomes most pronounced during severe, inflammatory and immunological stresses such as those caused by systemic Salmonella infection. This identifies a novel role for CD31 in regulating CD4 T cell homeostasis. PMID:21734076

  17. R-Ras Regulates Murine T Cell Migration and Intercellular Adhesion Molecule-1 Binding

    PubMed Central

    Yan, Xiaocai; Yan, Mingfei; Guo, Yihe; Singh, Gobind; Chen, Yuhong; Yu, Mei; Wang, Demin; Hillery, Cheryl A.; Chan, Andrew M.

    2015-01-01

    The trafficking of T-lymphocytes to peripheral draining lymph nodes is crucial for mounting an adaptive immune response. The role of chemokines in the activation of integrins via Ras-related small GTPases has been well established. R-Ras is a member of the Ras-subfamily of small guanosine-5’-triphosphate-binding proteins and its role in T cell trafficking has been investigated in R-Ras null mice (Rras−/−). An examination of the lymphoid organs of Rras−/− mice revealed a 40% reduction in the cellularity of the peripheral lymph nodes. Morphologically, the high endothelial venules of Rras−/− mice were more disorganized and less mature than those of wild-type mice. Furthermore, CD4+ and CD8+ T cells from Rras−/− mice had approximately 42% lower surface expression of L-selectin/CD62L. These aberrant peripheral lymph node phenotypes were associated with proliferative and trafficking defects in Rras−/− T cells. Furthermore, R-Ras could be activated by the chemokine, CCL21. Indeed, Rras−/− T cells had approximately 14.5% attenuation in binding to intercellular adhesion molecule 1 upon CCL21 stimulation. Finally, in a graft-versus host disease model, recipient mice that were transfused with Rras−/− T cells showed a significant reduction in disease severity when compared with mice transplanted with wild-type T cells. These findings implicate a role for R-Ras in T cell trafficking in the high endothelial venules during an effective immune response. PMID:26710069

  18. Loss of PTEN promotes resistance to T cell-mediated immunotherapy

    PubMed Central

    Peng, Weiyi; Chen, Jie Qing; Liu, Chengwen; Malu, Shruti; Creasy, Caitlin; Tetzlaff, Michael T; Xu, Chunyu; McKenzie, Jodi A; Zhang, Chunlei; Liang, Xiaoxuan; Williams, Leila J; Deng, Wanleng; Chen, Guo; Mbofung, Rina; Lazar, Alexander J; Torres-Cabala, Carlos A; Cooper, Zachary A; Chen, Pei-Ling; Tieu, Trang N; Spranger, Stefani; Yu, Xiaoxing; Bernatchez, Chantale; Forget, Marie-Andree; Haymaker, Cara; Amaria, Rodabe; McQuade, Jennifer L; Glitza, Isabella C; Cascone, Tina; Li, Haiyan S; Kwong, Lawrence N; Heffernan, Timothy P; Hu, Jianhua; Bassett, Roland L; Bosenberg, Marcus W; Woodman, Scott E; Overwijk, Willem W; Lizée, Gregory; Roszik, Jason; Gajewski, Thomas F; Wargo, Jennifer A; Gershenwald, Jeffrey E; Radvanyi, Laszlo; Davies, Michael A; Hwu, Patrick

    2015-01-01

    T cell-mediated immunotherapies are promising cancer treatments. However, most patients still fail to respond to these therapies. The molecular determinants of immune resistance are poorly understood. We show that loss of PTEN in tumor cells in preclinical models of melanoma inhibits T cell-mediated tumor killing and decreases T cell trafficking into tumors. In patients, PTEN loss correlates with decreased T cell infiltration at tumor sites, reduced likelihood of successful T cell expansion from resected tumors, and inferior outcomes with PD-1 inhibitor therapy. PTEN loss in tumor cells increased the expression of immunosuppressive cytokines, resulting in decreased T cell infiltration in tumors, and inhibited autophagy, which decreased T cell-mediated cell death. Treatment with a selective PI3Kβ inhibitor improved the efficacy of both anti-PD-1 and anti-CTLA4 antibodies in murine models. Together these findings demonstrate that PTEN loss promotes immune resistance and support the rationale to explore combinations of immunotherapies and PI3K-AKT pathway inhibitors. PMID:26645196

  19. Hydrophobic CDR3 residues promote the development of self-reactive T cells.

    PubMed

    Stadinski, Brian D; Shekhar, Karthik; Gómez-Touriño, Iria; Jung, Jonathan; Sasaki, Katsuhiro; Sewell, Andrew K; Peakman, Mark; Chakraborty, Arup K; Huseby, Eric S

    2016-08-01

    Studies of individual T cell antigen receptors (TCRs) have shed some light on structural features that underlie self-reactivity. However, the general rules that can be used to predict whether TCRs are self-reactive have not been fully elucidated. Here we found that the interfacial hydrophobicity of amino acids at positions 6 and 7 of the complementarity-determining region CDR3β robustly promoted the development of self-reactive TCRs. This property was found irrespective of the member of the β-chain variable region (Vβ) family present in the TCR or the length of the CDR3β. An index based on these findings distinguished Vβ2(+), Vβ6(+) and Vβ8.2(+) regulatory T cells from conventional T cells and also distinguished CD4(+) T cells selected by the major histocompatibility complex (MHC) class II molecule I-A(g7) (associated with the development of type 1 diabetes in NOD mice) from those selected by a non-autoimmunity-promoting MHC class II molecule I-A(b). Our results provide a means for distinguishing normal T cell repertoires versus autoimmunity-prone T cell repertoires. PMID:27348411

  20. Progesterone and HMOX-1 promote fetal growth by CD8+ T cell modulation

    PubMed Central

    Solano, María Emilia; Kowal, Mirka Katharina; O’Rourke, Greta Eugenia; Horst, Andrea Kristina; Modest, Kathrin; Plösch, Torsten; Barikbin, Roja; Remus, Chressen Catharina; Berger, Robert G.; Jago, Caitlin; Ho, Hoang; Sass, Gabriele; Parker, Victoria J.; Lydon, John P.; DeMayo, Francesco J.; Hecher, Kurt; Karimi, Khalil; Arck, Petra Clara

    2015-01-01

    Intrauterine growth restriction (IUGR) affects up to 10% of pregnancies in Western societies. IUGR is a strong predictor of reduced short-term neonatal survival and impairs long-term health in children. Placental insufficiency is often associated with IUGR; however, the molecular mechanisms involved in the pathogenesis of placental insufficiency and IUGR are largely unknown. Here, we developed a mouse model of fetal-growth restriction and placental insufficiency that is induced by a midgestational stress challenge. Compared with control animals, pregnant dams subjected to gestational stress exhibited reduced progesterone levels and placental heme oxygenase 1 (Hmox1) expression and increased methylation at distinct regions of the placental Hmox1 promoter. These stress-triggered changes were accompanied by an altered CD8+ T cell response, as evidenced by a reduction of tolerogenic CD8+CD122+ T cells and an increase of cytotoxic CD8+ T cells. Using progesterone receptor– or Hmox1-deficient mice, we identified progesterone as an upstream modulator of placental Hmox1 expression. Supplementation of progesterone or depletion of CD8+ T cells revealed that progesterone suppresses CD8+ T cell cytotoxicity, whereas the generation of CD8+CD122+ T cells is supported by Hmox1 and ameliorates fetal-growth restriction in Hmox1 deficiency. These observations in mice could promote the identification of pregnancies at risk for IUGR and the generation of clinical interventional strategies. PMID:25774501

  1. Hotspot autoimmune T cell receptor binding underlies pathogen and insulin peptide cross-reactivity

    PubMed Central

    Cole, David K.; Bulek, Anna M.; Dolton, Garry; Schauenberg, Andrea J.; Szomolay, Barbara; Trimby, Andrew; Jothikumar, Prithiviraj; Fuller, Anna; Skowera, Ania; Rossjohn, Jamie; Zhu, Cheng; Miles, John J.; Wooldridge, Linda; Rizkallah, Pierre J.; Sewell, Andrew K.

    2016-01-01

    The cross-reactivity of T cells with pathogen- and self-derived peptides has been implicated as a pathway involved in the development of autoimmunity. However, the mechanisms that allow the clonal T cell antigen receptor (TCR) to functionally engage multiple peptide–major histocompatibility complexes (pMHC) are unclear. Here, we studied multiligand discrimination by a human, preproinsulin reactive, MHC class-I–restricted CD8+ T cell clone (1E6) that can recognize over 1 million different peptides. We generated high-resolution structures of the 1E6 TCR bound to 7 altered peptide ligands, including a pathogen-derived peptide that was an order of magnitude more potent than the natural self-peptide. Evaluation of these structures demonstrated that binding was stabilized through a conserved lock-and-key–like minimal binding footprint that enables 1E6 TCR to tolerate vast numbers of substitutions outside of this so-called hotspot. Highly potent antigens of the 1E6 TCR engaged with a strong antipathogen-like binding affinity; this engagement was governed though an energetic switch from an enthalpically to entropically driven interaction compared with the natural autoimmune ligand. Together, these data highlight how T cell cross-reactivity with pathogen-derived antigens might break self-tolerance to induce autoimmune disease. PMID:27183389

  2. Hotspot autoimmune T cell receptor binding underlies pathogen and insulin peptide cross-reactivity.

    PubMed

    Cole, David K; Bulek, Anna M; Dolton, Garry; Schauenberg, Andrea J; Szomolay, Barbara; Rittase, William; Trimby, Andrew; Jothikumar, Prithiviraj; Fuller, Anna; Skowera, Ania; Rossjohn, Jamie; Zhu, Cheng; Miles, John J; Peakman, Mark; Wooldridge, Linda; Rizkallah, Pierre J; Sewell, Andrew K

    2016-06-01

    The cross-reactivity of T cells with pathogen- and self-derived peptides has been implicated as a pathway involved in the development of autoimmunity. However, the mechanisms that allow the clonal T cell antigen receptor (TCR) to functionally engage multiple peptide-major histocompatibility complexes (pMHC) are unclear. Here, we studied multiligand discrimination by a human, preproinsulin reactive, MHC class-I-restricted CD8+ T cell clone (1E6) that can recognize over 1 million different peptides. We generated high-resolution structures of the 1E6 TCR bound to 7 altered peptide ligands, including a pathogen-derived peptide that was an order of magnitude more potent than the natural self-peptide. Evaluation of these structures demonstrated that binding was stabilized through a conserved lock-and-key-like minimal binding footprint that enables 1E6 TCR to tolerate vast numbers of substitutions outside of this so-called hotspot. Highly potent antigens of the 1E6 TCR engaged with a strong antipathogen-like binding affinity; this engagement was governed though an energetic switch from an enthalpically to entropically driven interaction compared with the natural autoimmune ligand. Together, these data highlight how T cell cross-reactivity with pathogen-derived antigens might break self-tolerance to induce autoimmune disease. PMID:27183389

  3. β-Catenin induces T-cell transformation by promoting genomic instability

    PubMed Central

    Dose, Marei; Emmanuel, Akinola Olumide; Chaumeil, Julie; Zhang, Jiangwen; Sun, Tianjiao; Germar, Kristine; Aghajani, Katayoun; Davis, Elizabeth M.; Keerthivasan, Shilpa; Bredemeyer, Andrea L.; Sleckman, Barry P.; Rosen, Steven T.; Skok, Jane A.; Le Beau, Michelle M.; Georgopoulos, Katia; Gounari, Fotini

    2014-01-01

    Deregulated activation of β-catenin in cancer has been correlated with genomic instability. During thymocyte development, β-catenin activates transcription in partnership with T-cell–specific transcription factor 1 (Tcf-1). We previously reported that targeted activation of β-catenin in thymocytes (CAT mice) induces lymphomas that depend on recombination activating gene (RAG) and myelocytomatosis oncogene (Myc) activities. Here we show that these lymphomas have recurring Tcra/Myc translocations that resulted from illegitimate RAG recombination events and resembled oncogenic translocations previously described in human T-ALL. We therefore used the CAT animal model to obtain mechanistic insights into the transformation process. ChIP-seq analysis uncovered a link between Tcf-1 and RAG2 showing that the two proteins shared binding sites marked by trimethylated histone-3 lysine-4 (H3K4me3) throughout the genome, including near the translocation sites. Pretransformed CAT thymocytes had increased DNA damage at the translocating loci and showed altered repair of RAG-induced DNA double strand breaks. These cells were able to survive despite DNA damage because activated β-catenin promoted an antiapoptosis gene expression profile. Thus, activated β-catenin promotes genomic instability that leads to T-cell lymphomas as a consequence of altered double strand break repair and increased survival of thymocytes with damaged DNA. PMID:24371308

  4. Differential utilization of binding loop flexibility in T cell receptor ligand selection and cross-reactivity.

    PubMed

    Ayres, Cory M; Scott, Daniel R; Corcelli, Steven A; Baker, Brian M

    2016-01-01

    Complementarity determining region (CDR) loop flexibility has been suggested to play an important role in the selection and binding of ligands by T cell receptors (TCRs) of the cellular immune system. However, questions remain regarding the role of loop motion in TCR binding, and crystallographic structures have raised questions about the extent to which generalizations can be made. Here we studied the flexibility of two structurally well characterized αβ TCRs, A6 and DMF5. We found that the two receptors utilize loop motion very differently in ligand binding and cross-reactivity. While the loops of A6 move rapidly in an uncorrelated fashion, those of DMF5 are substantially less mobile. Accordingly, the mechanisms of binding and cross-reactivity are very different between the two TCRs: whereas A6 relies on conformational selection to select and bind different ligands, DMF5 uses a more rigid, permissive architecture with greater reliance on slower motions or induced-fit. In addition to binding site flexibility, we also explored whether ligand-binding resulted in common dynamical changes in A6 and DMF5 that could contribute to TCR triggering. Although binding-linked motional changes propagated throughout both receptors, no common features were observed, suggesting that changes in nanosecond-level TCR structural dynamics do not contribute to T cell signaling. PMID:27118724

  5. Differential utilization of binding loop flexibility in T cell receptor ligand selection and cross-reactivity

    PubMed Central

    Ayres, Cory M.; Scott, Daniel R.; Corcelli, Steven A.; Baker, Brian M.

    2016-01-01

    Complementarity determining region (CDR) loop flexibility has been suggested to play an important role in the selection and binding of ligands by T cell receptors (TCRs) of the cellular immune system. However, questions remain regarding the role of loop motion in TCR binding, and crystallographic structures have raised questions about the extent to which generalizations can be made. Here we studied the flexibility of two structurally well characterized αβ TCRs, A6 and DMF5. We found that the two receptors utilize loop motion very differently in ligand binding and cross-reactivity. While the loops of A6 move rapidly in an uncorrelated fashion, those of DMF5 are substantially less mobile. Accordingly, the mechanisms of binding and cross-reactivity are very different between the two TCRs: whereas A6 relies on conformational selection to select and bind different ligands, DMF5 uses a more rigid, permissive architecture with greater reliance on slower motions or induced-fit. In addition to binding site flexibility, we also explored whether ligand-binding resulted in common dynamical changes in A6 and DMF5 that could contribute to TCR triggering. Although binding-linked motional changes propagated throughout both receptors, no common features were observed, suggesting that changes in nanosecond-level TCR structural dynamics do not contribute to T cell signaling. PMID:27118724

  6. Impaired T-cell survival promotes mucosal inflammatory disease in SHIP1-deficient mice.

    PubMed

    Park, M Y; Srivastava, N; Sudan, R; Viernes, D R; Chisholm, J D; Engelman, R W; Kerr, W G

    2014-11-01

    T cells have a critical role in immune surveillance at mucosal surfaces. SHIP1(-/-) mice succumb to mucosal inflammatory disease that afflicts the lung and small intestine (SI). The basis of this condition has not been defined. Here we show that SHIP1 is required for the normal persistence and survival of T cells in mucosal tissues. We find that CD4 and CD8 effector T cells are reduced; however, Treg cells are increased in the SI and lungs of SHIP1(-/-) and CD4CreSHIP(flox/flox) mice. Furthermore, a subset of T cells in the SI of SHIP1(-/-) mice are FasL(+) and are more susceptible to extrinsic cell death. Mechanistic analyses showed that SHIP1 associates with the death receptor CD95/Fas and treatment with a Caspase 8 inhibitor prevents SHIP1 inhibitor-mediated T-cell death. Notably, mucosal inflammation in SHIP1(-/-) mice is reduced by treatment with a Caspase 8 inhibitor. We also find that the incidence of Crohn's disease (CD) and pneumonia is significantly increased in mice with dual T and myeloid lineage SHIP1 deletion but not in single lineage-deleted mice. Thus, by promoting survival of protective T cells, thereby preventing an inflammatory myeloid response, SHIP1 maintains an appropriate balance of innate immune function at mucosal surfaces necessary for immune homeostasis. PMID:24781051

  7. PD-1 promotes immune exhaustion by inducing antiviral T cell motility paralysis

    PubMed Central

    Zinselmeyer, Bernd H.; Heydari, Sara; Sacristán, Catarina; Nayak, Debasis; Cammer, Michael; Herz, Jasmin; Cheng, Xiaoxiao; Davis, Simon J.; Dustin, Michael L.

    2013-01-01

    Immune responses to persistent viral infections and cancer often fail because of intense regulation of antigen-specific T cells—a process referred to as immune exhaustion. The mechanisms that underlie the induction of exhaustion are not completely understood. To gain novel insights into this process, we simultaneously examined the dynamics of virus-specific CD8+ and CD4+ T cells in the living spleen by two-photon microscopy (TPM) during the establishment of an acute or persistent viral infection. We demonstrate that immune exhaustion during viral persistence maps anatomically to the splenic marginal zone/red pulp and is defined by prolonged motility paralysis of virus-specific CD8+ and CD4+ T cells. Unexpectedly, therapeutic blockade of PD-1–PD-L1 restored CD8+ T cell motility within 30 min, despite the presence of high viral loads. This result was supported by planar bilayer data showing that PD-L1 localizes to the central supramolecular activation cluster, decreases antiviral CD8+ T cell motility, and promotes stable immunological synapse formation. Restoration of T cell motility in vivo was followed by recovery of cell signaling and effector functions, which gave rise to a fatal disease mediated by IFN-γ. We conclude that motility paralysis is a manifestation of immune exhaustion induced by PD-1 that prevents antiviral CD8+ T cells from performing their effector functions and subjects them to prolonged states of negative immune regulation. PMID:23530125

  8. Interleukin-1-induced promotion of T-cell differentiation in mice immunized with killed Listeria monocytogenes.

    PubMed Central

    Igarashi, K; Mitsuyama, M; Muramori, K; Tsukada, H; Nomoto, K

    1990-01-01

    We studied the effects of administration of recombinant interleukin-1 alpha (rIL-1 alpha) to mice after immunization with killed Listeria monocytogenes cells on the promotion of the functional differentiation of T cells in vivo. Mice immunized with killed L. monocytogenes were unable to express cell-mediated immunity to specific antigen in vivo, as determined by delayed-type hypersensitivity (DTH) and acquired cellular resistance (ACR), and splenic T cells obtained from such mice were unable to respond to rIL-2 and specific antigen and to produce IL-2 after antigenic restimulation in vitro. When rIL-1 alpha was given to mice after immunization with killed bacteria. T cells became capable of responding to rIL-2 and specific antigen in vitro. These functions of T cells were similar to those from mice immunized with viable listeriae. Moreover, using a local passive transfer system, it was found that effector T cells mediating DTH but not ACR to L. monocytogenes were generated in mice treated with rIL-1 alpha after immunization with killed bacteria. These T cells were able to produce macrophage chemotactic factor but not macrophage-activating factor or gamma interferon in vitro in response to stimulation with specific antigen. These results suggest that in vivo administration of rIL-1 alpha facilitates the maturation of antigen-specific T cells mediating DTH and that different effector T cells mediating DTH or ACR are involved in cell-mediated immunity to L. monocytogenes. PMID:2123829

  9. Fibrocyte and T cell interactions promote disease pathogenesis in rheumatoid arthritis.

    PubMed

    Galligan, Carole L; Keystone, Edward C; Fish, Eleanor N

    2016-05-01

    Rheumatoid arthritis (RA) is a systemic autoimmune disease. We previously identified a circulating cell population, fibrocytes, which is activated early in disease. As RA is characterized by the formation of autoantibodies and autoreactive T cells, which often precede symptom onset, the objective of these studies was to characterize fibrocyte activation in the context of T cell activation. Multidimensional flow cytometry was used to characterize the activation status of peripheral blood (PB) fibrocytes and T cells derived from RA patients with different levels of disease activity. Compared to healthy controls, fibrocytes from RA patients exhibited increased activation, denoted as elevated levels of phosphorylation of STAT3 and NF-κB. RA patients had higher numbers of circulating activated Th17 cells and Tregs compared with healthy controls, Th17 cell numbers being higher in patients with moderate to high disease activity. Additionally, increased numbers of FOXP3+ RORγt+ double positive CD4+ T cells were observed in RA patients with more severe disease. Our data confirm that circulating fibrocytes are expanded in RA and that there is a direct correlation between the increase in number of activated fibrocytes and increased number of CD4+ T cells. Moreover, our data suggest that interactions between circulating fibrocytes and activated T cells may promote disease activity. Specifically, we provide in vitro evidence that mouse-derived CD4+ T cells produce GM-CSF which induces fibrocyte proliferation. In turn, activated fibrocytes produce IL-6, promoting Th17 polarization. PMID:26948996

  10. Interplay between T Cell Receptor Binding Kinetics and the Level of Cognate Peptide Presented by Major Histocompatibility Complexes Governs CD8+ T Cell Responsiveness*

    PubMed Central

    Irving, Melita; Zoete, Vincent; Hebeisen, Michael; Schmid, Daphné; Baumgartner, Petra; Guillaume, Philippe; Romero, Pedro; Speiser, Daniel; Luescher, Immanuel; Rufer, Nathalie; Michielin, Olivier

    2012-01-01

    Through a rational design approach, we generated a panel of HLA-A*0201/NY-ESO-1157–165-specific T cell receptors (TCR) with increasing affinities of up to 150-fold from the wild-type TCR. Using these TCR variants which extend just beyond the natural affinity range, along with an extreme supraphysiologic one having 1400-fold enhanced affinity, and a low-binding one, we sought to determine the effect of TCR binding properties along with cognate peptide concentration on CD8+ T cell responsiveness. Major histocompatibility complexes (MHC) expressed on the surface of various antigen presenting cells were peptide-pulsed and used to stimulate human CD8+ T cells expressing the different TCR via lentiviral transduction. At intermediate peptide concentration we measured maximum cytokine/chemokine secretion, cytotoxicity, and Ca2+ flux for CD8+ T cells expressing TCR within a dissociation constant (KD) range of ∼1–5 μm. Under these same conditions there was a gradual attenuation in activity for supraphysiologic affinity TCR with KD < ∼1 μm, irrespective of CD8 co-engagement and of half-life (t1/2 = ln 2/koff) values. With increased peptide concentration, however, the activity levels of CD8+ T cells expressing supraphysiologic affinity TCR were gradually restored. Together our data support the productive hit rate model of T cell activation arguing that it is not the absolute number of TCR/pMHC complexes formed at equilibrium, but rather their productive turnover, that controls levels of biological activity. Our findings have important implications for various immunotherapies under development such as adoptive cell transfer of TCR-engineered CD8+ T cells, as well as for peptide vaccination strategies. PMID:22549784

  11. Jun Kinase Phosphorylates and Regulates the DNA Binding Activity of an Octamer Binding Protein, T-Cell Factor β1†

    PubMed Central

    Kasibhatla, Shailaja; Tailor, Pankaj; Bonefoy-Berard, Nathalie; Mustelin, Tomas; Altman, Amnon; Fotedar, Arun

    1999-01-01

    POU domain proteins have been implicated as key regulators during development and lymphocyte activation. The POU domain protein T-cell factor β1 (TCFβ1), which binds octamer and octamer-related sequences, is a potent transactivator. In this study, we showed that TCFβ1 is phosphorylated following activation via the T-cell receptor or by stress-induced signals. Phosphorylation of TCFβ1 occurred predominantly at serine and threonine residues. Signals which upregulate Jun kinase (JNK)/stress-activated protein kinase activity also lead to association of JNK with TCFβ1. JNK associates with the activation domain of TCFβ1 and phosphorylates its DNA binding domain. The phosphorylation of recombinant TCFβ1 by recombinant JNK enhances the ability of TCFβ1 to bind to a consensus octamer motif. Consistent with this conclusion, TCFβ1 upregulates reporter gene transcription in an activation- and JNK-dependent manner. In addition, inhibition of JNK activity by catalytically inactive MEKK (in which methionine was substituted for the lysine at position 432) also inhibits the ability of TCFβ1 to drive inducible transcription from the interleukin-2 promoter. These results suggest that stress-induced signals and T-cell activation induce JNK, which then acts on multiple cis sequences by modulating distinct transactivators like c-Jun and TCFβ1. This demonstrates a coupling between the JNK activation pathway and POU domain proteins and implicates TCFβ1 as a physiological target in the JNK signal transduction pathway leading to coordinated biological responses. PMID:10022889

  12. Ezrin/Radixin/Moesin Proteins and Flotillins Cooperate to Promote Uropod Formation in T Cells

    PubMed Central

    Martinelli, Sibylla; Chen, Emily J. H.; Clarke, Fiona; Lyck, Ruth; Affentranger, Sarah; Burkhardt, Janis K.; Niggli, Verena

    2013-01-01

    T cell uropods are enriched in specific proteins including adhesion receptors such as P-selectin glycoprotein ligand-1 (PSGL-1), lipid raft-associated proteins such as flotillins and ezrin/radixin/moesin (ERM) proteins which associate with cholesterol-rich raft domains and anchor adhesion receptors to the actin cytoskeleton. Using dominant mutants and siRNA technology we have tested the interactions among these proteins and their role in shaping the T cell uropod. Expression of wild type (WT) ezrin-EGFP failed to affect the morphology of human T cells or chemokine-induced uropod recruitment of PSGL-1 and flotillin-1 and -2. In contrast, expression of constitutively active T567D ezrin-EGFP induced a motile, polarized phenotype in some of the transfected T cells, even in the absence of chemokine. These cells featured F-actin-rich ruffles in the front and uropod enrichment of PSGL-1 and flotillins. T567D ezrin-EGFP was itself strongly enriched in the rear of the polarized T cells. Uropod formation induced by T567D ezrin-EGFP was actin-dependent as it was attenuated by inhibition of Rho-kinase or myosin II, and abolished by disruption of actin filaments. While expression of constitutively active ezrin enhanced cell polarity, expression of a dominant-negative deletion mutant of ezrin, 1–310 ezrin-EGFP, markedly reduced uropod formation induced by the chemokine SDF-1, T cell front-tail polarity, and capping of PSGL-1 and flotillins. Transfection of T cells with WT or T567D ezrin did not affect chemokine-mediated chemotaxis whereas 1–310 ezrin significantly impaired spontaneous 2D migration and chemotaxis. siRNA-mediated downregulation of flotillins in murine T cells attenuated moesin capping and uropod formation, indicating that ERM proteins and flotillins cooperate in uropod formation. In summary, our results indicate that activated ERM proteins function together with flotillins to promote efficient chemotaxis of T cells by structuring the uropod of migrating T

  13. Tumor sialylation impedes T cell mediated anti-tumor responses while promoting tumor associated-regulatory T cells

    PubMed Central

    Perdicchio, Maurizio; Cornelissen, Lenneke A. M.; Streng-Ouwehand, Ingeborg; Engels, Steef; Verstege, Marleen I.; Boon, Louis; Geerts, Dirk

    2016-01-01

    The increased presence of sialylated glycans on the tumor surface has been linked to poor prognosis, yet the effects on tumor-specific T cell immunity are hardly studied. We here show that hypersialylation of B16 melanoma substantially influences tumor growth by preventing the formation of effector T cells and facilitating the presence of high regulatory T cell (Treg) frequencies. Knock-down of the sialic acid transporter created “sialic acid low” tumors, that grew slower in-vivo than hypersialylated tumors, altered the Treg/Teffector balance, favoring immunological tumor control. The enhanced effector T cell response in developing “sialic acid low” tumors was preceded by and dependent on an increased influx and activity of Natural Killer (NK) cells. Thus, tumor hypersialylation orchestrates immune escape at the level of NK and Teff/Treg balance within the tumor microenvironment, herewith dampening tumor-specific T cell control. Reducing sialylation provides a therapeutic option to render tumors permissive to immune attack. PMID:26741508

  14. Tumor sialylation impedes T cell mediated anti-tumor responses while promoting tumor associated-regulatory T cells.

    PubMed

    Perdicchio, Maurizio; Cornelissen, Lenneke A M; Streng-Ouwehand, Ingeborg; Engels, Steef; Verstege, Marleen I; Boon, Louis; Geerts, Dirk; van Kooyk, Yvette; Unger, Wendy W J

    2016-02-23

    The increased presence of sialylated glycans on the tumor surface has been linked to poor prognosis, yet the effects on tumor-specific T cell immunity are hardly studied. We here show that hypersialylation of B16 melanoma substantially influences tumor growth by preventing the formation of effector T cells and facilitating the presence of high regulatory T cell (Treg) frequencies. Knock-down of the sialic acid transporter created "sialic acid low" tumors, that grew slower in-vivo than hypersialylated tumors, altered the Treg/Teffector balance, favoring immunological tumor control. The enhanced effector T cell response in developing "sialic acid low" tumors was preceded by and dependent on an increased influx and activity of Natural Killer (NK) cells. Thus, tumor hypersialylation orchestrates immune escape at the level of NK and Teff/Treg balance within the tumor microenvironment, herewith dampening tumor-specific T cell control. Reducing sialylation provides a therapeutic option to render tumors permissive to immune attack. PMID:26741508

  15. LMO2 Oncoprotein Stability in T-Cell Leukemia Requires Direct LDB1 Binding

    PubMed Central

    Layer, Justin H.; Alford, Catherine E.; McDonald, W. Hayes

    2015-01-01

    LMO2 is a component of multisubunit DNA-binding transcription factor complexes that regulate gene expression in hematopoietic stem and progenitor cell development. Enforced expression of LMO2 causes leukemia by inducing hematopoietic stem cell-like features in T-cell progenitor cells, but the biochemical mechanisms of LMO2 function have not been fully elucidated. In this study, we systematically dissected the LMO2/LDB1-binding interface to investigate the role of this interaction in T-cell leukemia. Alanine scanning mutagenesis of the LIM interaction domain of LDB1 revealed a discrete motif, R320LITR, required for LMO2 binding. Most strikingly, coexpression of full-length, wild-type LDB1 increased LMO2 steady-state abundance, whereas coexpression of mutant proteins deficient in LMO2 binding compromised LMO2 stability. These mutant LDB1 proteins also exerted dominant negative effects on growth and transcription in diverse leukemic cell lines. Mass spectrometric analysis of LDB1 binding partners in leukemic lines supports the notion that LMO2/LDB1 function in leukemia occurs in the context of multisubunit complexes, which also protect the LMO2 oncoprotein from degradation. Collectively, these data suggest that the assembly of LMO2 into complexes, via direct LDB1 interaction, is a potential molecular target that could be exploited in LMO2-driven leukemias resistant to existing chemotherapy regimens. PMID:26598604

  16. CTLA-4 promotes Foxp3 induction and regulatory T cell accumulation in the intestinal lamina propria.

    PubMed

    Barnes, M J; Griseri, T; Johnson, A M F; Young, W; Powrie, F; Izcue, A

    2013-03-01

    Thymic induction of CD4(+)Foxp3(+) regulatory T (Treg) cells relies on CD28 costimulation and high-affinity T-cell receptor (TCR) signals, whereas Foxp3 (forkhead box P3) induction on activated peripheral CD4(+) T cells is inhibited by these signals. Accordingly, the inhibitory molecule CTLA-4 (cytotoxic T-lymphocyte antigen 4) promoted, but was not essential for CD4(+) T-cell Foxp3 induction in vitro. We show that CTLA-4-deficient cells are equivalent to wild-type cells in the thymic induction of Foxp3 and maintenance of Foxp3 populations in the spleen and mesenteric lymph nodes, but their accumulation in the colon, where Treg cells specific for commensal bacteria accumulate, is impaired. In a T cell-transfer model of colitis, the two known CTLA-4 ligands, B7-1 and B7-2, had largely redundant roles in inducing inflammation and promoting Treg cell function. However, B7-2 proved more efficient than B7-1 in inducing Foxp3 in vitro and in vivo. Our data reveal an unappreciated role for CTLA-4 in establishing the Foxp3(+) compartment in the intestine. PMID:22910217

  17. Insulin promotes T cell recovery in a murine model of autoimmune myocarditis.

    PubMed

    Zhang, Y; Zhuang, R; Geng, C; Cai, X; Lei, W; Tian, N; Gao, F

    2013-01-01

    Glucose-insulin-potassium (GIK) is a useful adjunct to myocarditis. Besides its essential action in energy metabolism, insulin also exerts an anti-inflammatory effect. This study investigated the effect of insulin on myocardial inflammation in experimental autoimmune myocarditis (EAM) in mice and its potential role in T cell regulation. Mice were divided randomly into a normal control group, a saline-treated EAM group and an insulin-treated EAM group. The histopathological changes of myocardium, α-myosin heavy chain (MyHCα)(614-629) antigen-specific autoantibody titre, the serum level of cardiac troponin I (cTnI), mitogen-activated protein kinase (MAPK) family members' activity and content were measured. Furthermore, the phenotype of T lymphocyte subsets in splenocytes was analysed to evaluate the immune status of mice. Insulin reduced serum cTnI of EAM mice on days 14 and 21 (P < 0·05) after immunization, with no changes in blood glucose and autoantibody production. Western blot revealed that extracellular signal-regulated protein kinase (ERK1/2) may be a determining factor in this process. Total ERK1/2 and phospho-ERK1/2 (p-ERK1/2) were both up-regulated in insulin-treated mice after immunization. We also found that insulin treatment promoted T cell recovery without changing the naive-to-memory T-cell ratio; in particular, CD3(+) T cells in insulin-treated mice proliferated more vigorously than in control mice (P < 0·05). We report here for the first time that insulin alleviates myocarditis in the EAM model. These data show that insulin has a direct effect on T cell proliferation in EAM. It is possible that GIK or insulin may assist T cell recovery towards normal in myocarditis, especially for diabetic or hyperglycaemic patients. PMID:23199322

  18. Controlled delivery of a metabolic modulator promotes regulatory T cells and restrains autoimmunity.

    PubMed

    Gammon, Joshua M; Tostanoski, Lisa H; Adapa, Arjun R; Chiu, Yu-Chieh; Jewell, Christopher M

    2015-07-28

    Autoimmune disorders occur when the immune system abnormally recognizes and attacks self-molecules. Dendritic cells (DCs) play a powerful role in initiating adaptive immune response, and are therefore a recent target for autoimmune therapies. N-Phenyl-7-(hydroxyimino)cyclopropa[b]chromen-1a-carboxamide (PHCCC), a small molecule glutamate receptor enhancer, alters how DCs metabolize glutamate, skewing cytokine secretion to bias T cell function. These effects provide protection in mouse models of multiple sclerosis (MS) by polarizing T cells away from inflammatory TH17 cells and toward regulatory T cells (TREG) when mice receive daily systemic injections of PHCCC. However, frequent, continued treatment is required to generate and maintain therapeutic benefits. Thus, the use of PHCCC is limited by poor solubility, the need for frequent dosing, and cell toxicity. We hypothesized that controlled release of PHCCC from degradable nanoparticles (NPs) might address these challenges by altering DC function to maintain efficacy with reduced treatment frequency and toxicity. This idea could serve as a new strategy for harnessing biomaterials to polarize immune function through controlled delivery of metabolic modulators. PHCCC was readily encapsulated in nanoparticles, with controlled release of 89% of drug into media over three days. Culture of primary DCs or DC and T cell co-cultures with PHCCC NPs reduced DC activation and secretion of pro-inflammatory cytokines, while shifting T cells away from TH17 and toward TREG phenotypes. Importantly, PHCCC delivered to cells in NPs was 36-fold less toxic compared with soluble PHCCC. Treatment of mice with PHCCC NPs every three days delayed disease onset and decreased disease severity compared with mice treated with soluble drug at the same dose and frequency. These results highlight the potential to promote tolerance through controlled delivery of metabolic modulators that alter DC signaling to polarize T cells, and suggest future

  19. IL-15 promotes the survival of naive and memory phenotype CD8+ T cells.

    PubMed

    Berard, Marion; Brandt, Katja; Bulfone-Paus, Silvia; Tough, David F

    2003-05-15

    IL-15 stimulates the proliferation of memory phenotype CD44(high)CD8(+) T cells and is thought to play a key role in regulating the turnover of these cells in vivo. We have investigated whether IL-15 also has the capacity to affect the life span of naive phenotype (CD44(low)) CD8(+) T cells. We report that IL-15 promotes the survival of both CD44(low) and CD44(high) CD8(+) T cells, doing so at much lower concentrations than required to induce proliferation of CD44(high) cells. Rescue from apoptosis was associated with the up-regulation of Bcl-2 in both cell types, whereas elevated expression of Bcl-x(L) was observed among CD44(high) but not CD44(low) CD8(+) cells. An investigation into the role of IL-15R subunits in mediating the effects of IL-15 revealed distinct contributions of the alpha- and beta- and gamma-chains. Most strikingly, IL-15R alpha was not essential for either induction of proliferation or promotion of survival by IL-15, but did greatly enhance the sensitivity of cells to low concentrations of IL-15. By contrast, the beta- and gamma-chains of the IL-15R were absolutely required for the proliferative and pro-survival effects of IL-15, although it was not necessary for CD44(high)CD8(+) cells to express higher levels of IL-15R beta than CD44(low) cells to proliferate in response to IL-15. These results show that IL-15 has multiple effects on CD8 T cells and possesses the potential to regulate the life span of naive as well as memory CD8(+) T cells. PMID:12734346

  20. trans activation of the tumor necrosis factor alpha promoter by the human T-cell leukemia virus type I Tax1 protein.

    PubMed Central

    Albrecht, H; Shakhov, A N; Jongeneel, C V

    1992-01-01

    In a cotransfection assay, the human T-cell leukemia virus type I Tax1 gene product specifically activated transcription from the mouse tumor necrosis factor alpha promoter. The activation patterns of 5' deletion mutants, artificial enhancer constructs, and point mutations in the promoter indicate that the major Tax1-responsive element is a site at position -655 which binds the NF-kappa B/rel and NF-GMa transcription factors. Images PMID:1527856

  1. CTLA-4 blockade plus adoptive T cell transfer promotes optimal melanoma immunity in mice

    PubMed Central

    Mahvi, David A.; Meyers, Justin V.; Tatar, Andrew J.; Contreras, Amanda; Suresh, M.; Leverson, Glen E.; Sen, Siddhartha; Cho, Clifford S.

    2014-01-01

    Immunotherapeutic approaches to the treatment of advanced melanoma have relied on strategies that augment the responsiveness of endogenous tumor-specific T cell populations (e.g., CTLA-4 blockade-mediated checkpoint inhibition) or introduce exogenously-prepared tumor-specific T cell populations (e.g., adoptive cell transfer). Although both approaches have shown considerable promise, response rates to these therapies remain suboptimal. We hypothesized that a combinatorial approach to immunotherapy using both CTLA-4 blockade and non-lymphodepletional adoptive cell transfer could offer additive therapeutic benefit. C57BL/6 mice were inoculated with syngeneic B16F10 melanoma tumors transfected to express low levels of the lymphocytic choriomeningitis virus peptide GP33 (B16GP33), and treated with no immunotherapy, CTLA-4 blockade, adoptive cell transfer, or combination immunotherapy of CTLA-4 blockade with adoptive cell transfer. Combination immunotherapy resulted in optimal control of B16GP33 melanoma tumors. Combination immunotherapy promoted a stronger local immune response reflected by enhanced tumor-infiltrating lymphocyte populations, as well as a stronger systemic immune responses reflected by more potent tumor antigen-specific T cell activity in splenocytes. In addition, whereas both CTLA-4 blockade and combination immunotherapy were able to promote long-term immunity against B16GP33 tumors, only combination immunotherapy was capable of promoting immunity against parental B16F10 tumors as well. Our findings suggest that a combinatorial approach using CTLA-4 blockade with non-lymphodepletional adoptive cell transfer may promote additive endogenous and exogenous T cell activities that enable greater therapeutic efficacy in the treatment of melanoma. PMID:25658614

  2. Single MHC Mutation Eliminates Enthalpy Associated with T Cell Receptor Binding

    PubMed Central

    Miller, Peter J.; Pazy, Yael; Conti, Brian; Riddle, David; Appella, Ettore; Collins, Edward J.

    2007-01-01

    SUMMARY The keystone of the adaptive immune response is T cell receptor (TCR) recognition of peptide presented by Major Histocompatibility Complex (pMHC) molecules. The co-crystal structure of AHIII TCR bound to the MHC, HLA-A2, showed a large interface with an atypical binding orientation. MHC mutations in the interface of the proteins were tested for changes in TCR recognition. From the range of responses observed, three representative HLA-A2 mutants, T163A, W167A, and K66A, was selected for further study. Binding constants and co-crystal structures of the AHIII TCR and the three mutants were determined. K66 in HLA-A2 makes contacts with both peptide and TCR and previously has been identified as a critical residue for recognition by numerous TCR. The K66A mutation resulted in the lowest AHIII T cell response and the lowest binding affinity, which suggests T cell response may correlate with affinity. Importantly, the K66A mutation does not affect the conformation of the peptide. The change in affinity appears to be due to a loss in hydrogen bonds in the interface as a result of a conformational change in the TCR complementarity-determining region 3 (CDR3) loop. Isothermal titration calorimetry confirmed the loss of hydrogen bonding by a large loss in enthalpy. Our findings are inconsistent with the notion that the CDR1 and CDR2 loops of the TCR are responsible for MHC restriction, while the CDR3 loops interact solely with the peptide. Instead, we present here a MHC mutation that does not change the conformation of the peptide, yet results in an altered conformation of a CDR3. PMID:17825839

  3. Characterization of human platelet binding of recombinant T cell receptor ligand

    PubMed Central

    2010-01-01

    Background Recombinant T cell receptor ligands (RTLs) are bio-engineered molecules that may serve as novel therapeutic agents for the treatment of neuroinflammatory conditions such as multiple sclerosis (MS). RTLs contain membrane distal α1 plus β1 domains of class II major histocompatibility complex linked covalently to specific peptides that can be used to regulate T cell responses and inhibit experimental autoimmune encephalomyelitis (EAE). The mechanisms by which RTLs impede local recruitment and retention of inflammatory cells in the CNS, however, are not completely understood. Methods We have recently shown that RTLs bind strongly to B cells, macrophages, and dendritic cells, but not to T cells, in an antigenic-independent manner, raising the question whether peripheral blood cells express a distinct RTL-receptor. Our study was designed to characterize the molecular mechanisms by which RTLs bind human blood platelets, and the ability of RTL to modulate platelet function. Results Our data demonstrate that human blood platelets support binding of RTL. Immobilized RTL initiated platelet intracellular calcium mobilization and lamellipodia formation through a pathway dependent upon Src and PI3 kinases signaling. The presence of RTL in solution reduced platelet aggregation by collagen, while treatment of whole blood with RTL prolonged occlusive thrombus formation on collagen. Conclusions Platelets, well-known regulators of hemostasis and thrombosis, have been implicated in playing a major role in inflammation and immunity. This study provides the first evidence that blood platelets express a functional RTL-receptor with a putative role in modulating pathways of neuroinflammation. PMID:21059245

  4. MicroRNA-155 promotes autoimmune inflammation by enhancing inflammatory T cell development.

    PubMed

    O'Connell, Ryan M; Kahn, Daniel; Gibson, William S J; Round, June L; Scholz, Rebecca L; Chaudhuri, Aadel A; Kahn, Melissa E; Rao, Dinesh S; Baltimore, David

    2010-10-29

    Mammalian noncoding microRNAs (miRNAs) are a class of gene regulators that have been linked to immune system function. Here, we have investigated the role of miR-155 during an autoimmune inflammatory disease. Consistent with a positive role for miR-155 in mediating inflammatory responses, Mir155(-/-) mice were highly resistant to experimental autoimmune encephalomyelitis (EAE). miR-155 functions in the hematopoietic compartment to promote the development of inflammatory T cells including the T helper 17 (Th17) cell and Th1 cell subsets. Furthermore, the major contribution of miR-155 to EAE was CD4(+) T cell intrinsic, whereas miR-155 was also required for optimum dendritic cell production of cytokines that promoted Th17 cell formation. Our study shows that one aspect of miR-155 function is the promotion of T cell-dependent tissue inflammation, suggesting that miR-155 might be a promising therapeutic target for the treatment of autoimmune disorders. PMID:20888269

  5. T-cell hybridoma specific for a cytochrome c peptide: specific antigen binding and interleukin 2 production.

    PubMed Central

    Carel, S; Bron, C; Corradin, G

    1983-01-01

    T-cell hybridomas were obtained after fusion of BW 5147 thymoma and long-term cultured T cells specific for cytochrome c peptide 66-80 derivatized with a 2,4-dinitroaminophenyl (DNAP) group. The resulting hybridomas were selected for their capacity to specifically bind to soluble radiolabeled peptide antigen. One T-cell hybrid was positive for antigen binding. This hybrid T cell exhibits surface phenotypic markers of the parent antigen-specific T cells. The binding could be inhibited either by an excess of unlabeled homologous antigen or by cytochrome c peptide 11-25 derivatized with a 2-nitrophenylsulfenyl group. Several other peptide antigens tested failed to inhibit binding of the radioactive peptide. This suggests that a specific amino acid sequence, modified by a DNAP group, is the antigenic structure recognized by the putative T-cell receptor. In addition, direct interaction of DNAP-66-80 peptide with the hybridoma cell line induced production of the T-cell growth factor interleukin 2. Furthermore, supernatants derived from syngeneic macrophages pulsed with the relevant peptide also induced the antigen-specific hybridoma to produce interleukin 2. Images PMID:6192442

  6. T cell binding to B lymphoid cell lines in humans: a marker for T-B cell interaction?

    PubMed

    Goust, J M; Fudenberg, H H

    1983-04-15

    Binding of human circulating T cells to established normal and malignant B cell lines results in rosette formation. The percentage of B cells, circulating T cells, and thymocytes able to bind to the B-LCL Raji were 0%, 59 +/- 4% and 61 +/- 6%, respectively. The percentage of rosettes formed between Raji cells and circulating mononuclear cells from 92 normal individuals was 27.8 +/- 5.3%, and remained stable over several months. This phenomenon seems to involve relatively mature B cells, and a T cell marker which appears early in T cell ontogeny. In the peripheral blood, most of the B-LCL binding T cells exhibit a 'helper-inducer' phenotype, as determined with the monoclonal antibodies Leu 3a and OKT4. However, a significant percentage of T cells with so-called 'cytotoxic-suppressor' markers (Leu 2a and OKT8) also bind to B-LCL. The T cells involved in this morphological interactive reaction with B cells might conceivably be specifically involved in regulating B cell functions. Enumeration of this particular subset may be useful in conditions where abnormal T-B cell interactions are suspected. PMID:6601166

  7. Testing for HLA/peptide tetramer-binding to the T cell receptor complex on human T lymphocytes.

    PubMed

    Weder, Pauline; Schumacher, Ton N M; Spits, Hergen; Luiten, Rosalie M

    2012-01-01

    HLA/peptide tetramers are frequently used for ex vivo monitoring of disease- or vaccine-induced T cell immune responses and for T cell epitope identification. However, when low-levels HLA/peptide tetramer-positive T cell populations are encountered, it is difficult to ascertain whether this represents a true T cell receptor (TCR)-mediated interaction or background signal. To address this issue, we have developed a method for both HLA class I and class II tetramer assays to confirm tetramer-binding to the TCR/CD3 complex. Preincubation of T cells with anti-CD3 mAb SPV-T3b and subsequent crosslinking interferes with the binding of HLA/peptide tetramers to the TCR/CD3 complex and thereby indicates to what extent HLA/peptide tetramer binds through interaction with TCR/CD3 complex. SPV-T3b pretreatment results in a 2- to 10-fold decrease in tetramer-binding intensity to antigen-specific CD8+ or CD4+ T cells, whereas background reactivity of HLA/peptide tetramers containing HIV-derived peptide in HIV-negative donors remained unchanged. SPV-T3b pretreatment forms a valuable tool to verify tetramer-based detection of antigen-specific T cells during the monitoring of immune responses in clinical studies. PMID:24371571

  8. T Cell Immunoglobulin Mucin-3 Crystal Structure Reveals a Galectin-9-Independent Ligand-Binding Surface

    SciTech Connect

    Cao,E.; Zang, X.; Ramagopal, U.; Mukhopadhaya, A.; Fedorov, A.; Fedorov, E.; Zencheck, W.; Lary, J.; Cole, J.; et al.

    2007-01-01

    The T cell immunoglobulin mucin (Tim) family of receptors regulates effector CD4+ T cell functions and is implicated in autoimmune and allergic diseases. Tim-3 induces immunological tolerance, and engagement of the Tim-3 immunoglobulin variable (IgV) domain by galectin-9 is important for appropriate termination of T helper 1-immune responses. The 2 {angstrom} crystal structure of the Tim-3 IgV domain demonstrated that four cysteines, which are invariant within the Tim family, form two noncanonical disulfide bonds, resulting in a surface not present in other immunoglobulin superfamily members. Biochemical and biophysical studies demonstrated that this unique structural feature mediates a previously unidentified galectin-9-independent binding process and suggested that this structural feature is conserved within the entire Tim family. The current work provided a graphic example of the relationship between sequence, structure, and function and suggested that the interplay between multiple Tim-3-binding activities contributes to the regulated assembly of signaling complexes required for effective Th1-mediated immunity.

  9. Phase separation of signaling molecules promotes T cell receptor signal transduction.

    PubMed

    Su, Xiaolei; Ditlev, Jonathon A; Hui, Enfu; Xing, Wenmin; Banjade, Sudeep; Okrut, Julia; King, David S; Taunton, Jack; Rosen, Michael K; Vale, Ronald D

    2016-04-29

    Activation of various cell surface receptors triggers the reorganization of downstream signaling molecules into micrometer- or submicrometer-sized clusters. However, the functional consequences of such clustering have been unclear. We biochemically reconstituted a 12-component signaling pathway on model membranes, beginning with T cell receptor (TCR) activation and ending with actin assembly. When TCR phosphorylation was triggered, downstream signaling proteins spontaneously separated into liquid-like clusters that promoted signaling outputs both in vitro and in human Jurkat T cells. Reconstituted clusters were enriched in kinases but excluded phosphatases and enhanced actin filament assembly by recruiting and organizing actin regulators. These results demonstrate that protein phase separation can create a distinct physical and biochemical compartment that facilitates signaling. PMID:27056844

  10. ETS1 transactivates the human GM-CSF promoter in Jurkat T cells stimulated with PMA and ionomycin.

    PubMed

    Thomas, R S; Tymms, M J; Seth, A; Shannon, M F; Kola, I

    1995-11-16

    Activation of T helper cells results in coordinate expression of a number of cytokines involved in differentiation, proliferation and activation of the haematopoietic system. Granulocyte-macrophage colony-stimulating factor (GM-CSF) is one such cytokine whose increased expression results partly from increases in transcription. Cis-acting elements with NF kappa B, AP-1 and ETS-like motifs have been identified in the promoter region of the GM-CSF gene, which are important for transcriptional activity following PMA and ionomycin stimulation. A number of the ETS family of transcription factors are expressed in T cells, including ETS1 and ELF1. Here we describe the ability of these factors to interact with a site (GM5), located within the CLE0 element, -47 to -40 upstream of the GM-CSF transcription initiation site. Exogenous ETS1, but not ELF1, can transactivate GM-CSF, through the GM5 site, in a PMA/ionomycin dependent manner. Other unidentified ETS-like factors present in Jurkat cells are also capable of binding GM5. Mutation of the core ETS binding site from -GGAA- to -GGAT- prevents the binding of ETS-like factors with the exception of ETS1. The GM-CSF promoter, modified in this way to be ETS1 specific, is fully responsive to PMA/ionomycin induction, in addition to ETS1 transactivation in the presence of PMA and ionomycin. Together these data suggest that ETS1 may be involved in mediating the increased GM-CSF production associated with T cell activation. PMID:7478534

  11. Galectin-8 promotes regulatory T cell differentiation by modulating IL-2 and TGFβ signaling

    PubMed Central

    Sampson, James F.; Suryawanshi, Amol; Chen, Wei-Sheng; Rabinovich, Gabriel A.; Panjwani, Noorjahan

    2015-01-01

    Galectins have emerged as potent immunoregulatory molecules that control chronic inflammation through distinct mechanisms. Galectin-8 (Gal-8), a tandem-repeat type galectin with unique preference for α2,3-sialylated glycans, is ubiquitously expressed, but little is known about its role in T cell differentiation. Here, we report that Gal-8 promotes the polyclonal differentiation of primary mouse Treg cells in vitro. We further show that Gal-8 also facilitates antigen-specific differentiation of regulatory T (Treg) cells, and that Treg cells polarized in the presence of Gal-8 express cytotoxic T lymphocyte antigen-4 (CTLA-4) and IL-10 at a higher frequency than control Treg cells, and efficiently inhibit proliferation of activated T cells in vitro. Investigation of the mechanism by which Gal-8 promotes Treg conversion revealed that Gal-8 activates TGFβ signaling and promotes sustained IL-2R signaling. Taken together, these data suggest that Gal-8 promotes the differentiation of highly suppressive Treg cells, which has implications for the treatment of inflammatory and autoimmune diseases. PMID:26282995

  12. 1,25-Dihydroxyvitamin D3 and its analog TX527 promote a stable regulatory T cell phenotype in T cells from type 1 diabetes patients.

    PubMed

    Van Belle, Tom L; Vanherwegen, An-Sofie; Feyaerts, Dorien; De Clercq, Pierre; Verstuyf, Annemieke; Korf, Hannelie; Gysemans, Conny; Mathieu, Chantal

    2014-01-01

    The emergence of regulatory T cells (Tregs) as central mediators of peripheral tolerance in the immune system has led to an important area of clinical investigation to target these cells for the treatment of autoimmune diseases such as type 1 diabetes. We have demonstrated earlier that in vitro treatment of T cells from healthy individuals with TX527, a low-calcemic analog of bioactive vitamin D, can promote a CD4+ CD25high CD127low regulatory profile and imprint a migratory signature specific for homing to sites of inflammation. Towards clinical application of vitamin D-induced Tregs in autologous adoptive immunotherapy for type 1 diabetes, we show here that 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] and TX527 similarly imprint T cells from type 1 diabetes patients with a CD4+ CD25high CD127low regulatory profile, modulate surface expression of skin- and inflammation-homing receptors, and increase expression of CTLA-4 and OX-40. Also, 1,25(OH)2D3 and TX527 treatment inhibit the production of effector cytokines IFN-γ, IL-9, and IL-17. Importantly, 1,25(OH)2D3 and TX527 promote the induction of IL-10-producing CD4+ CD25high CD127low T cells with a stable phenotype and the functional capacity to suppress proliferation of autologous responder T cells in vitro. These findings warrant additional validation of vitamin D-induced Tregs in view of future autologous adoptive immunotherapy in type 1 diabetes. PMID:25279717

  13. 1,25-Dihydroxyvitamin D3 and Its Analog TX527 Promote a Stable Regulatory T Cell Phenotype in T Cells from Type 1 Diabetes Patients

    PubMed Central

    De Clercq, Pierre; Verstuyf, Annemieke; Korf, Hannelie; Gysemans, Conny; Mathieu, Chantal

    2014-01-01

    The emergence of regulatory T cells (Tregs) as central mediators of peripheral tolerance in the immune system has led to an important area of clinical investigation to target these cells for the treatment of autoimmune diseases such as type 1 diabetes. We have demonstrated earlier that in vitro treatment of T cells from healthy individuals with TX527, a low-calcemic analog of bioactive vitamin D, can promote a CD4+CD25highCD127low regulatory profile and imprint a migratory signature specific for homing to sites of inflammation. Towards clinical application of vitamin D-induced Tregs in autologous adoptive immunotherapy for type 1 diabetes, we show here that 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] and TX527 similarly imprint T cells from type 1 diabetes patients with a CD4+CD25highCD127low regulatory profile, modulate surface expression of skin- and inflammation-homing receptors, and increase expression of CTLA-4 and OX-40. Also, 1,25(OH)2D3 and TX527 treatment inhibit the production of effector cytokines IFN-γ, IL-9, and IL-17. Importantly, 1,25(OH)2D3 and TX527 promote the induction of IL-10-producing CD4+CD25highCD127low T cells with a stable phenotype and the functional capacity to suppress proliferation of autologous responder T cells in vitro. These findings warrant additional validation of vitamin D-induced Tregs in view of future autologous adoptive immunotherapy in type 1 diabetes. PMID:25279717

  14. Class I T-cell epitope prediction: improvements using a combination of proteasome cleavage, TAP affinity, and MHC binding.

    PubMed

    Doytchinova, Irini A; Flower, Darren R

    2006-05-01

    Cleavage by the proteasome is responsible for generating the C terminus of T-cell epitopes. Modeling the process of proteasome cleavage as part of a multi-step algorithm for T-cell epitope prediction will reduce the number of non-binders and increase the overall accuracy of the predictive algorithm. Quantitative matrix-based models for prediction of the proteasome cleavage sites in a protein were developed using a training set of 489 naturally processed T-cell epitopes (nonamer peptides) associated with HLA-A and HLA-B molecules. The models were validated using an external test set of 227 T-cell epitopes. The performance of the models was good, identifying 76% of the C-termini correctly. The best model of proteasome cleavage was incorporated as the first step in a three-step algorithm for T-cell epitope prediction, where subsequent steps predicted TAP affinity and MHC binding using previously derived models. PMID:16524630

  15. Mcl-1 antagonizes Bax/Bak to promote effector CD4+ and CD8+ T-cell responses

    PubMed Central

    Tripathi, P; Koss, B; Opferman, J T; Hildeman, D A

    2013-01-01

    Members of the Bcl-2 family have critical roles in regulating tissue homeostasis by modulating apoptosis. Anti-apoptotic molecules physically interact and restrain pro-apoptotic family members preventing the induction of cell death. However, the specificity of the functional interactions between pro- and anti-apoptotic Bcl-2 family members remains unclear. The pro-apoptotic Bcl-2 family member Bcl-2 interacting mediator of death (Bim) has a critical role in promoting the death of activated, effector T cells following viral infections. Although Bcl-2 is an important Bim antagonist in effector T cells, and Bcl-xL is not required for effector T-cell survival, the roles of other anti-apoptotic Bcl-2 family members remain unclear. Here, we investigated the role of myeloid cell leukemia sequence 1 (Mcl-1) in regulating effector T-cell responses in vivo. We found, at the peak of the response to lymphocytic choriomeningitis virus (LCMV) infection, that Mcl-1 expression was increased in activated CD4+ and CD8+ T cells. Retroviral overexpression of Mcl-1-protected activated T cells from death, whereas deletion of Mcl-1 during the course of infection led to a massive loss of LCMV-specific CD4+ and CD8+ T cells. Interestingly, the co-deletion of Bim failed to prevent the loss of Mcl-1-deficient T cells. Furthermore, lck-driven overexpression of a Bcl-xL transgene only partially rescued Mcl-1-deficient effector T cells suggesting a lack of redundancy between the family members. In contrast, additional loss of Bax and Bak completely rescued Mcl-1-deficient effector T-cell number and function, without enhancing T-cell proliferation. These data suggest that Mcl-1 is critical for promoting effector T-cell responses, but does so by combating pro-apoptotic molecules beyond Bim. PMID:23558951

  16. Egr3 Induces a Th17 Response by Promoting the Development of γδ T Cells

    PubMed Central

    Parkinson, Rose M.; Collins, Samuel L.; Horton, Maureen R.; Powell, Jonathan D.

    2014-01-01

    The transcription factor Early Growth Response 3 (Egr3) has been shown to play an important role in negatively regulating T cell activation and promoting T cell anergy in Th1 cells. However, its role in regulating other T helper subsets has yet to be described. We sought to determine the role of Egr3 in a Th17 response using transgenic mice that overexpress Egr3 in T cells (Egr3 TG). Splenocytes from Egr3 TG mice demonstrated more robust generation of Th17 cells even under non-Th17 skewing conditions. We found that while Egr3 TG T cells were not intrinsically more likely to become Th17 cells, the environment encountered by these cells was more conducive to Th17 development. Further analysis revealed a considerable increase in the number of γδ T cells in both the peripheral lymphoid organs and mucosal tissues of Egr3 TG mice, a cell type which normally accounts for only a small fraction of peripheral lymphocytes. Consistent with this marked increase in peripheral γδ T cells, thymocytes from Egr3 TG mice also appear biased toward γδ T cell development. Coculture of these Egr3-induced γδ T cells with wildtype CD4+ T cells increases Th17 differentiation, and Egr3 TG mice are more susceptible to bleomycin-induced lung inflammation. Overall our findings strengthen the role for Egr3 in promoting γδ T cell development and show that Egr3-induced γδ T cells are both functional and capable of altering the adaptive immune response in a Th17-biased manner. Our data also demonstrates that the role played by Egr3 in T cell activation and differentiation is more complex than previously thought. PMID:24475259

  17. Hypoxia promotes Mycobacterium tuberculosis-specific up-regulation of granulysin in human T cells.

    PubMed

    Zenk, Sebastian F; Vollmer, Michael; Schercher, Esra; Kallert, Stephanie; Kubis, Jan; Stenger, Steffen

    2016-06-01

    Oxygen tension affects local immune responses in inflammation and infection. In tuberculosis mycobacteria avoid hypoxic areas and preferentially persist and reactivate in the oxygen-rich apex of the lung. Oxygen restriction activates antimicrobial effector mechanisms in macrophages and restricts growth of intracellular Mycobacterium tuberculosis (M.Tb). The effect of oxygen restriction on T cell-mediated antimicrobial effector mechanisms is unknown. Therefore we determined the influence of hypoxia on the expression of granulysin, an antimicrobial peptide of lymphocytes. Hypoxia increased the antigen-specific up-regulation of granulysin mRNA and protein in human CD4(+) and CD8(+) T lymphocytes. This observation was functionally relevant, because oxygen restriction supported the growth-limiting effect of antigen-specific T cells against virulent M.Tb residing in primary human macrophages. Our results provide evidence that oxygen restriction promotes the expression of granulysin and suggest that this effect-in conjunction with additional T cell-mediated immune responses-supports protection against mycobacteria. The therapeutic modulation of oxygen availability may offer a new strategy for the host-directed therapy of infectious diseases with intracellular pathogens. PMID:26613797

  18. Human leukocyte antigen (HLA)-binding epitopes dataset for the newly identified T-cell antigens of Mycobacterium immunogenum.

    PubMed

    Chandra, Harish; Yadav, Jagjit S

    2016-09-01

    The dataset described herein is related to our article entitled "T-cell antigens of Mycobacterium immunogenum (MI), an etiological agent of occupational hypersensitivity pneumonitis'' (Chandra and Yadav, 2016) [1]. The data include in silico-predicted T-cell epitopes of the T-cell antigens AgA and AgD of MI predicted to bind to HLA-I or HLA-II alleles. Data on two reference T-cell antigens ESAT-6 and CFP-10 of Mycobacterium tuberculosis H37Rv are included for comparison. The data for each antigen include the predicted epitope׳s amino acid sequence, its first amino acid position, and its ability to bind HLA-I or HLA-II allele(s). PMID:27508266

  19. Promotion of regulatory T cell induction by immunomodulatory herbal medicine licorice and its two constituents

    PubMed Central

    Guo, Ao; He, Dongming; Xu, Hong-Bo; Geng, Chang-An; Zhao, Jian

    2015-01-01

    Regulatory T cells (Treg) play a critical role to control immune responses and to prevent autoimmunity, thus selective increase of Treg cells in vivo has broad therapeutic implications for autoimmune and inflammatory diseases. Licorice is a well-known herbal medicine used worldwide for over thousands of years, and accumulating evidence has shown its immunomodulatory potential. However, it is not clear whether licorice could regulate the induction and function of Treg cells. Here we found licorice extract could promote Treg cell induction, and then we used a rational approach to isolate its functional fractions and constituents. The results showed that two constituents, isoliquiritigenin and naringenin, promoted Treg cell induction both in vitro and in vivo. The effective fractions and two constituents of licorice also enhanced immune suppression of Treg cells, and they further reduced severity of DSS-induced colitis in mice. This study suggested that promotion of regulatory T cell induction could be an underlying mechanism of the historically and widely used herbal medicine licorice, providing its two effective molecules against autoimmune and inflammatory diseases. PMID:26370586

  20. Impaired memory CD8 T cell development in the absence of methyl-CpG-binding domain protein 2.

    PubMed

    Kersh, Ellen N

    2006-09-15

    Intracellular differentiation events that determine which cells develop into memory CD8 T cells are currently incompletely understood. Methyl-CpG-binding domain protein 2 (MBD2) is a transcriptional repressor that binds to methylated DNA and mediates the biological consequences of epigenetic gene methylation. The role of MBD2 during the differentiation of naive CD8 T cells into effector and memory cells was determined following acute infection of MBD2-deficient mice with lymphocytic choriomeningitis virus. Despite rapid viral clearance and an efficient primary effector CD8 T cell response, reduced numbers of Ag-specific memory CD8 T cells were observed. Importantly, the appearance of precursor memory cells (IL-7Ralphahigh) was delayed. The remaining MBD2(-/-) memory cells were not fully protective during rechallenge, and memory cell characteristics were altered with regard to surface markers (IL-7Ralpha, KLRG-1, CD27, and others) and cytokine production. The defect was CD8 T cell intrinsic, because memory cell development was also delayed when MBD2(-/-) CD8 T cells were adoptively transferred into SCID mice. These data demonstrate that MBD2 is a previously unrecognized intracellular factor required for the efficient generation of protective memory CD8 T cells. PMID:16951344

  1. Stimulation of T-cells with OKT3 antibodies increases forskolin binding and cyclic AMP accumulation.

    PubMed

    Kvanta, A; Gerwins, P; Jondal, M; Fredholm, B B

    1990-01-01

    It has recently been shown that elevation of cAMP by adenosine receptor stimulation may be potentiated by stimulation of the T-cell receptor/CD3 complex on human T-cells with the monoclonal antibody OKT3, and that this is mimicked by activation of protein kinase C [Kvanta, A. et al. (1989) Naunyn-Schmeideberg's Arch. Pharmac. 340, 715-717]. In this study the diterpene forskolin, which binds to and activates the adenylate cyclase, has been used to examine further how the CD3 complex may influence the adenylate cyclase pathway. Stimulation with OKT3 alone was found to cause a small dose-dependent increase in basal cAMP accumulation. When combining OKT3 with a concentration of forskolin (10 microM), which by itself had little effect on the cyclase activity, the cAMP accumulation was markedly potentiated. This potentiation was paralleled by an increase in [3H]forskolin binding to saponine permeabilized Jurkat cells from 24 to 41 fmol/10(6) cells. The OKT3 effect on cAMP was blocked by chelating extracellular Ca2+ with EGTA or intracellular Ca2+ with BAPTA and also by W-7, an inhibitor of calmodulin, but was unaffected by H-7, an inhibitor of protein kinase C. Even though OKT3 caused an increase in inositolphosphate turnover, and activated protein kinase C, neither phorbol 12,13 dibutyrate (PDBu) nor the Ca2(+)-ionophore A23187 could mimic the OKT3 effect, whereas a combination of PDBu and A23187 at high concentrations could potentiate forskolin stimulated cyclase activity. Together, these results indicated that stimulation of the CD3 complex could influence the adenylate cyclase by two different mechanisms, one involving activation of protein kinase C and another which does not. PMID:2177619

  2. Kinetics of T-cell receptor binding to peptide/I-Ek complexes: correlation of the dissociation rate with T-cell responsiveness.

    PubMed Central

    Matsui, K; Boniface, J J; Steffner, P; Reay, P A; Davis, M M

    1994-01-01

    Recognition by T-cell antigen receptors (TCRs) of processed peptides bound to major histocompatibility complex (MHC) molecules is required for the initiation of most T-lymphocyte responses. Despite the availability of soluble forms of TCRs and MHC heterodimers, this interaction has proven difficult to study directly due to the very low affinity. We report here on the kinetics of TCR binding to peptide/MHC complexes in a cell-free system using surface plasmon resonance. The apparent association rates for the interactions of related peptide/MHC complexes to one such TCR are relatively slow (900-3000 M-1.s-1) and dissociation rates are very fast (0.3-0.06 s-1) with t1/2 of 2-12 s at 25 degrees C. The calculated affinity of the engineered soluble molecules compares well with previously reported competition data for native TCRs or competition data reported here for native peptide/MHC complexes, indicating that these soluble heterodimers bind in the same manner as the original molecules expressed on cells. We also find that the peptide variants which give weaker T-cell stimulatory responses have similar affinities but distinctly faster dissociation rates compared with the original peptide (when loaded onto the MHC molecule) and that this later property may be responsible for their lower activity. This has implications for both downstream signaling events and models of TCR-peptide antagonists. PMID:7809136

  3. Activation‐Induced Killer Cell Immunoglobulin‐like Receptor 3DL2 Binding to HLA–B27 Licenses Pathogenic T Cell Differentiation in Spondyloarthritis

    PubMed Central

    Ridley, Anna; Hatano, Hiroko; Wong‐Baeza, Isabel; Shaw, Jacqueline; Matthews, Katherine K.; Al‐Mossawi, Hussein; Ladell, Kristin; Price, David A.; Bowness, Paul

    2016-01-01

    Objective In the spondyloarthritides (SpA), increased numbers of CD4+ T cells express killer cell immunoglobulin‐like receptor 3DL2 (KIR‐3DL2). The aim of this study was to determine the factors that induce KIR‐3DL2 expression, and to characterize the relationship between HLA–B27 and the phenotype and function of KIR‐3DL2–expressing CD4+ T cells in SpA. Methods In total, 34 B27+ patients with SpA, 28 age‐ and sex‐matched healthy controls (20 B27− and 8 B27+), and 9 patients with rheumatoid arthritis were studied. KIR-3DL2 expression and other phenotypic characteristics of peripheral blood and synovial fluid CD4+ T cells were studied by flow cytometry, quantitative polymerase chain reaction, and Western blotting. T cell receptor clonality was determined by template‐switch anchored reverse transcription–polymerase chain reaction and sequencing analysis. Cytokines were measured by enzyme‐linked immunosorbent assay. Results Cellular activation induced KIR‐3DL2 expression on both naive and effector CD4+ T cells. KIR‐3DL2 binding to B27+ cells promoted expression of KIR‐3DL2, the Th17‐specific transcription factor retinoic acid receptor–related orphan nuclear receptor γt, and the antiapoptotic factor B cell lymphoma 2. KIR‐3DL2+CD4+ T cells in patients with ankylosing spondylitis were oligoclonal and enriched for markers of T cell activation and for the gut homing receptor CCR9. In the presence of B27+ antigen‐presenting cells, KIR‐3DL2+CD4+ T cells produced less interleukin‐2 (IL‐2) but more IL‐17. This effect was blocked by HC10, an antibody that inhibits the binding of KIR‐3DL2 to B27 heavy chains. Conclusion KIR‐3DL2 binding to HLA–B27 licenses Th17 cell differentiation in SpA. These findings raise the therapeutic potential of targeting HLA–B27–KIR‐3DL2 interactions for the treatment of B27+ patients with SpA. PMID:26841353

  4. Defect density in multiwalled carbon nanotubes influences ovalbumin adsorption and promotes macrophage activation and CD4(+) T-cell proliferation.

    PubMed

    Bai, Wei; Raghavendra, Achyut; Podila, Ramakrishna; Brown, Jared M

    2016-01-01

    Carbon nanotubes (CNTs) are of great interest for the development of drugs and vaccines due to their unique physicochemical properties. The high surface area to volume ratio and delocalized pi-electron cloud of CNTs promote binding of proteins to the surface forming a protein corona. This unique feature of CNTs has been recognized for potential delivery of antigens for strong and long-lasting antigen-specific immune responses. Based on an earlier study that demonstrated increased protein binding, we propose that carboxylated multiwalled CNTs (MWCNTs) can function as an improved carrier to deliver antigens such as ovalbumin (OVA). To test this hypothesis, we coated carboxylated MWCNTs with OVA and measured uptake and activation of antigen-presenting cells (macrophages) and their ability to stimulate CD4(+) T-cell proliferation. We employed two types of carboxylated MWCNTs with different surface areas and defects (MWCNT-2 and MWCNT-30). MWCNT-2 and MWCNT-30 have surface areas of ~215 m(2)/g and 94 m(2)/g, respectively. The ratios of D- to G-band areas (I D/I G) were 0.97 and 1.37 for MWCNT-2 and MWCNT-30, respectively, samples showing that MWCNT-30 contained more defects. The increase in defects in MWCNT-30 led to increased binding of OVA as compared to MWCNT-2 (1,066±182 μg/mL vs 582±41 μg/mL, respectively). Both types of MWCNTs, along with MWCNT-OVA complexes, showed no observable toxicity to bone-marrow-derived macrophages up to 5 days. Surprisingly, we found that MWCNT-OVA complex significantly increased the expression of major histocompatibility complex class II on macrophages and production of pro-inflammatory cytokines (tumor necrosis factor-α and interleukin 6), while MWCNTs without OVA protein corona did not. The coculture of MWCNT-OVA-complex-treated macrophages and OVA-specific CD4(+) T-cells isolated from OT-II mice demonstrated robust proliferation of CD4(+) T-cells. This study provides strong evidence for a role for defects in carboxylated MWCNTs

  5. Defect density in multiwalled carbon nanotubes influences ovalbumin adsorption and promotes macrophage activation and CD4+ T-cell proliferation

    PubMed Central

    Bai, Wei; Raghavendra, Achyut; Podila, Ramakrishna; Brown, Jared M

    2016-01-01

    Carbon nanotubes (CNTs) are of great interest for the development of drugs and vaccines due to their unique physicochemical properties. The high surface area to volume ratio and delocalized pi-electron cloud of CNTs promote binding of proteins to the surface forming a protein corona. This unique feature of CNTs has been recognized for potential delivery of antigens for strong and long-lasting antigen-specific immune responses. Based on an earlier study that demonstrated increased protein binding, we propose that carboxylated multiwalled CNTs (MWCNTs) can function as an improved carrier to deliver antigens such as ovalbumin (OVA). To test this hypothesis, we coated carboxylated MWCNTs with OVA and measured uptake and activation of antigen-presenting cells (macrophages) and their ability to stimulate CD4+ T-cell proliferation. We employed two types of carboxylated MWCNTs with different surface areas and defects (MWCNT-2 and MWCNT-30). MWCNT-2 and MWCNT-30 have surface areas of ~215 m2/g and 94 m2/g, respectively. The ratios of D- to G-band areas (ID/IG) were 0.97 and 1.37 for MWCNT-2 and MWCNT-30, respectively, samples showing that MWCNT-30 contained more defects. The increase in defects in MWCNT-30 led to increased binding of OVA as compared to MWCNT-2 (1,066±182 μg/mL vs 582±41 μg/mL, respectively). Both types of MWCNTs, along with MWCNT–OVA complexes, showed no observable toxicity to bone-marrow-derived macrophages up to 5 days. Surprisingly, we found that MWCNT–OVA complex significantly increased the expression of major histocompatibility complex class II on macrophages and production of pro-inflammatory cytokines (tumor necrosis factor-α and interleukin 6), while MWCNTs without OVA protein corona did not. The coculture of MWCNT–OVA-complex-treated macrophages and OVA-specific CD4+ T-cells isolated from OT-II mice demonstrated robust proliferation of CD4+ T-cells. This study provides strong evidence for a role for defects in carboxylated MWCNTs and

  6. Lung-resident tissue macrophages generate Foxp3+ regulatory T cells and promote airway tolerance.

    PubMed

    Soroosh, Pejman; Doherty, Taylor A; Duan, Wei; Mehta, Amit Kumar; Choi, Heonsik; Adams, Yan Fei; Mikulski, Zbigniew; Khorram, Naseem; Rosenthal, Peter; Broide, David H; Croft, Michael

    2013-04-01

    Airway tolerance is the usual outcome of inhalation of harmless antigens. Although T cell deletion and anergy are likely components of tolerogenic mechanisms in the lung, increasing evidence indicates that antigen-specific regulatory T cells (inducible Treg cells [iTreg cells]) that express Foxp3 are also critical. Several lung antigen-presenting cells have been suggested to contribute to tolerance, including alveolar macrophages (MØs), classical dendritic cells (DCs), and plasmacytoid DCs, but whether these possess the attributes required to directly promote the development of Foxp3(+) iTreg cells is unclear. Here, we show that lung-resident tissue MØs coexpress TGF-β and retinal dehydrogenases (RALDH1 and RALDH 2) under steady-state conditions and that their sampling of harmless airborne antigen and presentation to antigen-specific CD4 T cells resulted in the generation of Foxp3(+) Treg cells. Treg cell induction in this model depended on both TGF-β and retinoic acid. Transfer of the antigen-pulsed tissue MØs into the airways correspondingly prevented the development of asthmatic lung inflammation upon subsequent challenge with antigen. Moreover, exposure of lung tissue MØs to allergens suppressed their ability to generate iTreg cells coincident with blocking airway tolerance. Suppression of Treg cell generation required proteases and TLR-mediated signals. Therefore, lung-resident tissue MØs have regulatory functions, and strategies to target these cells might hold promise for prevention or treatment of allergic asthma. PMID:23547101

  7. V{delta}1 T cell receptor binds specifically to MHC I chain related A: Molecular and biochemical evidences

    SciTech Connect

    Zhao Jianqing; Huang Jie; Chen Hui; Cui Lianxian; He Wei . E-mail: heweiimu@public.bta.net.cn

    2006-01-06

    Human MHC class I chain-related A (MICA) is a tumor-associated antigen that can be recognized by V{delta}1 subset of tumor-infiltrating {gamma}{delta} T cells. We previously reported that immobilized recombinant MICA protein could induce the proliferation of tumor-infiltrating V{delta}1 {gamma}{delta} T cells in vitro. But there has been no direct evidence showing the engagement of {gamma}{delta} T cell receptors (TCR) of the induced cells with MICA. In the current investigation, we show that MICA induces specific cytolytic activity of the expanded {gamma}{delta} T cells. We expressed the coupled V domains from the MICA-induced T cells as a single polypeptide chain V{delta}V{gamma} TCR ({gamma}{delta} scTCR). Such scTCR can specifically bind MICA of HeLa cells. Direct interaction of {gamma}{delta} scTCRs with in vitro expressed MICA was monitored using an IAsys biosensor. We found that the V{delta}1 scTCR can specifically bind to immobilized MICA molecule and MICA{alpha}1{alpha}2 domains are responsible for the binding reaction.

  8. A miRNA upregulated in asthma airway T cells promotes TH2 cytokine production

    PubMed Central

    Simpson, Laura J.; Patel, Sana; Bhakta, Nirav R.; Choy, David F.; Brightbill, Hans D.; Ren, Xin; Wang, Yanli; Pua, Heather H.; Baumjohann, Dirk; Montoya, Misty M.; Panduro, Marisella; Remedios, Kelly A.; Huang, Xiaozhu; Fahy, John V.; Arron, Joseph R.; Woodruff, Prescott G.; Ansel., Karl M.

    2014-01-01

    MicroRNAs (miRNAs) exert powerful effects on immune function by tuning networks of target genes that orchestrate cell behavior. We sought to uncover miRNAs and miRNA-regulated pathways that control the TH2 responses that drive pathogenic inflammation in asthma. Profiling miRNA expression in human airway-infiltrating T cells revealed miR-19a elevation in asthma. Modulating miR-19 activity altered TH2 cytokine production in both human and mouse T cells, and TH2 cell responses were markedly impaired in cells lacking the entire miR-17∼92 cluster. miR-19 promotes TH2 cytokine production and amplifies PI(3)K, JAK-STAT, and NF-κB signaling by direct targeting of PTEN, SOCS1, and A20. Thus, miR-19a up regulation in asthma may be an indicator and a cause of increased TH2 cytokine production in the airways. PMID:25362490

  9. MicroRNA-26a Promotes Regulatory T cells and Suppresses Autoimmune Diabetes in Mice.

    PubMed

    Ma, Hui; Zhang, Shoutao; Shi, Doufei; Mao, Yanhua; Cui, Jianguo

    2016-02-01

    Type-1 diabetes (TID) is an autoimmune disease in which the body's own immune cells attack islet β cells, the cells in the pancreas that produce and release the hormone insulin. Mir-26a has been reported to play functions in cellular differentiation, cell growth, cell apoptosis, and metastasis. However, the role of microRNA-26a (Mir-26a) in autoimmune TID has never been investigated. In our current study, we found that pre-Mir-26a (LV-26a)-treated mice had significantly longer normoglycemic time and lower frequency of autoreactive IFN-γ-producing CD4(+) cells compared with an empty lentiviral vector (LV-Con)-treated non-obese diabetic (NOD) mice. Mir-26a suppresses autoreactive T cells and expands Tregs in vivo and in vitro. Furthermore, in our adoptive transfer study, the groups receiving whole splenocytes and CD25-depleted splenocytes from LV-Con-treated diabetic NOD mice develop diabetes at 3 to 4 weeks of age. In comparison, mice injected with undepleted splenocytes obtained from LV-26a-treated reversal NOD mice develop diabetes after 6-8 weeks. And depletion of CD25(+) cells in the splenocytes of reversed mice abrogates the delay in diabetes onset. In conclusion, Mir-26a suppresses autoimmune diabetes in NOD mice in part through promoted regulatory T cells (Tregs) expression. PMID:26208605

  10. CD4-Negative Cells Bind Human Immunodeficiency Virus Type 1 and Efficiently Transfer Virus to T Cells

    PubMed Central

    Olinger, Gene G.; Saifuddin, Mohammed; Spear, Gregory T.

    2000-01-01

    The ability of human immunodeficiency virus strain MN (HIVMN), a T-cell line-adapted strain of HIV, and X4 and R5 primary isolates to bind to various cell types was investigated. In general, HIVMN bound to cells at higher levels than did the primary isolates. Virus bound to both CD4-positive (CD4+) and CD4-negative (CD4−) cells, including neutrophils, Raji cells, tonsil mononuclear cells, erythrocytes, platelets, and peripheral blood mononuclear cells (PBMC), although virus bound at significantly higher levels to PBMC. However, there was no difference in the amount of HIV that bound to CD4-enriched or CD4-depleted PBMC. Virus bound to CD4− cells was up to 17 times more infectious for T cells in cocultures than was the same amount of cell-free virus. Virus bound to nucleated cells was significantly more infectious than virus bound to erythrocytes or platelets. The enhanced infection of T cells by virus bound to CD4− cells was not due to stimulatory signals provided by CD4− cells or infection of CD4− cells. However, anti-CD18 antibody substantially reduced the enhanced virus replication in T cells, suggesting that virus that bound to the surface of CD4− cells is efficiently passed to CD4+ T cells during cell-cell adhesion. These studies show that HIV binds at relatively high levels to CD4− cells and, once bound, is highly infectious for T cells. This suggests that virus binding to the surface of CD4− cells is an important route for infection of T cells in vivo. PMID:10954556

  11. Binding of Soluble Natural Ligands to a Soluble Human T-Cell Receptor Fragment Produced in Escherichia coli

    NASA Astrophysics Data System (ADS)

    Hilyard, Katherine L.; Reyburn, Hugh; Chung, Shan; Bell, John I.; Strominger, Jack L.

    1994-09-01

    An Escherichia coli expression system has been developed to produce milligram quantities of the variable domains of a human T-cell receptor from a cytotoxic T cell that recognizes the HLA-A2-influenza matrix peptide complex as a single polypeptide chain. The recombinant protein was purified by metal-chelate chromatography and then refolded in a redox buffer system. The refolded protein was shown to directly bind both Staphylococcus aureus enterotoxin B and the major histocompatibility complex protein-peptide complex using a BIAcore biosensor. Thus this preparation of a single-chain, variable-domain, T-cell receptor fragment can bind both of its natural ligands and some of it is therefore a functional fragment of the receptor molecule.

  12. Transcriptional repressor ZEB2 promotes terminal differentiation of CD8+ effector and memory T cell populations during infection

    PubMed Central

    Omilusik, Kyla D.; Best, J. Adam; Yu, Bingfei; Goossens, Steven; Weidemann, Alexander; Nguyen, Jessica V.; Seuntjens, Eve; Stryjewska, Agata; Zweier, Christiane; Roychoudhuri, Rahul; Gattinoni, Luca; Bird, Lynne M.; Higashi, Yujiro; Kondoh, Hisato; Huylebroeck, Danny; Haigh, Jody

    2015-01-01

    ZEB2 is a multi-zinc-finger transcription factor known to play a significant role in early neurogenesis and in epithelial-mesenchymal transition–dependent tumor metastasis. Although the function of ZEB2 in T lymphocytes is unknown, activity of the closely related family member ZEB1 has been implicated in lymphocyte development. Here, we find that ZEB2 expression is up-regulated by activated T cells, specifically in the KLRG1hi effector CD8+ T cell subset. Loss of ZEB2 expression results in a significant loss of antigen-specific CD8+ T cells after primary and secondary infection with a severe impairment in the generation of the KLRG1hi effector memory cell population. We show that ZEB2, which can bind DNA at tandem, consensus E-box sites, regulates gene expression of several E-protein targets and may directly repress Il7r and Il2 in CD8+ T cells responding to infection. Furthermore, we find that T-bet binds to highly conserved T-box sites in the Zeb2 gene and that T-bet and ZEB2 regulate similar gene expression programs in effector T cells, suggesting that T-bet acts upstream and through regulation of ZEB2. Collectively, we place ZEB2 in a larger transcriptional network that is responsible for the balance between terminal differentiation and formation of memory CD8+ T cells. PMID:26503445

  13. PKCθ and HIV-1 Transcriptional Regulator Tat Co-exist at the LTR Promoter in CD4+ T Cells

    PubMed Central

    López-Huertas, María Rosa; Li, Jasmine; Zafar, Anjum; Rodríguez-Mora, Sara; García-Domínguez, Carlota; Mateos, Elena; Alcamí, José; Rao, Sudha; Coiras, Mayte

    2016-01-01

    PKCθ is essential for the activation of CD4+ T cells. Upon TCR/CD28 stimulation, PKCθ is phosphorylated and migrates to the immunological synapse, inducing the activation of cellular transcription factors such as NF-κB and kinases as ERK that are critical for HIV-1 replication. We previously demonstrated that PKCθ is also necessary for HIV-1 replication but the precise mechanism is unknown. Efficient HIV-1 transcription and elongation are absolutely dependent on the synergy between NF-κB and the viral regulator Tat. Tat exerts its function by binding a RNA stem-loop structure proximal to the viral mRNA cap site termed TAR. Besides, due to its effect on cellular metabolic pathways, Tat causes profound changes in infected CD4+ T cells such as the activation of NF-κB and ERK. We hypothesized that the aberrant upregulation of Tat-mediated activation of NF-κB and ERK occurred through PKCθ signaling. In fact, Jurkat TetOff cells with stable and doxycycline-repressible expression of Tat (Jurkat-Tat) expressed high levels of mRNA for PKCθ. In these cells, PKCθ located at the plasma membrane was phosphorylated at T538 residue in undivided cells, in the absence of stimulation. Treatment with doxycycline inhibited PKCθ phosphorylation in Jurkat-Tat, suggesting that Tat expression was directly related to the activation of PKCθ. Both NF-κB and Ras/Raf/MEK/ERK signaling pathway were significantly activated in Jurkat-Tat cells, and this correlated with high transactivation of HIV-1 LTR promoter. RNA interference for PKCθ inhibited NF-κB and ERK activity, as well as LTR-mediated transactivation even in the presence of Tat. In addition to Tat-mediated activation of PKCθ in the cytosol, we demonstrated by sequential ChIP that Tat and PKCθ coexisted in the same complex bound at the HIV-1 LTR promoter, specifically at the region containing TAR loop. In conclusion, PKCθ-Tat interaction seemed to be essential for HIV-1 replication in CD4+ T cells and could be used as a

  14. Natural killer cell activation enhances immune pathology and promotes chronic infection by limiting CD8+ T-cell immunity.

    PubMed

    Lang, Philipp A; Lang, Karl S; Xu, Haifeng C; Grusdat, Melanie; Parish, Ian A; Recher, Mike; Elford, Alisha R; Dhanji, Salim; Shaabani, Namir; Tran, Charles W; Dissanayake, Dilan; Rahbar, Ramtin; Ghazarian, Magar; Brüstle, Anne; Fine, Jason; Chen, Peter; Weaver, Casey T; Klose, Christoph; Diefenbach, Andreas; Häussinger, Dieter; Carlyle, James R; Kaech, Susan M; Mak, Tak W; Ohashi, Pamela S

    2012-01-24

    Infections with HIV, hepatitis B virus, and hepatitis C virus can turn into chronic infections, which currently affect more than 500 million patients worldwide. It is generally thought that virus-mediated T-cell exhaustion limits T-cell function, thus promoting chronic disease. Here we demonstrate that natural killer (NK) cells have a negative impact on the development of T-cell immunity by using the murine lymphocytic choriomeningitis virus. NK cell-deficient (Nfil3(-/-), E4BP4(-/-)) mice exhibited a higher virus-specific T-cell response. In addition, NK cell depletion caused enhanced T-cell immunity in WT mice, which led to rapid virus control and prevented chronic infection in lymphocytic choriomeningitis virus clone 13- and reduced viral load in DOCILE-infected animals. Further experiments showed that NKG2D triggered regulatory NK cell functions, which were mediated by perforin, and limited T-cell responses. Therefore, we identified an important role of regulatory NK cells in limiting T-cell immunity during virus infection. PMID:22167808

  15. Peptidoglycan-linked protein A promotes T cell-dependent antibody expansion during Staphylococcus aureus infection

    PubMed Central

    Kim, Hwan Keun; Falugi, Fabiana; Missiakas, Dominique M.; Schneewind, Olaf

    2016-01-01

    A hallmark of Staphylococcus aureus disease in humans is persistent infections without development of protective immune responses. Infected patients generate VH3 plasmablast expansions and increased VH3 idiotype Ig; however, the mechanisms for staphylococcal modification of immune responses are not known. We report here that S. aureus-infected mice generate VH3 antibody expansions via a mechanism requiring MHC-restricted antigen presentation to CD4+ T cells and staphylococcal protein A (SpA), a cell wall-anchored surface molecule that binds Fcγ and VH3 variant heavy chains of Ig. VH3 expansion occurred with peptidoglycan-linked SpA from the bacterial envelope but not with recombinant SpA, and optimally required five tandem repeats of its Ig-binding domains. Signaling via receptor-interacting serine/threonine protein kinase 2 (RIPK2) was essential for implementing peptidoglycan-linked SpA superantigen activity. VH3 clan IgG from S. aureus-infected or SpA-treated animals was not pathogen-specific, suggesting that SpA cross-linking of VH3 idiotype B-cell receptors and activation via attached peptidoglycan are the determinants of staphylococcal escape from adaptive immune responses. PMID:27140614

  16. Peptidoglycan-linked protein A promotes T cell-dependent antibody expansion during Staphylococcus aureus infection.

    PubMed

    Kim, Hwan Keun; Falugi, Fabiana; Missiakas, Dominique M; Schneewind, Olaf

    2016-05-17

    A hallmark of Staphylococcus aureus disease in humans is persistent infections without development of protective immune responses. Infected patients generate VH3 plasmablast expansions and increased VH3 idiotype Ig; however, the mechanisms for staphylococcal modification of immune responses are not known. We report here that S. aureus-infected mice generate VH3 antibody expansions via a mechanism requiring MHC-restricted antigen presentation to CD4(+) T cells and staphylococcal protein A (SpA), a cell wall-anchored surface molecule that binds Fcγ and VH3 variant heavy chains of Ig. VH3 expansion occurred with peptidoglycan-linked SpA from the bacterial envelope but not with recombinant SpA, and optimally required five tandem repeats of its Ig-binding domains. Signaling via receptor-interacting serine/threonine protein kinase 2 (RIPK2) was essential for implementing peptidoglycan-linked SpA superantigen activity. VH3 clan IgG from S. aureus-infected or SpA-treated animals was not pathogen-specific, suggesting that SpA cross-linking of VH3 idiotype B-cell receptors and activation via attached peptidoglycan are the determinants of staphylococcal escape from adaptive immune responses. PMID:27140614

  17. Histamine type I (H/sub 1/) receptor radioligand binding studies on normal T cell subsets, B cells, and monocytes

    SciTech Connect

    Cameron, W.; Doyle, K.; Rocklin, R.E.

    1986-03-15

    A single, specific binding site for (/sup 3/H)pyrilamine on normal human T helper, T suppressor, B cells, and monocytes was documented. The binding of the radioligand to its receptor is reversible with cold H/sub 1/ antagonist, saturates at 40 to 60 nM, and binding equilibrium is achieved in 2 to 4 min. Using a computer program (Ligand), the authors calculated the dissociation constants, binding capacities, and numbers of receptors per cell for each of the different cell types. Monocytes were found to have the highest affinity for (/sup 3/H)pyrilamine, followed by T helper cells, B cells and T suppressor cells (K/sub D/ = 44.6 +/- 49.4 nM). T suppressor cells were found to express the higher number of H/sub 1/ receptors per cell followed by B cells, T helper cells, and monocytes. The binding affinity for (/sup 3/H)pyrilamine increased over a 48-hr period, whereas the number of receptors per T cell was essentially unchanged. In contrast, T cells stimulated with Con A or PHA were shown to have a greater than fourfold increase in the number of receptors per cell, whereas the binding affinity for (/sup 3/H)pyrilamine decreased over the 48-hr period. Although the function of H/sub 1/ receptors on T cells, B cells, and monocytes has not been completely defined, this receptor has the potential of playing an important role in the modulating the immune response.

  18. Active role of T cells in promoting an in vitro autoantibody response to self erythrocytes in NZB mice.

    PubMed Central

    Miller, R D; Calkins, C E

    1988-01-01

    The in vitro anti-self erythrocyte antibody response of NZB spleen cells appears to be influenced directly by T cells. Thy-1+, L3T4+ helper T cells are required for: (i) the generation in vitro of MRBC-specific IgM and IgG AFC by spleen cells from 'autoimmune' (9-12-month old) NZB mice and (ii) the generation in vitro of MRBC-specific IgM and IgG AFC by spleen cells depleted of suppressor cells from pre-autoimmune (2-3-months-old) NZB mice which show no clinical signs of an anti-MRBC response. It is evident from the present and previous studies that the anti-MRBC autoantibody response is regulated in pre-autoimmune spleen cell populations by Ly2+ T cells. Ly2-T cells from both pre-autoimmune and autoimmune mice in sufficient numbers can overcome this normal regulation and promote the anti-MRBC response in cultures of unfractionated pre-autoimmune spleen cells. Ly2- T cells isolated from autoimmune NZB mice were consistently more active in this than the Ly2- T cells isolated from pre-autoimmune mice, suggesting an enrichment of MRBC-reactive Ly2- T cells in autoimmune NZB mice. The Ly2- T cells from autoimmune NZB mice greatly enhance the autoimmune anti-MRBC response relative to a modest enhancement of the response to a foreign antigen, SRBC, produced by the same cells. These data indicate that T cells play an important role both in supporting the autoantibody response to MRBC and in disrupting tolerance, leading to autoimmunity in NZB mice. PMID:2966765

  19. The alarmin IL-33 promotes regulatory T-cell function in the intestine.

    PubMed

    Schiering, Chris; Krausgruber, Thomas; Chomka, Agnieszka; Fröhlich, Anja; Adelmann, Krista; Wohlfert, Elizabeth A; Pott, Johanna; Griseri, Thibault; Bollrath, Julia; Hegazy, Ahmed N; Harrison, Oliver J; Owens, Benjamin M J; Löhning, Max; Belkaid, Yasmine; Fallon, Padraic G; Powrie, Fiona

    2014-09-25

    FOXP3(+) regulatory T cells (Treg cells) are abundant in the intestine, where they prevent dysregulated inflammatory responses to self and environmental stimuli. It is now appreciated that Treg cells acquire tissue-specific adaptations that facilitate their survival and function; however, key host factors controlling the Treg response in the intestine are poorly understood. The interleukin (IL)-1 family member IL-33 is constitutively expressed in epithelial cells at barrier sites, where it functions as an endogenous danger signal, or alarmin, in response to tissue damage. Recent studies in humans have described high levels of IL-33 in inflamed lesions of inflammatory bowel disease patients, suggesting a role for this cytokine in disease pathogenesis. In the intestine, both protective and pathological roles for IL-33 have been described in murine models of acute colitis, but its contribution to chronic inflammation remains ill defined. Here we show in mice that the IL-33 receptor ST2 is preferentially expressed on colonic Treg cells, where it promotes Treg function and adaptation to the inflammatory environment. IL-33 signalling in T cells stimulates Treg responses in several ways. First, it enhances transforming growth factor (TGF)-β1-mediated differentiation of Treg cells and, second, it provides a necessary signal for Treg-cell accumulation and maintenance in inflamed tissues. Strikingly, IL-23, a key pro-inflammatory cytokine in the pathogenesis of inflammatory bowel disease, restrained Treg responses through inhibition of IL-33 responsiveness. These results demonstrate a hitherto unrecognized link between an endogenous mediator of tissue damage and a major anti-inflammatory pathway, and suggest that the balance between IL-33 and IL-23 may be a key controller of intestinal immune responses. PMID:25043027

  20. Targeting HIV Reservoir in Infected CD4 T Cells by Dual-Affinity Re-targeting Molecules (DARTs) that Bind HIV Envelope and Recruit Cytotoxic T Cells

    PubMed Central

    Sloan, Derek D.; Lam, Chia-Ying Kao; Irrinki, Alivelu; Liu, Liqin; Tsai, Angela; Pace, Craig S.; Kaur, Jasmine; Murry, Jeffrey P.; Balakrishnan, Mini; Moore, Paul A.; Johnson, Syd; Nordstrom, Jeffrey L.; Cihlar, Tomas; Koenig, Scott

    2015-01-01

    HIV reservoirs and production of viral antigens are not eliminated in chronically infected participants treated with combination antiretroviral therapy (cART). Novel therapeutic strategies aiming at viral reservoir elimination are needed to address chronic immune dysfunction and non-AIDS morbidities that exist despite effective cART. The HIV envelope protein (Env) is emerging as a highly specific viral target for therapeutic elimination of the persistent HIV-infected reservoirs via antibody-mediated cell killing. Dual-Affinity Re-Targeting (DART) molecules exhibit a distinct mechanism of action via binding the cell surface target antigen and simultaneously engaging CD3 on cytotoxic T lymphocytes (CTLs). We designed and evaluated Env-specific DARTs (HIVxCD3 DARTs) derived from known antibodies recognizing diverse Env epitopes with or without broadly neutralizing activity. HIVxCD3 DARTs derived from PGT121, PGT145, A32, and 7B2, but not VRC01 or 10E8 antibodies, mediated potent CTL-dependent killing of quiescent primary CD4 T cells infected with diverse HIV isolates. Similar killing activity was also observed with DARTs structurally modified for in vivo half-life extension. In an ex vivo model using cells isolated from HIV-infected participants on cART, combinations of the most potent HIVxCD3 DARTs reduced HIV expression both in quiescent and activated peripheral blood mononuclear cell cultures isolated from HIV-infected participants on suppressive cART. Importantly, HIVxCD3 DARTs did not induce cell-to-cell virus spread in resting or activated CD4 T cell cultures. Collectively, these results provide support for further development of HIVxCD3 DARTs as a promising therapeutic strategy for targeting HIV reservoirs. PMID:26539983

  1. The sterol regulatory element binding proteins are essential for the metabolic programming of effector T cells and adaptive immunity

    PubMed Central

    Kidani, Yoko; Elsaesser, Heidi; Hock, M Benjamin; Vergnes, Laurent; Williams, Kevin J; Argus, Joseph P; Marbois, Beth N; Komisopoulou, Evangelia; Wilson, Elizabeth B; Osborne, Timothy F; Graeber, Thomas G; Reue, Karen; Brooks, David G; Bensinger, Steven J

    2013-01-01

    Newly activated CD8+ T cells reprogram their metabolism to meet the extraordinary biosynthetic demands of clonal expansion; however, the signals mediating metabolic reprogramming remain poorly defined. Herein, we demonstrate an essential role for sterol regulatory element binding proteins (SREBPs) in the acquisition of effector cell metabolism. Without SREBP signaling, CD8+ T cells are unable to blast, resulting in markedly attenuated clonal expansion during viral infection. Mechanistic studies indicate that SREBPs are essential to meet the heightened lipid requirements of membrane synthesis during blastogenesis. SREBPs are dispensable for homeostatic proliferation, indicating a context-specific requirement for SREBPs in effector responses. These studies provide insights into the molecular signals underlying metabolic reprogramming of CD8+ T cells during the transition from quiescence to activation. PMID:23563690

  2. CD8 T-cell recognition of acquired alloantigen promotes acute allograft rejection

    PubMed Central

    Harper, Simon J. F.; Ali, Jason M.; Wlodek, Elizabeth; Negus, Marg C.; Harper, Ines G.; Chhabra, Manu; Qureshi, M. Saeed; Mallik, Mekhola; Bolton, Eleanor; Bradley, J. Andrew; Pettigrew, Gavin J.

    2015-01-01

    Adaptive CD8 T-cell immunity is the principal arm of the cellular alloimmune response, but its development requires help. This can be provided by CD4 T cells that recognize alloantigen “indirectly,” as self-restricted allopeptide, but this process remains unexplained, because the target epitopes for CD4 and CD8 T-cell recognition are “unlinked” on different cells (recipient and donor antigen presenting cells (APCs), respectively). Here, we test the hypothesis that the presentation of intact and processed MHC class I alloantigen by recipient dendritic cells (DCs) (the “semidirect” pathway) allows linked help to be delivered by indirect-pathway CD4 T cells for generating destructive cytotoxic CD8 T-cell alloresponses. We show that CD8 T-cell–mediated rejection of murine heart allografts that lack hematopoietic APCs requires host secondary lymphoid tissue (SLT). SLT is necessary because within it, recipient dendritic cells can acquire MHC from graft parenchymal cells and simultaneously present it as intact protein to alloreactive CD8 T cells and as processed peptide alloantigen for recognition by indirect-pathway CD4 T cells. This enables delivery of essential help for generating cytotoxic CD8 T-cell responses that cause rapid allograft rejection. In demonstrating the functional relevance of the semidirect pathway to transplant rejection, our findings provide a solution to a long-standing conundrum as to why SLT is required for CD8 T-cell allorecognition of graft parenchymal cells and suggest a mechanism by which indirect-pathway CD4 T cells provide help for generating effector cytotoxic CD8 T-cell alloresponses at late time points after transplantation. PMID:26420874

  3. Coupling of HIV-1 Antigen to the Selective Autophagy Receptor SQSTM1/p62 Promotes T-Cell-Mediated Immunity

    PubMed Central

    Andersen, Aram Nikolai; Landsverk, Ole Jørgen; Simonsen, Anne; Bogen, Bjarne; Corthay, Alexandre; Øynebråten, Inger

    2016-01-01

    Vaccines aiming to promote T-cell-mediated immune responses have so far showed limited efficacy, and there is a need for novel strategies. Studies indicate that autophagy plays an inherent role in antigen processing and presentation for CD4+ and CD8+ T cells. Here, we report a novel vaccine strategy based on fusion of antigen to the selective autophagy receptor sequestosome 1 (SQSTM1)/p62. We hypothesized that redirection of vaccine antigen from proteasomal degradation into the autophagy pathway would increase the generation of antigen-specific T cells. A hybrid vaccine construct was designed in which the antigen is fused to the C-terminus of p62, a signaling hub, and a receptor that naturally delivers ubiquitinated cargo for autophagic degradation. Fusion of the human immunodeficiency virus-1 antigen Gagp24 to p62 resulted in efficient antigen delivery into the autophagy pathway. Intradermal immunization of mice revealed that, in comparison to Gagp24 delivered alone, fusion to p62 enhanced the number of Gagp24-specific interferon-γ-producing T cells, including CD8+ T cells. The strategy may also have the potential to modulate the antigenic peptide repertoire. Because p62 and autophagy are highly conserved between species, we anticipate this strategy to be a candidate for the development of T-cell-based vaccines in humans. PMID:27242780

  4. Id1 expression promotes peripheral CD4{sup +} T cell proliferation and survival upon TCR activation without co-stimulation

    SciTech Connect

    Liu, Chen; Jin, Rong; Wang, Hong-Cheng; Tang, Hui; Liu, Yuan-Feng; Qian, Xiao-Ping; Sun, Xiu-Yuan; Ge, Qing; Sun, Xiao-Hong; Zhang, Yu

    2013-06-21

    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{sup +} 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.

  5. Integrin α1β1 Promotes Caveolin-1 Dephosphorylation by Activating T Cell Protein-tyrosine Phosphatase*

    PubMed Central

    Borza, Corina M.; Chen, Xiwu; Mathew, Sijo; Mont, Stacey; Sanders, Charles R.; Zent, Roy; Pozzi, Ambra

    2010-01-01

    Integrin α1β1 is a collagen receptor that down-regulates collagen and reactive oxygen species (ROS) production, and mice lacking this receptor show increased ROS levels and exacerbated glomerular sclerosis following injury. Caveolin-1 (Cav-1) is a multifunctional protein that is tyrosine-phosphorylated in response to injury and has been implicated in ROS-mediated injury. Cav-1 interacts with integrins, and integrin α1β1 binds/activates T cell protein-tyrosine phosphatase (TCPTP), which is homologous to the tyrosine phosphatase PTP1B known to dephosphorylate Cav-1. In this study, we analyzed whether phosphorylated Cav-1 (pCav-1) is a substrate of TCPTP and if integrin α1β1 is essential for promoting TCPTP-mediated Cav-1 dephosphorylation. We found that Cav-1 phosphorylation is significantly higher in cells lacking integrin α1β1 at base line and following oxidative stress. Overexpression of TCPTP leads to reduced pCav-1 levels only in cells expressing integrin α1β1. Using solid phase binding assays, we demonstrated that 1) purified Cav-1 directly interacts with TCPTP and the integrin α1 subunit, 2) pCav-1 is a substrate of TCPTP, and 3) TCPTP-mediated Cav-1 dephosphorylation is highly increased by the addition of purified integrin α1β1 or an integrin α1 cytoplasmic peptide to which TCPTP has been shown to bind. Thus, our results demonstrate that pCav-1 is a new substrate of TCPTP and that integrin α1β1 acts as a negative regulator of Cav-1 phosphorylation by activating TCPTP. This could explain the protective function of integrin α1β1 in oxidative stress-mediated damage and why integrin α1-null mice are more susceptible to fibrosis following injury. PMID:20940300

  6. Integrin {alpha}1{beta}1 promotes caveolin-1 dephosphorylation by activating T cell protein-tyrosine phosphatase.

    PubMed

    Borza, Corina M; Chen, Xiwu; Mathew, Sijo; Mont, Stacey; Sanders, Charles R; Zent, Roy; Pozzi, Ambra

    2010-12-17

    Integrin α1β1 is a collagen receptor that down-regulates collagen and reactive oxygen species (ROS) production, and mice lacking this receptor show increased ROS levels and exacerbated glomerular sclerosis following injury. Caveolin-1 (Cav-1) is a multifunctional protein that is tyrosine-phosphorylated in response to injury and has been implicated in ROS-mediated injury. Cav-1 interacts with integrins, and integrin α1β1 binds/activates T cell protein-tyrosine phosphatase (TCPTP), which is homologous to the tyrosine phosphatase PTP1B known to dephosphorylate Cav-1. In this study, we analyzed whether phosphorylated Cav-1 (pCav-1) is a substrate of TCPTP and if integrin α1β1 is essential for promoting TCPTP-mediated Cav-1 dephosphorylation. We found that Cav-1 phosphorylation is significantly higher in cells lacking integrin α1β1 at base line and following oxidative stress. Overexpression of TCPTP leads to reduced pCav-1 levels only in cells expressing integrin α1β1. Using solid phase binding assays, we demonstrated that 1) purified Cav-1 directly interacts with TCPTP and the integrin α1 subunit, 2) pCav-1 is a substrate of TCPTP, and 3) TCPTP-mediated Cav-1 dephosphorylation is highly increased by the addition of purified integrin α1β1 or an integrin α1 cytoplasmic peptide to which TCPTP has been shown to bind. Thus, our results demonstrate that pCav-1 is a new substrate of TCPTP and that integrin α1β1 acts as a negative regulator of Cav-1 phosphorylation by activating TCPTP. This could explain the protective function of integrin α1β1 in oxidative stress-mediated damage and why integrin α1-null mice are more susceptible to fibrosis following injury. PMID:20940300

  7. Intestinal commensal bacteria promote T cell hyporesponsiveness and down-regulate the serum antibody responses induced by dietary antigen.

    PubMed

    Tsuda, Masato; Hosono, Akira; Yanagibashi, Tsutomu; Kihara-Fujioka, Miran; Hachimura, Satoshi; Itoh, Kikuji; Hirayama, Kazuhiro; Takahashi, Kyoko; Kaminogawa, Shuichi

    2010-08-16

    Colonization of the gut by commensal bacteria modulates the induction of oral tolerance and allergy. However, how these intestinal bacteria modulate antigen-specific T cell responses induced by oral antigens remains unclear. In order to investigate this, we used germ-free (GF) ovalbumin (OVA)-specific T cell receptor transgenic (OVA23-3) mice. Conventional (CV) or GF mice were administered an OVA-containing diet. Cytokine production by CD4(+) cells from spleen (SP), mesenteric lymph nodes (MLN) and Peyer's patches (PP) was evaluated by ELISA, as was the peripheral antibody titer. T cell phenotype was assessed by flow cytometry. CD4(+) cells from the SP and MLN of CV and GF mice fed an OVA diet for 3 weeks produced significantly less IL-2 than the corresponding cells from mice receiving a control diet, suggesting that oral tolerance could be induced at the T cell level in the systemic and intestinal immune systems of both bacterial condition of mice. However, we also observed that the T cell hyporesponsiveness induced by dietary antigen was delayed in the systemic immune tissues and was weaker in the intestinal immune tissues of the GF mice. Intestinal MLN and PP CD4(+) T cells from these animals also produced lower levels of IL-10, had less activated/memory type CD45RB(low) cells, and expressed lower levels of CTLA-4 but not Foxp3 compared to their CV counterparts. Furthermore, GF mice produced higher serum levels of OVA-specific antibodies than CV animals. CD40L expression by SP CD4(+) cells from GF mice fed OVA was higher than that of CV mice. These results suggest that intestinal commensal bacteria promote T cell hyporesponsiveness and down-regulate serum antibody responses induced by dietary antigens through modulation of the intestinal and systemic T cell phenotype. PMID:20621647

  8. Delineation of a T-cell activation motif required for binding of protein tyrosine kinases containing tandem SH2 domains.

    PubMed Central

    Koyasu, S; Tse, A G; Moingeon, P; Hussey, R E; Mildonian, A; Hannisian, J; Clayton, L K; Reinherz, E L

    1994-01-01

    To define the T-cell receptor signal transduction motif, we have transfected human and murine T-cell lines with a chimeric receptor consisting of the extracellular and transmembrane domains of human CD8 alpha and the membrane-proximal portion of CD3 zeta containing at its C terminus either an 18-amino acid segment (NQLYNELNLGRREEYDVL) or alanine-scanning point mutant derivatives. Crosslinking of the extracellular domain of the chimera is sufficient to initiate Ca2+ flux, interleukin 2 production, and tyrosine phosphorylation of cellular proteins including the chimera. Subsequently, the chimera becomes associated with several tyrosine-phosphorylated proteins, among them the 70-kDa protein tyrosine kinase ZAP70. Mutational data identify the T-cell activation motif as Y(X)2L(X)7Y(X)2L and show that each of the four designated residues is necessary for the above activation events. Recombinant protein containing the two tandem SH2 domains derived from ZAP70 binds to a synthetic peptide corresponding to the above 18-amino acid motif but only when both tyrosines are phosphorylated; in contrast, little or no binding is observed to monophosphorylated or nonphosphorylated analogues. These results imply that after receptor crosslinking in T cells, and by inference also in B cells and mast cells, the motif is phosphorylated on both tyrosine residues, thereafter serving as a docking site for protein tyrosine kinases containing tandem SH2 domains. Images PMID:7517560

  9. 5-Azacytidine Promotes an Inhibitory T-Cell Phenotype and Impairs Immune Mediated Antileukemic Activity

    PubMed Central

    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

  10. 5-azacytidine promotes an inhibitory T-cell phenotype and impairs immune mediated antileukemic activity.

    PubMed

    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

  11. Investigating potential exogenous tumor initiating and promoting factors for Cutaneous T-Cell Lymphomas (CTCL), a rare skin malignancy.

    PubMed

    Litvinov, Ivan V; Shtreis, Anna; Kobayashi, Kenneth; Glassman, Steven; Tsang, Matthew; Woetmann, Anders; Sasseville, Denis; Ødum, Niels; Duvic, Madeleine

    2016-07-01

    Most skin malignancies are caused by external and often preventable environmental agents. Multiple reports demonstrated that cutaneous T-cell lymphomas (CTCL) can occur in married couples and cluster in families. Furthermore, recent studies document geographic clustering of this malignancy in Texas as well as in other areas of the United States. Multiple infectious, occupational, and medication causes have been proposed as triggers or promoters of this malignancy including hydrochlorothiazide diuretics, Staphylococcus aureus, dermatophytes, Mycobacterium leprae, Chlamydia pneumoniae, human T-Cell lymphotropic virus type 1 (HTLV1), Epstein-Barr virus (EBV), and herpes simplex virus (HSV). In this report, we review recent evidence evaluating the involvement of these agents in cancer initiation/progression. Most importantly, recent molecular experimental evidence documented for the first time that S. aureus can activate oncogenic STAT3 signaling in malignant T cells. Specifically, S. aureus Enterotoxin type A (SEA) was recently shown to trigger non-malignant infiltrating T cells to release IL-2 and other cytokines. These signals upon binging to their cognate receptors on malignant T cells are then able to activate STAT3 and STAT5 oncogenic signaling and promote cancer progression and IL-17 secretion. In light of these findings, it might be important for patients with exacerbation of their CTCL symptoms to maintain high index of suspicion and treat these individuals for S. aureus colonization and/or sepsis with topical and systemic antibiotics. PMID:27622024

  12. RelB-Dependent Stromal Cells Promote T-Cell Leukemogenesis

    PubMed Central

    dos Santos, Nuno R.; Williame, Maryvonne; Gachet, Stéphanie; Cormier, Françoise; Janin, Anne; Weih, Debra; Weih, Falk; Ghysdael, Jacques

    2008-01-01

    Background The Rel/NF-κB transcription factors are often activated in solid or hematological malignancies. In most cases, NF-κB activation is found in malignant cells and results from activation of the canonical NF-κB pathway, leading to RelA and/or c-Rel activation. Recently, NF-κB activity in inflammatory cells infiltrating solid tumors has been shown to contribute to solid tumor initiation and progression. Noncanonical NF-κB activation, which leads to RelB activation, has also been reported in breast carcinoma, prostate cancer, and lymphoid leukemia. Methodology/Principal Findings Here we report a novel role for RelB in stromal cells that promote T-cell leukemogenesis. RelB deficiency delayed leukemia onset in the TEL-JAK2 transgenic mouse model of human T acute lymphoblastic leukemia. Bone marrow chimeric mouse experiments showed that RelB is not required in the hematopoietic compartment. In contrast, RelB plays a role in radio-resistant stromal cells to accelerate leukemia onset and increase disease severity. Conclusions/Significance The present results are the first to uncover a role for RelB in the crosstalk between non-hematopoietic stromal cells and leukemic cells. Thus, besides its previously reported role intrinsic to specific cancer cells, the noncanonical NF-κB pathway may also play a pro-oncogenic role in cancer microenvironmental cells. PMID:18596915

  13. Fluorine substitutions in an antigenic peptide selectively modulate T-cell receptor binding in a minimally perturbing manner

    SciTech Connect

    Piepenbrink, Kurt H.; Borbulevych, Oleg Y.; Sommese, Ruth F.; Clemens, John; Armstrong, Kathryn M.; Desmond, Clare; Do, Priscilla; Baker, Brian M.

    2010-08-17

    TCR (T-cell receptor) recognition of antigenic peptides bound and presented by MHC (major histocompatibility complex) molecules forms the basis of the cellular immune response to pathogens and cancer. TCRs bind peptide - MHC complexes weakly and with fast kinetics, features which have hindered detailed biophysical studies of these interactions. Modified peptides resulting in enhanced TCR binding could help overcome these challenges. Furthermore, there is considerable interest in using modified peptides with enhanced TCR binding as the basis for clinical vaccines. In the present study, we examined how fluorine substitutions in an antigenic peptide can selectively impact TCR recognition. Using a structure-guided design approach, we found that fluorination of the Tax peptide [HTLV (human T-cell lymphotropic virus)-1 Tax] enhanced binding by the Tax-specific TCR A6, yet weakened binding by the Tax-specific TCR B7. The changes in affinity were consistent with crystallographic structures and fluorine chemistry, and with the A6 TCR independent of other substitutions in the interface. Peptide fluorination thus provides a means to selectively modulate TCR binding affinity without significantly perturbing peptide composition or structure. Lastly, we probed the mechanism of fluorine's effect on TCR binding and we conclude that our results were most consistent with a 'polar hydrophobicity' mechanism, rather than a purely hydrophobic- or electrostatic-based mechanism. This finding should have an impact on other attempts to alter molecular recognition with fluorine.

  14. The RNA-Binding Protein, Polypyrimidine Tract-Binding Protein 1 (PTBP1) Is a Key Regulator of CD4 T Cell Activation

    PubMed Central

    Valentín-Acevedo, Aníbal

    2016-01-01

    We have previously shown that the RNA binding protein, polypyrimidine tract-binding protein (PTBP1) plays a critical role in regulating the expression of CD40L in activated CD4 T cells. This is achieved mechanistically through message stabilization at late times of activation as well as by altered distribution of CD40L mRNA within distinct cellular compartments. PTBP1 has been implicated in many different processes, however whether PTBP1 plays a broader role in CD4 T cell activation is not known. To examine this question, experiments were designed to introduce shRNA into primary human CD4 T cells to achieve decreased, but not complete ablation of PTBP1 expression. Analyses of shPTB-expressing CD4 T cells revealed multiple processes including cell proliferation, activation-induced cell death and expression of activation markers and cytokines that were regulated in part by PTBP1 expression. Although there was an overall decrease in the steady-state level of several activation genes, only IL-2 and CD40L appeared to be regulated by PTBP1 at the level of RNA decay suggesting that PTBP1 is critical at different regulatory steps of expression that is gene-specific. Importantly, even though the IL-2 protein levels were reduced in cells with lowered PTBP1, the steady-state level of IL-2 mRNA was significantly higher in these cells suggesting a block at the translational level. Evaluation of T cell activation in shPTB-expressing T cells revealed that PTBP1 was linked primarily to the activation of the PLCγ1/ERK1/2 and the NF-κB pathways. Overall, our results reveal the importance of this critical RNA binding protein in multiple steps of T cell activation. PMID:27513449

  15. Attenuation of microglial RANTES by NEMO-binding domain peptide inhibits the infiltration of CD8(+) T cells in the nigra of hemiparkinsonian monkey.

    PubMed

    Roy, A; Mondal, S; Kordower, J H; Pahan, K

    2015-08-27

    Parkinson's disease (PD) is a progressive neurodegenerative disease characterized by the loss of dopaminergic (DA) neurons in the substantia nigra pars compacta (SNpc). Despite intense investigations, little is known about its pathological mediators. Here, we report the marked upregulation of RANTES (regulated on activation, normal T cell expressed and secreted) and eotaxin, chemokines that are involved in T cell trafficking, in the serum of hemiparkinsonian monkeys. Interestingly, 1-methyl-4-phenylpyridinium (MPP(+)), a Parkinsonian toxin, increased the expression of RANTES and eotaxin in mouse microglial cells. The presence of NF-κB binding sites in promoters of RANTES and eotaxin and down-regulation of these genes by NEMO-binding domain (NBD) peptide, selective inhibitor of induced NF-κB activation, in MPP(+)-stimulated microglial cells suggest that the activation of NF-κB plays an important role in the upregulation of these two chemokines. Consistently, serum enzyme-linked immuno assay (ELISA) and nigral immunohistochemistry further confirmed that these chemokines were strongly upregulated in MPTP-induced hemiparkinsonian monkeys and that treatment with NBD peptides effectively inhibited the level of these chemokines. Furthermore, the microglial upregulation of RANTES in the nigra of hemiparkinsonian monkeys could be involved in the altered adaptive immune response in the brain as we observed greater infiltration of CD8(+) T cells around the perivascular niche and deep brain parenchyma of hemiparkinsonian monkeys as compared to control. The treatment of hemiparkinsonian monkeys with NBD peptides decreased the microglial expression of RANTES and attenuated the infiltration of CD8(+) T cells in nigra. These results indicate the possible involvement of chemokine-dependent adaptive immune response in Parkinsonism. PMID:25783477

  16. Differential transactivation of the intercellular adhesion molecule 1 gene promoter by Tax1 and Tax2 of human T-cell leukemia viruses.

    PubMed Central

    Tanaka, Y; Hayashi, M; Takagi, S; Yoshie, O

    1996-01-01

    Previously, we showed that surface expression of intercellular adhesion molecule 1 (ICAM-1) was strongly upregulated in T cells carrying proviral human T-cell leukemia virus type 1 (HTLV-1) and that the viral transactivator protein Tax1 was capable of inducing the ICAM-1 gene. To determine the responsive elements in the human ICAM-1 gene promoter, a reporter construct in which the 5'-flanking 4.4-kb region of the ICAM-1 gene was linked to the promoterless chloramphenicol acetyltransferase (CAT) gene was cotransfected with expression vectors for Tax1 and Tax2, both of which were separately confirmed to be potent transactivators of the HTLV-1 long terminal repeat (LTR). Tax1 strongly activated the ICAM-1 promoter in all the cell lines tested: three T-cell lines (Jurkat, MOLT-4, and CEM), one monocytoid cell line (U937), and HeLa. Unexpectedly, Tax2 activated the ICAM-1 promoter only in HeLa. By deletion and mutation analyses of the 1.3-kb 5'-flanking region, we found that Tax1 transactivated the ICAM-1 promoter mainly via a cyclic AMP-responsive element (CRE)-like site at -630 to -624 in the Jurkat T-cell line and via an NF-kappaB site at -185 to -177 and an SP-1 site at -59 to -54 in HeLa. On the other hand, Tax2 was totally inactive on the ICAM-1 promoter in Jurkat but transactivated the promoter via the NF-kappaB site at -185 to -177 in HeLa. Gel mobility shift assays demonstrated proteins specifically binding to the CRE-like site at -630 to -624 in Tax1-expressing T-cell lines. Stable expression of Tax1 but not Tax2 in Jurkat subclones enhanced the surface expression of ICAM-1. The differential ability of Tax1 and Tax2 in transactivation of the ICAM-1 gene may be related to the differential pathogenicity of HTLV-1 and HTLV-2. PMID:8970974

  17. Oral bacteria induce a differential activation of human immunodeficiency virus-1 promoter in T cells, macrophages and dendritic cells

    PubMed Central

    Huang, C. B.; Emerson, K. A.; Gonzalez, O. A.; Ebersole, J. L.

    2014-01-01

    Introduction The human immunodeficiency virus (HIV) can integrate into T cells, macrophages and dendritic cells resulting in a latent infection. Reports have also demonstrated that various microbial and host cell factors can trigger HIV reactivation leading to HIV recrudescence, potentially undermining highly active antiretroviral therapies. Methods This study evaluated the capacity of oral bacteria associated with chronic periodontal infections to stimulate HIV promoter activation in various cell models of HIV latency. Results T cells (1G5) challenged with oral bacteria demonstrated a dose–response of HIV promoter activation with a subset of the bacteria, as well as kinetics that were generally similar irrespective of the stimuli. Direct bacterial challenge of the T cells resulted in increased activation of approximately 1.5- to 7-fold over controls. Challenge of macrophages (BF24) indicated different kinetics for individual bacteria and resulted in consistent increases in promoter activation of five fold to six fold over basal levels for all bacteria except Streptococcus mutans. Dendritic cells showed increases in HIV reactivation of 7- to 34-fold specific for individual species of bacteria. Conclusion These results suggested that oral bacteria have the capability to reactivate HIV from latently infected cells, showing a relationship of mature dendritic cells > immature dendritic cells > macrophages ≥ T cells. Expression of various pattern recognition receptors on these various cell types may provide insight into the primary receptors/signaling pathways used for reactivation by the bacteria. PMID:19702954

  18. Adhesion- and Degranulation-Promoting Adapter Protein Promotes CD8 T Cell Differentiation and Resident Memory Formation and Function during an Acute Infection.

    PubMed

    Fiege, Jessica K; Beura, Lalit K; Burbach, Brandon J; Shimizu, Yoji

    2016-09-15

    During acute infections, naive Ag-specific CD8 T cells are activated and differentiate into effector T cells, most of which undergo contraction after pathogen clearance. A small population of CD8 T cells persists as memory to protect against future infections. We investigated the role of adhesion- and degranulation-promoting adapter protein (ADAP) in promoting CD8 T cell responses to a systemic infection. Naive Ag-specific CD8 T cells lacking ADAP exhibited a modest expansion defect early after Listeria monocytogenes or vesicular stomatitis virus infection but comparable cytolytic function at the peak of response. However, reduced numbers of ADAP-deficient CD8 T cells were present in the spleen after the peak of the response. ADAP deficiency resulted in a greater frequency of CD127(+) CD8 memory precursors in secondary lymphoid organs during the contraction phase. Reduced numbers of ADAP-deficient killer cell lectin-like receptor G1(-) CD8 resident memory T (TRM) cell precursors were present in a variety of nonlymphoid tissues at the peak of the immune response, and consequently the total numbers of ADAP-deficient TRM cells were reduced at memory time points. TRM cells that did form in the absence of ADAP were defective in effector molecule expression. ADAP-deficient TRM cells exhibited impaired effector function after Ag rechallenge, correlating with defects in their ability to form T cell-APC conjugates. However, ADAP-deficient TRM cells responded to TGF-β signals and recruited circulating memory CD8 T cells. Thus, ADAP regulates CD8 T cell differentiation events following acute pathogen challenge that are critical for the formation and selected functions of TRM cells in nonlymphoid tissues. PMID:27521337

  19. Endothelial-binding, proinflammatory T cells identified by MCAM (CD146) expression: Characterization and role in human autoimmune diseases.

    PubMed

    Dagur, Pradeep K; McCoy, J Philip

    2015-05-01

    A subset of T cells defined by the cell surface expression of MCAM (CD146) has been identified in the peripheral circulation of healthy individuals. These cells comprise approximately 3% of the pool of circulating T cells, have an effector memory phenotype, and are capable of producing several cytokines. Notably, the MCAM positive cells are enhanced for IL-17 production compared to MCAM negative effector memory T cells. These cells are committed to IL-17 production and do not require in vitro polarization with exogenous cytokines. MCAM positive T cells also demonstrate an increased ability to bind to endothelial monolayers. In numerous autoimmune diseases these cells are found at increased proportions in the peripheral circulation, and at the sites of active inflammation in patients with autoimmune disease, these cells appear in large numbers and are major contributors to IL-17 production. Studies to date have been performed with human subjects and it is uncertain if appropriate mouse models exist for this cell type. These cells could represent early components of the adaptive immune response and serve as targets of therapy in these diseases, although much work remains to be performed in order to discern the exact nature and function of these cells. PMID:25595133

  20. Oligoclonal CD4+ T Cells Promote Host Memory Immune Responses to Zwitterionic Polysaccharide of Streptococcus pneumoniae▿

    PubMed Central

    Groneck, Laura; Schrama, David; Fabri, Mario; Stephen, Tom Li; Harms, Fabian; Meemboor, Sonja; Hafke, Helena; Bessler, Martina; Becker, Jürgen C.; Kalka-Moll, Wiltrud M.

    2009-01-01

    Zwitterionic polysaccharides of the normal flora bacteria represent a novel class of antigens in that they correct systemic CD4+ T-cell deficiencies and direct lymphoid organogenesis during colonization of the host. Presentation of these polysaccharides to CD4+ T cells depends on major histocompatibility complex class II- and DM-dependent retrograde transport from lysosomes to the cell surface. Yet the phenotype and clonality of the immune response to the polysaccharide in the mature host immune system have not been studied. Using the zwitterionic capsular polysaccharide Sp1 of Streptococcus pneumoniae, a transient member of the bacterial flora, in an experimental mouse model of cellular immunity, we demonstrated the accumulation of TH1- and TH17-polarized CD4+ CD44high CD62low CD25− memory T cells. Subcutaneous immunization with Sp1 resulted in an increase of serum immunoglobulin G (IgG), predominantly of the IgG1 subclass, and suggested the presence of a humoral memory response to the polysaccharide. CD4+ T cells stimulated with polysaccharide in vitro and in vivo showed a nonrestricted pattern for the T-cell receptor (TCR) β-chain variable region, as demonstrated by semiquantitative reverse transcription-PCR and flow cytometry. Clonotype mapping of in vivo and in vitro polysaccharide-activated CD4+ T cells revealed clonotypic TCR transcripts. Taken together, the data show the induction of clonal expansion of CD4+ T cells by polysaccharides of commensal bacteria. Cellular and humoral memory host responses imply the ability of these polysaccharides to mediate the expansion of T cells via recognition within the CDR3 region of the TCR. PMID:19546196

  1. T cell- but not tumor cell-produced TGF-β1 promotes the development of spontaneous mammary cancer

    PubMed Central

    Sarkar, Abira; Donkor, Moses K.; Li, Ming O.

    2011-01-01

    During their development, tumors acquire multiple capabilities that enable them to proliferate, disseminate and evade immunosurveillance. A putative mechanism is through the production of the cytokine TGF-β1. We showed in our recent studies that T cell-produced TGF-β1 inhibits antitumor T cell responses to foster tumor growth raising the question of the precise function of TGF-β1 produced by tumor cells in tumor development. Here, using a transgenic model of mammary cancer, we report that deletion of TGF-β1 from tumor cells did not protect mice from tumor development. However, ablation of TGF-β1 from T cells significantly inhibited mammary tumor growth. Additionally, absence of TGF-β1 in T cells prevented tumors from advancing to higher pathological grades and further suppressed secondary tumor development in the lungs. These findings reveal T cells but not tumor cells as a critical source of TGF-β1 that promotes tumor development. PMID:22248703

  2. Schisandrin B inhibits Th1/Th17 differentiation and promotes regulatory T cell expansion in mouse lymphocytes.

    PubMed

    Chen, Zhaoyang; Guo, Min; Song, Guohua; Gao, Jiping; Zhang, Yinhong; Jing, Zhijie; Liu, Tianfu; Dong, Chuan

    2016-06-01

    Schisandrin B (Sch-B), the most abundant active ingredient of the fruit of Schisandra chinensis, has been proposed to have antioxidant, anti-tumor and anti-inflammatory effects. The present study was undertaken to investigate the effect of Sch-B on differentiation of T helper cells (Th). Using mouse splenic lymphocytes stimulated with concanavalin A (Con A) in vitro and ex vivo as inflammation models, we found that Sch-B significantly inhibited secretion of Th1 and Th17 related cytokines, such as IFN-γ and IL-17. In addition, we found that Sch-B suppressed the differentiation of naive CD4+ T cells into Th1 and Th17 cells, while promoted their differentiation into the regulatory T cells (Treg) in vitro. We further found that Sch-B suppressed transcription of Th1-related T-box transcription factor, T-bet, and Th17-related transcription factor, retinoid related orphan receptor gamma t (RORγt), while enhanced transcription of Treg-related transcription factor forkhead box protein 3 (Foxp3) in naive CD4+ T cells under Th cell polarization conditions. Furthermore, the effect of Sch-B on the T cell differentiation was abrogated by heme oxygenase-1 (HO-1) inhibitor zinc protoporphyrin. Taken together, we conclude that Sch-B can modulate differentiation of naïve CD4+ T cells into specific lineages of effector cells, which may have potential benefits for treatment of autoimmune diseases. PMID:27085037

  3. ICOS Promotes the Function of CD4+ Effector T Cells during Anti-OX40-Mediated Tumor Rejection.

    PubMed

    Metzger, Todd C; Long, Hua; Potluri, Shobha; Pertel, Thomas; Bailey-Bucktrout, Samantha L; Lin, John C; Fu, Tihui; Sharma, Padmanee; Allison, James P; Feldman, Reid M R

    2016-07-01

    ICOS is a T-cell coregulatory receptor that provides a costimulatory signal to T cells during antigen-mediated activation. Antitumor immunity can be improved by ICOS-targeting therapies, but their mechanism of action remains unclear. Here, we define the role of ICOS signaling in antitumor immunity using a blocking, nondepleting antibody against ICOS ligand (ICOS-L). ICOS signaling provided critical support for the effector function of CD4(+) Foxp3(-) T cells during anti-OX40-driven tumor immune responses. By itself, ICOS-L blockade reduced accumulation of intratumoral T regulatory cells (Treg), but it was insufficient to substantially inhibit tumor growth. Furthermore, it did not impede antitumor responses mediated by anti-4-1BB-driven CD8(+) T cells. We found that anti-OX40 efficacy, which is based on Treg depletion and to a large degree on CD4(+) effector T cell (Teff) responses, was impaired with ICOS-L blockade. In contrast, the provision of additional ICOS signaling through direct ICOS-L expression by tumor cells enhanced tumor rejection and survival when administered along with anti-OX40 therapy. Taken together, our results showed that ICOS signaling during antitumor responses acts on both Teff and Treg cells, which have opposing roles in promoting immune activation. Thus, effective therapies targeting the ICOS pathway should seek to promote ICOS signaling specifically in effector CD4(+) T cells by combining ICOS agonism and Treg depletion. Cancer Res; 76(13); 3684-9. ©2016 AACR. PMID:27197182

  4. Genetic absence of PD-1 promotes accumulation of terminally differentiated exhausted CD8+ T cells

    PubMed Central

    Odorizzi, Pamela M.; Pauken, Kristen E.; Paley, Michael A.; Sharpe, Arlene

    2015-01-01

    Programmed Death-1 (PD-1) has received considerable attention as a key regulator of CD8+ T cell exhaustion during chronic infection and cancer because blockade of this pathway partially reverses T cell dysfunction. Although the PD-1 pathway is critical in regulating established “exhausted” CD8+ T cells (TEX cells), it is unclear whether PD-1 directly causes T cell exhaustion. We show that PD-1 is not required for the induction of exhaustion in mice with chronic lymphocytic choriomeningitis virus (LCMV) infection. In fact, some aspects of exhaustion are more severe with genetic deletion of PD-1 from the onset of infection. Increased proliferation between days 8 and 14 postinfection is associated with subsequent decreased CD8+ T cell survival and disruption of a critical proliferative hierarchy necessary to maintain exhausted populations long term. Ultimately, the absence of PD-1 leads to the accumulation of more cytotoxic, but terminally differentiated, CD8+ TEX cells. These results demonstrate that CD8+ T cell exhaustion can occur in the absence of PD-1. They also highlight a novel role for PD-1 in preserving TEX cell populations from overstimulation, excessive proliferation, and terminal differentiation. PMID:26034050

  5. Necroptosis of Dendritic Cells Promotes Activation of γδ T Cells.

    PubMed

    Collins, Cheryl C; Bashant, Kathleen; Erikson, Cuixia; Thwe, Phyu Myat; Fortner, Karen A; Wang, Hong; Morita, Craig T; Budd, Ralph C

    2016-01-01

    γδ T cells function at the interface between innate and adaptive immunity and have well-demonstrated roles in response to infection, autoimmunity and tumors. A common characteristic of these seemingly disparate conditions may be cellular stress or death. However, the conditions under which ligands for γδ T cells are induced or exposed remain largely undefined. We observed that induction of necroptosis of murine or human dendritic cells (DC) by inhibition of caspase activity paradoxically augments their ability to activate γδ T cells. Furthermore, upregulation of the stabilizer of caspase-8 activity, c-FLIP, by IL-4, not only greatly reduced the susceptibility of DC to necroptosis, but also considerably decreased their ability to activate γδ T cells. Collectively, these findings suggest that the induction of necroptosis in DC upregulates or exposes the expression of γδ T cell ligands, and they support the view that γδ T cells function in the immune surveillance of cell stress. PMID:27431410

  6. The MAR-binding protein SATB1 orchestrates temporal and spatial expression of multiple genes during T-cell development

    SciTech Connect

    Alvarez, John D.; Yasui, Dag H.; Niida, Hiroyuki; Joh, Tadashi; Loh, Dennis Y.; Kowhi-Shigematsu, Terumi

    2000-02-24

    SATB1 is expressed primarily in thymocytes and can act as a transcriptional repressor. SATB1 binds in vivo to the matrix attachment regions (MARs) of DNA, which are implicated in the loop domain organization of chromatin. The role of MAR-binding proteins in specific cell lineages is unknown. We generated SATB1-null mice to determine how SATB1 functions in the T-cell lineage. SATB1-null mice are small in size, have disproportionately small thymi and spleens, and die at 3 weeks of age. At the cellular level, multiple defects in T-cell development were observed. Immature CD3-CD4-CD8 triple negative (TN) thymocytes were greatly reduced in number, and thymocyte development was blocked mainly at the DP stage. The few peripheral CD4{sup +} single positive (SP) cells underwent apoptosis and failed to proliferate in response to activating stimuli. At the molecular level, among 589 genes examined, at least 2% of genes including a proto-oncogene, cytokine receptor genes, and apoptosis-related genes were derepressed at inappropriate stages of T-cell development in SATB1-null mice. For example, IL-2R{alpha} and IL-7R{alpha} genes were ectopically transcribed in CD4{sup 4+}-CD{sup 8+} double positive (DP) thymocytes. SATB1 appears to orchestrate the temporal and spatial expression of genes during T-cell development, thereby ensuring the proper development of this lineage. Our data provide the first evidence that MAR-binding proteins can act as global regulators of cell function in specific cell lineages.

  7. Soluble T cell receptor Vβ domains engineered for high-affinity binding to staphylococcal or streptococcal superantigens.

    PubMed

    Sharma, Preeti; Wang, Ningyan; Kranz, David M

    2014-02-01

    Staphylococcus aureus and group A Streptococcus secrete a collection of toxins called superantigens (SAgs), so-called because they stimulate a large fraction of an individual's T cells. One consequence of this hyperactivity is massive cytokine release leading to severe tissue inflammation and, in some cases, systemic organ failure and death. The molecular basis of action involves the binding of the SAg to both a T cell receptor (TCR) on a T cell and a class II product of the major histocompatibility complex (MHC) on an antigen presenting cell. This cross-linking leads to aggregation of the TCR complex and signaling. A common feature of SAgs is that they bind with relatively low affinity to the variable region (V) of the beta chain of the TCR. Despite this low affinity binding, SAgs are very potent, as each T cell requires only a small fraction of their receptors to be bound in order to trigger cytokine release. To develop high-affinity agents that could neutralize the activity of SAgs, and facilitate the development of detection assays, soluble forms of the Vβ regions have been engineered to affinities that are up to 3 million-fold higher for the SAg. Over the past decade, six different Vβ regions against SAgs from S. aureus (SEA, SEB, SEC3, TSST-1) or S. pyogenes (SpeA and SpeC) have been engineered for high-affinity using yeast display and directed evolution. Here we review the engineering of these high-affinity Vβ proteins, structural features of the six different SAgs and the Vβ proteins, and the specific properties of the engineered Vβ regions that confer high-affinity and specificity for their SAg ligands. PMID:24476714

  8. The MAR-binding protein SATB1 orchestrates temporal and spatial expression of multiple genes during T-cell development

    PubMed Central

    Alvarez, John D.; Yasui, Dag H.; Niida, Hiroyuki; Joh, Tadashi; Loh, Dennis Y.; Kohwi-Shigematsu, Terumi

    2000-01-01

    SATB1 is expressed primarily in thymocytes and can act as a transcriptional repressor. SATB1 binds in vivo to the matrix attachment regions (MARs) of DNA, which are implicated in the loop domain organization of chromatin. The role of MAR-binding proteins in specific cell lineages is unknown. We generated SATB1-null mice to determine how SATB1 functions in the T-cell lineage. SATB1-null mice are small in size, have disproportionately small thymi and spleens, and die at 3 weeks of age. At the cellular level, multiple defects in T-cell development were observed. Immature CD3−CD4−CD8− triple negative (TN) thymocytes were greatly reduced in number, and thymocyte development was blocked mainly at the DP stage. The few peripheral CD4+ single positive (SP) cells underwent apoptosis and failed to proliferate in response to activating stimuli. At the molecular level, among 589 genes examined, at least 2% of genes including a proto-oncogene, cytokine receptor genes, and apoptosis-related genes were derepressed at inappropriate stages of T-cell development in SATB1-null mice. For example, IL-2Rα and IL-7Rα genes were ectopically transcribed in CD4+CD8+ double positive (DP) thymocytes. SATB1 appears to orchestrate the temporal and spatial expression of genes during T-cell development, thereby ensuring the proper development of this lineage. Our data provide the first evidence that MAR-binding proteins can act as global regulators of cell function in specific cell lineages. PMID:10716941

  9. Phosphoinositide binding by the SNX27 FERM domain regulates its localization at the immune synapse of activated T-cells

    PubMed Central

    Ghai, Rajesh; Tello-Lafoz, Maria; Norwood, Suzanne J.; Yang, Zhe; Clairfeuille, Thomas; Teasdale, Rohan D.; Mérida, Isabel; Collins, Brett M.

    2015-01-01

    ABSTRACT Sorting nexin 27 (SNX27) controls the endosomal-to-cell-surface recycling of diverse transmembrane protein cargos. Crucial to this function is the recruitment of SNX27 to endosomes which is mediated by the binding of phosphatidylinositol-3-phosphate (PtdIns3P) by its phox homology (PX) domain. In T-cells, SNX27 localizes to the immunological synapse in an activation-dependent manner, but the molecular mechanisms underlying SNX27 translocation remain to be clarified. Here, we examined the phosphoinositide-lipid-binding capabilities of full-length SNX27, and discovered a new PtdInsP-binding site within the C-terminal 4.1, ezrin, radixin, moesin (FERM) domain. This binding site showed a clear preference for bi- and tri-phosphorylated phophoinositides, and the interaction was confirmed through biophysical, mutagenesis and modeling approaches. At the immunological synapse of activated T-cells, cell signaling regulates phosphoinositide dynamics, and we find that perturbing phosphoinositide binding by the SNX27 FERM domain alters the SNX27 distribution in both endosomal recycling compartments and PtdIns(3,4,5)P3-enriched domains of the plasma membrane during synapse formation. Our results suggest that SNX27 undergoes dynamic partitioning between different membrane domains during immunological synapse assembly, and underscore the contribution of unique lipid interactions for SNX27 orchestration of cargo trafficking. PMID:25472716

  10. Daratumumab depletes CD38+ immune regulatory cells, promotes T-cell expansion, and skews T-cell repertoire in multiple myeloma.

    PubMed

    Krejcik, Jakub; Casneuf, Tineke; Nijhof, Inger S; Verbist, Bie; Bald, Jaime; Plesner, Torben; Syed, Khaja; Liu, Kevin; van de Donk, Niels W C J; Weiss, Brendan M; Ahmadi, Tahamtan; Lokhorst, Henk M; Mutis, Tuna; Sasser, A Kate

    2016-07-21

    Daratumumab targets CD38-expressing myeloma cells through a variety of immune-mediated mechanisms (complement-dependent cytotoxicity, antibody-dependent cell-mediated cytotoxicity, and antibody-dependent cellular phagocytosis) and direct apoptosis with crosslinking. These mechanisms may also target nonplasma cells that express CD38, which prompted evaluation of daratumumab's effects on CD38-positive immune subpopulations. Peripheral blood (PB) and bone marrow (BM) from patients with relapsed/refractory myeloma from 2 daratumumab monotherapy studies were analyzed before and during therapy and at relapse. Regulatory B cells and myeloid-derived suppressor cells, previously shown to express CD38, were evaluated for immunosuppressive activity and daratumumab sensitivity in the myeloma setting. A novel subpopulation of regulatory T cells (Tregs) expressing CD38 was identified. These Tregs were more immunosuppressive in vitro than CD38-negative Tregs and were reduced in daratumumab-treated patients. In parallel, daratumumab induced robust increases in helper and cytotoxic T-cell absolute counts. In PB and BM, daratumumab induced significant increases in CD8(+):CD4(+) and CD8(+):Treg ratios, and increased memory T cells while decreasing naïve T cells. The majority of patients demonstrated these broad T-cell changes, although patients with a partial response or better showed greater maximum effector and helper T-cell increases, elevated antiviral and alloreactive functional responses, and significantly greater increases in T-cell clonality as measured by T-cell receptor (TCR) sequencing. Increased TCR clonality positively correlated with increased CD8(+) PB T-cell counts. Depletion of CD38(+) immunosuppressive cells, which is associated with an increase in T-helper cells, cytotoxic T cells, T-cell functional response, and TCR clonality, represents possible additional mechanisms of action for daratumumab and deserves further exploration. PMID:27222480

  11. Daratumumab depletes CD38+ immune regulatory cells, promotes T-cell expansion, and skews T-cell repertoire in multiple myeloma

    PubMed Central

    Krejcik, Jakub; Casneuf, Tineke; Nijhof, Inger S.; Verbist, Bie; Bald, Jaime; Plesner, Torben; Syed, Khaja; Liu, Kevin; van de Donk, Niels W. C. J.; Weiss, Brendan M.; Ahmadi, Tahamtan; Lokhorst, Henk M.; Mutis, Tuna

    2016-01-01

    Daratumumab targets CD38-expressing myeloma cells through a variety of immune-mediated mechanisms (complement-dependent cytotoxicity, antibody-dependent cell-mediated cytotoxicity, and antibody-dependent cellular phagocytosis) and direct apoptosis with crosslinking. These mechanisms may also target nonplasma cells that express CD38, which prompted evaluation of daratumumab’s effects on CD38-positive immune subpopulations. Peripheral blood (PB) and bone marrow (BM) from patients with relapsed/refractory myeloma from 2 daratumumab monotherapy studies were analyzed before and during therapy and at relapse. Regulatory B cells and myeloid-derived suppressor cells, previously shown to express CD38, were evaluated for immunosuppressive activity and daratumumab sensitivity in the myeloma setting. A novel subpopulation of regulatory T cells (Tregs) expressing CD38 was identified. These Tregs were more immunosuppressive in vitro than CD38-negative Tregs and were reduced in daratumumab-treated patients. In parallel, daratumumab induced robust increases in helper and cytotoxic T-cell absolute counts. In PB and BM, daratumumab induced significant increases in CD8+:CD4+ and CD8+:Treg ratios, and increased memory T cells while decreasing naïve T cells. The majority of patients demonstrated these broad T-cell changes, although patients with a partial response or better showed greater maximum effector and helper T-cell increases, elevated antiviral and alloreactive functional responses, and significantly greater increases in T-cell clonality as measured by T-cell receptor (TCR) sequencing. Increased TCR clonality positively correlated with increased CD8+ PB T-cell counts. Depletion of CD38+ immunosuppressive cells, which is associated with an increase in T-helper cells, cytotoxic T cells, T-cell functional response, and TCR clonality, represents possible additional mechanisms of action for daratumumab and deserves further exploration. PMID:27222480

  12. T Cells

    MedlinePlus

    ... or turn off the immune response. Cytotoxic or “killer” T cells directly attack and destroy cells bearing ... involve selective activation of helper T cells and killer T cells, with a corresponding decrease in regulatory ...

  13. Mst1 Kinase Regulates the Actin-Bundling Protein L-Plastin To Promote T Cell Migration.

    PubMed

    Xu, Xiaolu; Wang, Xinxin; Todd, Elizabeth M; Jaeger, Emily R; Vella, Jennifer L; Mooren, Olivia L; Feng, Yunfeng; Hu, Jiancheng; Cooper, John A; Morley, Sharon Celeste; Huang, Yina H

    2016-09-01

    Exploring the mechanisms controlling lymphocyte trafficking is essential for understanding the function of the immune system and the pathophysiology of immunodeficiencies. The mammalian Ste20-like kinase 1 (Mst1) has been identified as a critical signaling mediator of T cell migration, and loss of Mst1 results in immunodeficiency disease. Although Mst1 is known to support T cell migration through induction of cell polarization and lamellipodial formation, the downstream effectors of Mst1 are incompletely defined. Mice deficient for the actin-bundling protein L-plastin (LPL) have phenotypes similar to mice lacking Mst1, including decreased T cell polarization, lamellipodial formation, and cell migration. We therefore asked whether LPL functions downstream of Mst1. The regulatory N-terminal domain of LPL contains a consensus Mst1 phosphorylation site at Thr(89) We found that Mst1 can phosphorylate LPL in vitro and that Mst1 can interact with LPL in cells. Removal of the Mst1 phosphorylation site by mutating Thr(89) to Ala impaired localization of LPL to the actin-rich lamellipodia of T cells. Expression of the T89A LPL mutant failed to restore migration of LPL-deficient T cells in vitro. Furthermore, expression of T89A LPL in LPL-deficient hematopoietic cells, using bone marrow chimeras, failed to rescue the phenotype of decreased thymic egress. These results identify LPL as a key effector of Mst1 and establish a novel mechanism linking a signaling intermediate to an actin-binding protein critical to T cell migration. PMID:27465533

  14. The Regulatory T Cell Lineage Factor Foxp3 Regulates Gene Expression through Several Distinct Mechanisms Mostly Independent of Direct DNA Binding

    PubMed Central

    Andersen, Kristian G.; Hebenstreit, Daniel; Teichmann, Sarah A.; Betz, Alexander G.

    2015-01-01

    The lineage factor Foxp3 is essential for the development and maintenance of regulatory T cells, but little is known about the mechanisms involved. Here, we demonstrate that an N-terminal proline-rich interaction region is crucial for Foxp3’s function. Subdomains within this key region link Foxp3 to several independent mechanisms of transcriptional regulation. Our study suggests that Foxp3, even in the absence of its DNA-binding forkhead domain, acts as a bridge between DNA-binding interaction partners and proteins with effector function permitting it to regulate a large number of genes. We show that, in one such mechanism, Foxp3 recruits class I histone deacetylases to the promoters of target genes, counteracting activation-induced histone acetylation and thereby suppressing their expression. PMID:26107960

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

    PubMed

    Côme, Christophe; 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; Hrabě de Angelis, Martin; 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

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

    PubMed Central

    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

  17. Local antigen in nonlymphoid tissue promotes resident memory CD8+ T cell formation during viral infection.

    PubMed

    Khan, Tahsin N; Mooster, Jana L; Kilgore, Augustus M; Osborn, Jossef F; Nolz, Jeffrey C

    2016-05-30

    Tissue-resident memory (Trm) CD8(+) T cells are functionally distinct from their circulating counterparts and are potent mediators of host protection against reinfection. Whether local recognition of antigen in nonlymphoid tissues during infection can impact the formation of Trm populations remains unresolved. Using skin infections with vaccinia virus (VacV)-expressing model antigens, we found that local antigen recognition had a profound impact on Trm formation. Activated CD8(+) T cells trafficked to VacV-infected skin in an inflammation-dependent, but antigen-independent, manner. However, after viral clearance, there was a subsequent ∼50-fold increase in Trm formation when antigen was present in the tissue microenvironment. Secondary antigen stimulation in nonlymphoid tissue caused CD8(+) T cells to rapidly express CD69 and be retained at the site of infection. Finally, Trm CD8(+) T cells that formed during VacV infection in an antigen-dependent manner became potent stimulators of localized antigen-specific inflammatory responses in the skin. Thus, our studies indicate that the presence of antigen in the nonlymphoid tissue microenvironment plays a critical role in the formation of functional Trm CD8(+) T cell populations, a finding with relevance for both vaccine design and prevention of inflammatory disorders. PMID:27217536

  18. IL-17-producing γδ T cells and neutrophils conspire to promote breast cancer metastasis.

    PubMed

    Coffelt, Seth B; Kersten, Kelly; Doornebal, Chris W; Weiden, Jorieke; Vrijland, Kim; Hau, Cheei-Sing; Verstegen, Niels J M; Ciampricotti, Metamia; Hawinkels, Lukas J A C; Jonkers, Jos; de Visser, Karin E

    2015-06-18

    Metastatic disease remains the primary cause of death for patients with breast cancer. The different steps of the metastatic cascade rely on reciprocal interactions between cancer cells and their microenvironment. Within this local microenvironment and in distant organs, immune cells and their mediators are known to facilitate metastasis formation. However, the precise contribution of tumour-induced systemic inflammation to metastasis and the mechanisms regulating systemic inflammation are poorly understood. Here we show that tumours maximize their chance of metastasizing by evoking a systemic inflammatory cascade in mouse models of spontaneous breast cancer metastasis. We mechanistically demonstrate that interleukin (IL)-1β elicits IL-17 expression from gamma delta (γδ) T cells, resulting in systemic, granulocyte colony-stimulating factor (G-CSF)-dependent expansion and polarization of neutrophils in mice bearing mammary tumours. Tumour-induced neutrophils acquire the ability to suppress cytotoxic T lymphocytes carrying the CD8 antigen, which limit the establishment of metastases. Neutralization of IL-17 or G-CSF and absence of γδ T cells prevents neutrophil accumulation and downregulates the T-cell-suppressive phenotype of neutrophils. Moreover, the absence of γδ T cells or neutrophils profoundly reduces pulmonary and lymph node metastases without influencing primary tumour progression. Our data indicate that targeting this novel cancer-cell-initiated domino effect within the immune system--the γδ T cell/IL-17/neutrophil axis--represents a new strategy to inhibit metastatic disease. PMID:25822788

  19. 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.

    PubMed

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

    2010-08-25

    Recombinant T cell ligands (RTLs) ameliorate experimental autoimmune encephalomyelitis (EAE) in an antigen-specific manner. We evaluated effects of RTL401 (I-A(s) alpha1beta1+PLP-139-151) on splenocytes from SJL/J mice with EAE to study RTL-T cell tolerance-inducing mechanisms. RTLs bound to B, macrophages and DCs, through RTL-MHC-alpha1beta1 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 RTL- incubated splenocytes to transfer EAE was likely mediated through macrophages/DC, since B cells were unnecessary for RTL treatment of EAE. These results demonstrate a novel pathway of T cell regulation by RTL-bound APCs. PMID:20546940

  20. 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

    PubMed Central

    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 RTL- incubated splenocytes to transfer EAE was likely mediated through macrophages/DC, since B cells were unnecessary for RTL treatment of EAE. These results demonstrate novel pathway of T cell regulation by RTL bound APCs. PMID:20546940

  1. Characterization of palytoxin binding to HaCaT cells using a monoclonal anti-palytoxin antibody.

    PubMed

    Pelin, Marco; Boscolo, Sabrina; Poli, Mark; Sosa, Silvio; Tubaro, Aurelia; Florio, Chiara

    2013-03-01

    Palytoxin (PLTX) is the reference compound for a group of potent marine biotoxins, for which the molecular target is Na+/K+-ATPase. Indeed, ouabain (OUA), a potent blocker of the pump, is used to inhibit some PLTX effects in vitro. However, in an effort to explain incomplete inhibition of PLTX cytotoxicity, some studies suggest the possibility of two different binding sites on Na+/K+-ATPase. Hence, this study was performed to characterize PLTX binding to intact HaCaT keratinocytes and to investigate the ability of OUA to compete for this binding. PLTX binding to HaCaT cells was demonstrated by immunocytochemical analysis after 10 min exposure. An anti-PLTX monoclonal antibody-based ELISA showed that the binding was saturable and reversible, with a K(d) of 3 × 10-10 M. However, kinetic experiments revealed that PLTX binding dissociation was incomplete, suggesting an additional, OUA-insensitive, PLTX binding site. Competitive experiments suggested that OUA acts as a negative allosteric modulator against high PLTX concentrations (0.3-1.0 × 10-7 M) and possibly as a non-competitive antagonist against low PLTX concentrations (0.1-3.0 × 10-9 M). Antagonism was supported by PLTX cytotoxicity inhibition at OUA concentrations that displaced PLTX binding (1 × 10-5 M). However, this inhibition was incomplete, supporting the existence of both OUA-sensitive and -insensitive PLTX binding sites. PMID:23442788

  2. Non-small-cell lung cancer-induced immunosuppression by increased human regulatory T cells via Foxp3 promoter demethylation.

    PubMed

    Ke, Xing; Zhang, Shuping; Xu, Jian; Liu, Genyan; Zhang, Lixia; Xie, Erfu; Gao, Li; Li, Daqian; Sun, Ruihong; Wang, Fang; Pan, Shiyang

    2016-05-01

    Patients with non-small-cell lung cancer (NSCLC) have immune defects that are poorly understood. Forkhead box protein P3 (Foxp3) is crucial for immunosuppression by CD4(+) regulatory T cells (Tregs). It is not well known how NSCLC induces Foxp3 expression and causes immunosuppression in tumor-bearing patients. Our study found a higher percentage of CD4(+) Tregs in the peripheral blood of NSCLC compared with healthy donors. NSCLC patients showed demethylation of eight CpG sites within the Foxp3 promoter with methylation ratios negatively correlated with CD4(+)CD25(+)Foxp3(+) T levels. Foxp3 expression in CD4(+) Tregs was directly regulated by Foxp3 promoter demethylation and was involved in immunosuppression by NSCLC. To verify the effect of tumor cells on the phenotype and function of CD4(+) Tregs, we established a coculture system using NSCLC cell line and healthy CD4(+) T cells and showed that SPC-A1 induced IL-10 and TGF-β1 secretion by affecting the function of CD4(+) Tregs. The activity of DNA methyltransferases from CD4(+) T was decreased during this process. Furthermore, eight CpG sites within the Foxp3 promoter also appeared to have undergone demethylation. Foxp3 is highly expressed in CD4(+) T cells, and this may be caused by gene promoter demethylation. These induced Tregs are highly immunosuppressive and dramatically inhibit the proliferative activity of naïve CD4(+) T cells. Our study provides one possible mechanism describing Foxp3 promoter demethylation changes by which NSCLC down-regulates immune responses and contributes to tumor progression. Foxp3 represents an important target for NSCLC anti-tumor immunotherapy. PMID:27000869

  3. CD4+ T Cells Promote Antibody Production but Not Sustained Affinity Maturation during Borrelia burgdorferi Infection

    PubMed Central

    Elsner, Rebecca A.; Hastey, Christine J.

    2014-01-01

    CD4 T cells are crucial for enhancing B cell-mediated immunity, supporting the induction of high-affinity, class-switched antibody responses, long-lived plasma cells, and memory B cells. Previous studies showed that the immune response to Borrelia burgdorferi appears to lack robust T-dependent B cell responses, as neither long-lived plasma cells nor memory B cells form for months after infection, and nonswitched IgM antibodies are produced continuously during this chronic disease. These data prompted us to evaluate the induction and functionality of B. burgdorferi infection-induced CD4 TFH cells. We report that CD4 T cells were effectively primed and TFH cells induced after B. burgdorferi infection. These CD4 T cells contributed to the control of B. burgdorferi burden and supported the induction of B. burgdorferi-specific IgG responses. However, while affinity maturation of antibodies against a prototypic T-dependent B. burgdorferi protein, Arthritis-related protein (Arp), were initiated, these increases were reversed later, coinciding with the previously observed involution of germinal centers. The cessation of affinity maturation was not due to the appearance of inhibitory or exhausted CD4 T cells or a strong induction of regulatory T cells. In vitro T-B cocultures demonstrated that T cells isolated from B. burgdorferi-infected but not B. burgdorferi-immunized mice supported the rapid differentiation of B cells into antibody-secreting plasma cells rather than continued proliferation, mirroring the induction of rapid short-lived instead of long-lived T-dependent antibody responses in vivo. The data further suggest that B. burgdorferi infection drives the humoral response away from protective, high-affinity, and long-lived antibody responses and toward the rapid induction of strongly induced, short-lived antibodies of limited efficacy. PMID:25312948

  4. HIRF: a novel nuclear factor that binds to the human T-cell leukemia virus type I internal regulatory element (HIRE).

    PubMed

    Ariumi, Y; Copeland, T D; Nosaka, T; Hatanaka, M

    1997-04-01

    The transcription of human T-cell leukemia virus type I (HTLV-I) provirus starts from a promoter located in the 5' long terminal repeat (LTR). We have identified a second promoter at the 3' end of the pol gene. This internal promoter expresses the Tax transactivator protein, but does not require Tax for its activity. Furthermore, we have found the novel enhancer motif AGTTCTGCCC, which are located near the initiation site. We have named the sequence HIRE (HTLV-I internal regulatory element). The HIRE binding protein is a ubiquitous protein. We purified this protein from the HTLV-I producing cell line MT-2 cells by DNA affinity chromatography. SDS-PAGE analysis revealed four major bands (70, 85, 115 and more than 200 kDa) and some minor bands on the gel. We renatured each major protein and showed the 70 and 115 kDa proteins bind to DNA, although the 115 kDa protein seemed to bind nonspecifically. We have designated these components as HIRF (HTLV-I internal regulatory factor). PMID:9209287

  5. Glycan modification of antigen alters its intracellular routing in dendritic cells, promoting priming of T cells

    PubMed Central

    Streng-Ouwehand, Ingeborg; Ho, Nataschja I; Litjens, Manja; Kalay, Hakan; Boks, Martine Annemarie; Cornelissen, Lenneke AM; Kaur Singh, Satwinder; Saeland, Eirikur; Garcia-Vallejo, Juan J; Ossendorp, Ferry A; Unger, Wendy WJ; van Kooyk, Yvette

    2016-01-01

    Antigen uptake by dendritic cells and intracellular routing of antigens to specific compartments is regulated by C-type lectin receptors that recognize glycan structures. We show that the modification of Ovalbumin (OVA) with the glycan-structure LewisX (LeX) re-directs OVA to the C-type lectin receptor MGL1. LeX-modification of OVA favored Th1 skewing of CD4+ T cells and enhanced cross-priming of CD8+ T cells. While cross-presentation of native OVA requires high antigen dose and TLR stimuli, LeX modification reduces the required amount 100-fold and obviates its dependence on TLR signaling. The OVA-LeX-induced enhancement of T cell cross-priming is MGL1-dependent as shown by reduced CD8+ effector T cell frequencies in MGL1-deficient mice. Moreover, MGL1-mediated cross-presentation of OVA-LeX neither required TAP-transporters nor Cathepsin-S and was still observed after prolonged intracellular storage of antigen in Rab11+LAMP1+ compartments. We conclude that controlled neo-glycosylation of antigens can crucially influence intracellular routing of antigens, the nature and strength of immune responses and should be considered for optimizing current vaccination strategies. DOI: http://dx.doi.org/10.7554/eLife.11765.001 PMID:26999763

  6. Tumor-associated macrophages promote tumor cell proliferation in nasopharyngeal NK/T-cell lymphoma

    PubMed Central

    Liu, Yixiong; Fan, Linni; Wang, Yingmei; Li, Peifeng; Zhu, Jin; Wang, Lu; Zhang, Weichen; Zhang, Yuehua; Huang, Gaosheng

    2014-01-01

    Objective: To explore the relationship between the number of tumor-associated macrophages (TAMs) and proliferative activity of tumor cells and the relationship between two macrophage biomarkers CD68 and CD163 in nasopharyngeal NK/T-cell lymphoma. Methods: Immunohistochemistry was used to reconfirm the diagnosis of nasal NK/T-cell lymphoma and detect the numbers of TAMs and the ki-67 label index of the tumor cells in all 31 cases. In addition, 12 cases of inflammatory cases were collected as controls, for which the immunostaining of CD68 and CD163 were done as well. Then staining results were analyzed with Pearson correlation and t test. Results: The number of TAMs was positively correlated with tumor proliferative activity (P = 0.024) in nasopharyngeal NK/T-cell lymphoma. The expression of CD68 and CD163 was closely related (P = 0.009), and the positive rate of CD68 was generally higher than CD163, however there is no statistical significance. Conclusion: The increase in numbers of TAMs in nasopharyngeal NK/T-cell lymphoma is related to higher proliferative index, indicating the TAMs play an important role in tumor proliferation. Meanwhile both CD68 and CD163 might be the markers for TAMs but CD163 would be the better one. PMID:25337185

  7. Pediatric T-cell lymphoblastic leukemia evolves into relapse by clonal selection, acquisition of mutations and promoter hypomethylation

    PubMed Central

    Kunz, Joachim B.; Rausch, Tobias; Bandapalli, Obul R.; Eilers, Juliane; Pechanska, Paulina; Schuessele, Stephanie; Assenov, Yassen; Stütz, Adrian M.; Kirschner-Schwabe, Renate; Hof, Jana; Eckert, Cornelia; von Stackelberg, Arend; Schrappe, Martin; Stanulla, Martin; Koehler, Rolf; Avigad, Smadar; Elitzur, Sarah; Handgretinger, Rupert; Benes, Vladimir; Weischenfeldt, Joachim; Korbel, Jan O.; Muckenthaler, Martina U.; Kulozik, Andreas E.

    2015-01-01

    Relapsed precursor T-cell acute lymphoblastic leukemia is characterized by resistance against chemotherapy and is frequently fatal. We aimed at understanding the molecular mechanisms resulting in relapse of T-cell acute lymphoblastic leukemia and analyzed 13 patients at first diagnosis, remission and relapse by whole exome sequencing, targeted ultra-deep sequencing, multiplex ligation dependent probe amplification and DNA methylation array. Compared to primary T-cell acute lymphoblastic leukemia, in relapse the number of single nucleotide variants and small insertions and deletions approximately doubled from 11.5 to 26. Targeted ultra-deep sequencing sensitively detected subclones that were selected for in relapse. The mutational pattern defined two types of relapses. While both are characterized by selection of subclones and acquisition of novel mutations, ‘type 1’ relapse derives from the primary leukemia whereas ‘type 2’ relapse originates from a common pre-leukemic ancestor. Relapse-specific changes included activation of the nucleotidase NT5C2 resulting in resistance to chemotherapy and mutations of epigenetic modulators, exemplified by SUZ12, WHSC1 and SMARCA4. While mutations present in primary leukemia and in relapse were enriched for known drivers of leukemia, relapse-specific changes revealed an association with general cancer-promoting mechanisms. This study thus identifies mechanisms that drive progression of pediatric T-cell acute lymphoblastic leukemia to relapse and may explain the characteristic treatment resistance of this condition. PMID:26294725

  8. Pediatric T-cell lymphoblastic leukemia evolves into relapse by clonal selection, acquisition of mutations and promoter hypomethylation.

    PubMed

    Kunz, Joachim B; Rausch, Tobias; Bandapalli, Obul R; Eilers, Juliane; Pechanska, Paulina; Schuessele, Stephanie; Assenov, Yassen; Stütz, Adrian M; Kirschner-Schwabe, Renate; Hof, Jana; Eckert, Cornelia; von Stackelberg, Arend; Schrappe, Martin; Stanulla, Martin; Koehler, Rolf; Avigad, Smadar; Elitzur, Sarah; Handgretinger, Rupert; Benes, Vladimir; Weischenfeldt, Joachim; Korbel, Jan O; Muckenthaler, Martina U; Kulozik, Andreas E

    2015-11-01

    Relapsed precursor T-cell acute lymphoblastic leukemia is characterized by resistance against chemotherapy and is frequently fatal. We aimed at understanding the molecular mechanisms resulting in relapse of T-cell acute lymphoblastic leukemia and analyzed 13 patients at first diagnosis, remission and relapse by whole exome sequencing, targeted ultra-deep sequencing, multiplex ligation dependent probe amplification and DNA methylation array. Compared to primary T-cell acute lymphoblastic leukemia, in relapse the number of single nucleotide variants and small insertions and deletions approximately doubled from 11.5 to 26. Targeted ultra-deep sequencing sensitively detected subclones that were selected for in relapse. The mutational pattern defined two types of relapses. While both are characterized by selection of subclones and acquisition of novel mutations, 'type 1' relapse derives from the primary leukemia whereas 'type 2' relapse originates from a common pre-leukemic ancestor. Relapse-specific changes included activation of the nucleotidase NT5C2 resulting in resistance to chemotherapy and mutations of epigenetic modulators, exemplified by SUZ12, WHSC1 and SMARCA4. While mutations present in primary leukemia and in relapse were enriched for known drivers of leukemia, relapse-specific changes revealed an association with general cancer-promoting mechanisms. This study thus identifies mechanisms that drive progression of pediatric T-cell acute lymphoblastic leukemia to relapse and may explain the characteristic treatment resistance of this condition. PMID:26294725

  9. Vitamin D Regulates Cytokine Patterns Secreted by Dendritic Cells to Promote Differentiation of IL-22-Producing T Cells

    PubMed Central

    Sommer, Andrea; Fabri, Mario

    2015-01-01

    One central mechanism, by which vitamin D regulates human immune responses, is the direct modulation of dendritic cells (DCs). However, the effect of vitamin D on several key DC functions, such as the secretion of central inflammatory cytokines, remains controversial. Moreover, whether vitamin D treatment of DCs regulates their ability to promote differentiation of IL-17-/IL-22-producing T cell subsets, such as Th17 and Th22 cell, is not known. Here, we report that vitamin D treatment during differentiation of monocytes into DCs markedly enhanced their ability to secrete TNF-α, IL-6, IL-1β and IL-23. Cytokines secreted by vitamin D-treated DC were significantly more potent in driving differentiation of IL-22-producing T cells, but not IL-17-producing T cells, as compared to secreted cytokines of not-vitamin D-treated DCs. Finally, we found that the differentiation of IL-22-producing T cells mediated by supernatants of vitamin D-treated DCs was dependent on TNF-α IL-6 and IL-23. In summary, our study suggests a novel role of vitamin D in regulating DC-mediated immune responses in humans. PMID:26107738

  10. PD-1 alters T-cell metabolic reprogramming by inhibiting glycolysis and promoting lipolysis and fatty acid oxidation

    PubMed Central

    Patsoukis, Nikolaos; Bardhan, Kankana; Chatterjee, Pranam; Sari, Duygu; Liu, Bianling; Bell, Lauren N.; Karoly, Edward D.; Freeman, Gordon J.; Petkova, Victoria; Seth, Pankaj; Li, Lequn; Boussiotis, Vassiliki A.

    2015-01-01

    During activation, T cells undergo metabolic reprogramming, which imprints distinct functional fates. We determined that on PD-1 ligation, activated T cells are unable to engage in glycolysis or amino acid metabolism but have an increased rate of fatty acid β-oxidation (FAO). PD-1 promotes FAO of endogenous lipids by increasing expression of CPT1A, and inducing lipolysis as indicated by elevation of the lipase ATGL, the lipolysis marker glycerol and release of fatty acids. Conversely, CTLA-4 inhibits glycolysis without augmenting FAO, suggesting that CTLA-4 sustains the metabolic profile of non-activated cells. Because T cells utilize glycolysis during differentiation to effectors, our findings reveal a metabolic mechanism responsible for PD-1-mediated blockade of T-effector cell differentiation. The enhancement of FAO provides a mechanistic explanation for the longevity of T cells receiving PD-1 signals in patients with chronic infections and cancer, and for their capacity to be reinvigorated by PD-1 blockade. PMID:25809635

  11. Molecular analysis of tumor-promoting CD8+ T cells in two-stage cutaneous chemical carcinogenesis.

    PubMed

    Kwong, Bernice Y; Roberts, Scott J; Silberzahn, Tobias; Filler, Renata B; Neustadter, Jason H; Galan, Anjela; Reddy, Swapna; Lin, William M; Ellis, Peter D; Langford, Cordelia F; Hayday, Adrian C; Girardi, Michael

    2010-06-01

    T-pro are tumor-infiltrating TCRalphabeta(+)CD8(+) cells of reduced cytotoxic potential that promote experimental two-stage chemical cutaneous carcinogenesis. Toward understanding their mechanism of action, this study uses whole-genome expression analysis to compare T-pro with systemic CD8(+) T cells from multiple groups of tumor-bearing mice. T-pro show an overt T helper 17-like profile (high retinoic acid-related orphan receptor-(ROR)gammat, IL-17A, IL-17F; low T-bet and eomesodermin), regulatory potential (high FoxP3, IL-10, Tim-3), and transcripts encoding epithelial growth factors (amphiregulin, Gro-1, Gro-2). Tricolor flow cytometry subsequently confirmed the presence of TCRbeta(+) CD8(+) IL-17(+) T cells among tumor-infiltrating lymphocytes (TILs). Moreover, a time-course analysis of independent TIL isolates from papillomas versus carcinomas exposed a clear association of the "T-pro phenotype" with malignant progression. This molecular characterization of T-pro builds a foundation for elucidating the contributions of inflammation to cutaneous carcinogenesis, and may provide useful biomarkers for cancer immunotherapy in which the widely advocated use of tumor-specific CD8(+) cytolytic T cells should perhaps accommodate the cells' potential corruption toward the T-pro phenotype. The data are also likely germane to psoriasis, in which the epidermis may be infiltrated by CD8(+) IL-17-producing T cells. PMID:19924136

  12. Characterization of a Putative Receptor Binding Surface on Skint-1, a Critical Determinant of Dendritic Epidermal T Cell Selection*

    PubMed Central

    Salim, Mahboob; Knowles, Timothy J.; Hart, Rosie; Mohammed, Fiyaz; Woodward, Martin J.; Willcox, Carrie R.; Overduin, Michael; Hayday, Adrian C.; Willcox, Benjamin E.

    2016-01-01

    Dendritic epidermal T cells (DETC) form a skin-resident γδ T cell population that makes key contributions to cutaneous immune stress surveillance, including non-redundant contributions to protection from cutaneous carcinogens. How DETC become uniquely associated with the epidermis was in large part solved by the identification of Skint-1, the prototypic member of a novel B7-related multigene family. Expressed only by thymic epithelial cells and epidermal keratinocytes, Skint-1 drives specifically the development of DETC progenitors, making it the first clear candidate for a selecting ligand for non-MHC/CD1-restricted T cells. However, the molecular mechanisms underpinning Skint-1 activity are unresolved. Here, we provide evidence that DETC selection requires Skint-1 expression on the surface of thymic epithelial cells, and depends upon specific residues on the CDR3-like loop within the membrane-distal variable domain of Skint-1 (Skint-1 DV). Nuclear magnetic resonance of Skint-1 DV revealed a core tertiary structure conserved across the Skint family, but a highly distinct surface charge distribution, possibly explaining its unique function. Crucially, the CDR3-like loop formed an electrostatically distinct surface, featuring key charged and hydrophobic solvent-exposed residues, at the membrane-distal tip of DV. These results provide the first structural insights into the Skint family, identifying a putative receptor binding surface that directly implicates Skint-1 in receptor-ligand interactions crucial for DETC selection. PMID:26917727

  13. Characterization of a Putative Receptor Binding Surface on Skint-1, a Critical Determinant of Dendritic Epidermal T Cell Selection.

    PubMed

    Salim, Mahboob; Knowles, Timothy J; Hart, Rosie; Mohammed, Fiyaz; Woodward, Martin J; Willcox, Carrie R; Overduin, Michael; Hayday, Adrian C; Willcox, Benjamin E

    2016-04-22

    Dendritic epidermal T cells (DETC) form a skin-resident γδ T cell population that makes key contributions to cutaneous immune stress surveillance, including non-redundant contributions to protection from cutaneous carcinogens. How DETC become uniquely associated with the epidermis was in large part solved by the identification of Skint-1, the prototypic member of a novel B7-related multigene family. Expressed only by thymic epithelial cells and epidermal keratinocytes, Skint-1 drives specifically the development of DETC progenitors, making it the first clear candidate for a selecting ligand for non-MHC/CD1-restricted T cells. However, the molecular mechanisms underpinning Skint-1 activity are unresolved. Here, we provide evidence that DETC selection requires Skint-1 expression on the surface of thymic epithelial cells, and depends upon specific residues on the CDR3-like loop within the membrane-distal variable domain of Skint-1 (Skint-1 DV). Nuclear magnetic resonance of Skint-1 DV revealed a core tertiary structure conserved across the Skint family, but a highly distinct surface charge distribution, possibly explaining its unique function. Crucially, the CDR3-like loop formed an electrostatically distinct surface, featuring key charged and hydrophobic solvent-exposed residues, at the membrane-distal tip of DV. These results provide the first structural insights into the Skint family, identifying a putative receptor binding surface that directly implicates Skint-1 in receptor-ligand interactions crucial for DETC selection. PMID:26917727

  14. A77 1726, the active metabolite of leflunomide, attenuates lupus nephritis by promoting the development of regulatory T cells and inhibiting IL-17-producing double negative T cells.

    PubMed

    Qiao, Guilin; Yang, Lifen; Li, Zhenping; Williams, James W; Zhang, Jian

    2015-04-01

    Lupus nephritis (LN) is a challenging problem that affects 50% of patients with systemic lupus erythematosus (SLE) without effective therapy. Here, we report that A77 1726, the active metabolite of leflunomide, effectively inhibits development of LN and attenuates the generalized autoimmune features. A77 1726 suppresses the expansion of double negative (DN) T cells, and inhibits T and B cell activation. Intriguingly, A77 1726 treatment significantly increases CD4(+)Foxp3(+) regulatory T cells but suppresses potential "pathogenic" IL-17-producing DN T cells in lymph nodes. In vitro experiment shows that A77 1726 potentiates the conversion of naive CD4(+)CD25(-) T cells into CD4(+)CD25(+)Foxp3(+) inducible regulatory T cells (iTregs) by inhibiting Akt. Taken together, our data indicate that the therapeutic effects of A77 1726 in murine LN are mediated, at least in part, by augmenting iTregs which suppress pathogenic IL-17-producing DN T cells through an Akt-dependent mechanism. PMID:25638413

  15. CADM1 Interacts with Tiam1 and Promotes Invasive Phenotype of Human T-cell Leukemia Virus Type I-transformed Cells and Adult T-cell Leukemia Cells*

    PubMed Central

    Masuda, Mari; Maruyama, Tomoko; Ohta, Tsutomu; Ito, Akihiko; Hayashi, Tomayoshi; Tsukasaki, Kunihiko; Kamihira, Shimeru; Yamaoka, Shoji; Hoshino, Hiroo; Yoshida, Teruhiko; Watanabe, Toshiki; Stanbridge, Eric J.; Murakami, Yoshinori

    2010-01-01

    CADM1 encodes a multifunctional immunoglobulin-like cell adhesion molecule whose cytoplasmic domain contains a type II PSD95/Dlg/ZO-1 (PDZ)-binding motif (BM) for associating with other intracellular proteins. Although CADM1 lacks expression in T lymphocytes of healthy individuals, it is overexpressed in adult T-cell leukemia-lymphoma (ATL) cells. It has been suggested that the expression of CADM1 protein promotes infiltration of leukemic cells into various organs and tissues, which is one of the frequent clinical manifestations of ATL. Amino acid sequence alignment revealed that Tiam1 (T-lymphoma invasion and metastasis 1), a Rac-specific guanine nucleotide exchange factor, has a type II PDZ domain similar to those of membrane-associated guanylate kinase homologs (MAGUKs) that are known to bind to the PDZ-BM of CADM1. In this study, we demonstrated that the cytoplasmic domain of CADM1 directly interacted with the PDZ domain of Tiam1 and induced formation of lamellipodia through Rac activation in HTLV-I-transformed cell lines as well as ATL cell lines. Our results indicate that Tiam1 integrates signals from CADM1 to regulate the actin cytoskeleton through Rac activation, which may lead to tissue infiltration of leukemic cells in ATL patients. PMID:20215110

  16. Themis1 enhances T cell receptor signaling during thymocyte development by promoting Vav1 activity and Grb2 stability.

    PubMed

    Zvezdova, Ekaterina; Mikolajczak, Judith; Garreau, Anne; Marcellin, Marlène; Rigal, Lise; Lee, Jan; Choi, Seeyoung; Blaize, Gaëtan; Argenty, Jérémy; Familiades, Julien; Li, Liqi; Gonzalez de Peredo, Anne; Burlet-Schiltz, Odile; Love, Paul E; Lesourne, Renaud

    2016-01-01

    The T cell signaling protein Themis1 is essential for the positive and negative selection of thymocytes in the thymus. Although the developmental defect that results from the loss of Themis1 suggests that it enhances T cell receptor (TCR) signaling, Themis1 also recruits Src homology 2 domain-containing phosphatase-1 (SHP-1) to the vicinity of TCR signaling complexes, suggesting that it has an inhibitory role in TCR signaling. We used TCR signaling reporter mice and quantitative proteomics to explore the role of Themis1 in developing T cells. We found that Themis1 acted mostly as a positive regulator of TCR signaling in vivo when receptors were activated by positively selecting ligands. Proteomic analysis of the Themis1 interactome identified SHP-1, the TCR-associated adaptor protein Grb2, and the guanine nucleotide exchange factor Vav1 as the principal interacting partners of Themis1 in isolated mouse thymocytes. Analysis of TCR signaling in Themis1-deficient and Themis1-overexpressing mouse thymocytes demonstrated that Themis1 promoted Vav1 activity both in vitro and in vivo. The reduced activity of Vav1 and the impaired T cell development in Themis1(-/-) mice were due in part to increased degradation of Grb2, which suggests that Themis1 is required to maintain the steady-state abundance of Grb2 in thymocytes. Together, these data suggest that Themis1 acts as a positive regulator of TCR signaling in developing T cells, and identify a mechanism by which Themis1 regulates thymic selection. PMID:27188442

  17. CD4+ T cells promote the pathogenesis of Staphylococcus aureus pneumonia.

    PubMed

    Parker, Dane; Ryan, Chanelle L; Alonzo, Francis; Torres, Victor J; Planet, Paul J; Prince, Alice S

    2015-03-01

    We postulated that the activation of proinflammatory signaling by methicillin-resistant Staphylococcus aureus (MRSA) strain USA300 is a major factor in the pathogenesis of severe pneumonia and a target for immunomodulation. Local activation of T cells in the lung was a conserved feature of multiple strains of S. aureus, in addition to USA300. The pattern of Vβ chain activation was consistent with known superantigens, but deletion of SelX or SEK and SEQ was not sufficient to prevent T-cell activation, indicating the participation of multiple genes. Using Rag2(-/-), Cd4(-/-), and Cd28(-/-) mice, we observed significantly improved clearance of MRSA from the airways and decreased lung pathology, compared with findings for wild-type controls. The improved outcome correlated with decreased production of proinflammatory cytokines (tumor necrosis factor, KC, interleukin 6, and interleukin 1β). Our data suggest that T-cell-mediated hypercytokinemia induced by infection with MRSA strain USA300 contributes to pathogenesis and may be a therapeutic target for improving outcomes of this common infection in a clinical setting. PMID:25240171

  18. Galectin-8 Ameliorates Murine Autoimmune Ocular Pathology and Promotes a Regulatory T Cell Response

    PubMed Central

    Sampson, James F.; Hasegawa, Eiichi; Mulki, Lama; Suryawanshi, Amol; Jiang, Shuhong; Chen, Wei-Sheng; Rabinovich, Gabriel A.; Connor, Kip M.; Panjwani, Noorjahan

    2015-01-01

    Galectins have emerged as potent immunoregulatory agents that control chronic inflammation through distinct mechanisms. Here, we report that treatment with Galectin-8 (Gal-8), a tandem-repeat member of the galectin family, reduces retinal pathology and prevents photoreceptor cell damage in a murine model of experimental autoimmune uveitis. Gal-8 treatment increased the number of regulatory T cells (Treg) in both the draining lymph node (dLN) and the inflamed retina. Moreover, a greater percentage of Treg cells in the dLN and retina of Gal-8 treated animals expressed the inhibitory coreceptor cytotoxic T lymphocyte antigen (CTLA)-4, the immunosuppressive cytokine IL-10, and the tissue-homing integrin CD103. Treg cells in the retina of Gal-8-treated mice were primarily inducible Treg cells that lack the expression of neuropilin-1. In addition, Gal-8 treatment blunted production of inflammatory cytokines by retinal T helper type (TH) 1 and TH17 cells. The effect of Gal-8 on T cell differentiation and/or function was specific for tissues undergoing an active immune response, as Gal-8 treatment had no effect on T cell populations in the spleen. Given the need for rational therapies for managing human uveitis, Gal-8 emerges as an attractive therapeutic candidate not only for treating retinal autoimmune diseases, but also for other TH1- and TH17-mediated inflammatory disorders. PMID:26126176

  19. A THEMIS:SHP1 complex promotes T-cell survival

    PubMed Central

    Paster, Wolfgang; Bruger, Annika M; Katsch, Kristin; Grégoire, Claude; Roncagalli, Romain; Fu, Guo; Gascoigne, Nicholas RJ; Nika, Konstantina; Cohnen, Andre; Feller, Stephan M; Simister, Philip C; Molder, Kelly C; Cordoba, Shaun-Paul; Dushek, Omer; Malissen, Bernard; Acuto, Oreste

    2015-01-01

    THEMIS is critical for conventional T-cell development, but its precise molecular function remains elusive. Here, we show that THEMIS constitutively associates with the phosphatases SHP1 and SHP2. This complex requires the adapter GRB2, which bridges SHP to THEMIS in a Tyr-phosphorylation-independent fashion. Rather, SHP1 and THEMIS engage with the N-SH3 and C-SH3 domains of GRB2, respectively, a configuration that allows GRB2-SH2 to recruit the complex onto LAT. Consistent with THEMIS-mediated recruitment of SHP to the TCR signalosome, THEMIS knock-down increased TCR-induced CD3-ζ phosphorylation, Erk activation and CD69 expression, but not LCK phosphorylation. This generalized TCR signalling increase led to augmented apoptosis, a phenotype mirrored by SHP1 knock-down. Remarkably, a KI mutation of LCK Ser59, previously suggested to be key in ERK-mediated resistance towards SHP1 negative feedback, did not affect TCR signalling nor ligand discrimination in vivo. Thus, the THEMIS:SHP complex dampens early TCR signalling by a previously unknown molecular mechanism that favours T-cell survival. We discuss possible implications of this mechanism in modulating TCR output signals towards conventional T-cell development and differentiation. PMID:25535246

  20. T cell-intrinsic ASC critically promotes TH17-mediated experimental autoimmune encephalomyelitis.

    PubMed

    Martin, Bradley N; Wang, Chenhui; Zhang, Cun-Jin; Kang, Zizhen; Gulen, Muhammet Fatih; Zepp, Jarod A; Zhao, Junjie; Bian, Guanglin; Do, Jeong-Su; Min, Booki; Pavicic, Paul G; El-Sanadi, Caroline; Fox, Paul L; Akitsu, Aoi; Iwakura, Yoichiro; Sarkar, Anasuya; Wewers, Mark D; Kaiser, William J; Mocarski, Edward S; Rothenberg, Marc E; Hise, Amy G; Dubyak, George R; Ransohoff, Richard M; Li, Xiaoxia

    2016-05-01

    Interleukin 1β (IL-1β) is critical for the in vivo survival, expansion and effector function of IL-17-producing helper T (TH17) cells during autoimmune responses, including experimental autoimmune encephalomyelitis (EAE). However, the spatiotemporal role and cellular source of IL-1β during EAE pathogenesis are poorly defined. In the present study, we uncovered a T cell-intrinsic inflammasome that drives IL-1β production during TH17-mediated EAE pathogenesis. Activation of T cell antigen receptors induced expression of pro-IL-1β, whereas ATP stimulation triggered T cell production of IL-1β via ASC-NLRP3-dependent caspase-8 activation. IL-1R was detected on TH17 cells but not on type 1 helper T (TH1) cells, and ATP-treated TH17 cells showed enhanced survival compared with ATP-treated TH1 cells, suggesting autocrine action of TH17-derived IL-1β. Together these data reveal a critical role for IL-1β produced by a TH17 cell-intrinsic ASC-NLRP3-caspase-8 inflammasome during inflammation of the central nervous system. PMID:26998763

  1. Epidermal Fatty Acid Binding Protein (E-FABP) Is Not Required for the Generation or Maintenance of Effector and Memory T Cells following Infection with Listeria monocytogenes.

    PubMed

    Li, Bing; Schmidt, Nathan W

    2016-01-01

    Following activation of naïve T cells there are dynamic changes in the metabolic pathways used by T cells to support both the energetic needs of the cell and the macromolecules required for growth and proliferation. Among other changes, lipid metabolism undergoes dynamic transitions between fatty acid oxidation and fatty acid synthesis as cells progress from naïve to effector and effector to memory T cells. The hydrophobic nature of lipids requires that they be bound to protein chaperones within a cell. Fatty acid binding proteins (FABPs) represent a large class of lipid chaperones, with epidermal FABP (E-FABP) expressed in T cells. The objective of this study was to determine the contribution of E-FABP in antigen-specific T cell responses. Following infection with Listeria monocytogenes, we observed similar clonal expansion, contraction and formation of memory CD8 T cells in WT and E-FABP-/- mice, which also exhibited similar phenotypic and functional characteristics. Analysis of Listeria-specific CD4 T cells also revealed no defect in the expansion, contraction, and formation of memory CD4 T cells in E-FABP-/- mice. These data demonstrate that E-FABP is dispensable for antigen-specific T cell responses following a bacterial infection. PMID:27588422

  2. Membrane binding mode of intrinsically disordered cytoplasmic domains of T cell receptor signaling subunits depends on lipid composition

    SciTech Connect

    Sigalov, Alexander B.; Hendricks, Gregory M.

    2009-11-13

    Intrinsically disordered cytoplasmic domains of T cell receptor (TCR) signaling subunits including {zeta}{sub cyt} and CD3{epsilon}{sub cyt} all contain one or more copies of an immunoreceptor tyrosine-based activation motif (ITAM), tyrosine residues of which are phosphorylated upon receptor triggering. Membrane binding-induced helical folding of {zeta}{sub cyt} and CD3{epsilon}{sub cyt} ITAMs is thought to control TCR activation. However, the question whether or not lipid binding of {zeta}{sub cyt} and CD3{epsilon}{sub cyt} is necessarily accompanied by a folding transition of ITAMs remains open. In this study, we investigate whether the membrane binding mechanisms of {zeta}{sub cyt} and CD3{epsilon}{sub cyt} depend on the membrane model used. Circular dichroic and fluorescence data indicate that binding of {zeta}{sub cyt} and CD3{epsilon}{sub cyt} to detergent micelles and unstable vesicles is accompanied by a disorder-to-order transition, whereas upon binding to stable vesicles these proteins remain unfolded. Using electron microscopy and dynamic light scattering, we show that upon protein binding, unstable vesicles fuse and rupture. In contrast, stable vesicles remain intact under these conditions. This suggests different membrane binding modes for {zeta}{sub cyt} and CD3{epsilon}{sub cyt} depending on the bilayer stability: (1) coupled binding and folding, and (2) binding without folding. These findings explain the long-standing puzzle in the literature and highlight the importance of the choice of an appropriate membrane model for protein-lipid interactions studies.

  3. Components of chicken egg white extract smaller than 3 kDa in size promote 293T cell proliferation.

    PubMed

    Ruan, Guang-Ping; Yao, Xiang; Wang, Jin-Xiang; Liu, Ju-Fen; Shu, Fan; Li, Zi-An; Pang, Rong-Qing; Pan, Xing-Hua

    2016-08-01

    We previously found that chicken egg white extract could promote cell survival and proliferation. In the present study, we further separated this extract into its components to identify those primarily responsible for promoting cell proliferation. Components of differing molecular weight were separated from chicken egg white extract by ultrafiltration and 293T cell cultures were supplemented with various concentrations. The effects on cell proliferation were subsequently determined by a CellTiter 96 Aqueous One Solution Cell Proliferation Assay kit (Promega). We demonstrate that components from chicken egg white smaller than 3 kDa in size are able to function as active ingredients promoting cellular proliferation. This discovery may identify a new and convenient additive for cell culture media to promote cell growth and proliferation. PMID:26541834

  4. [Lipopolysaccharide promotes the proliferation of CD4(+) T cells by modulating the survive and cytokine secretion of dendritic cells].

    PubMed

    Chen, Song; Lu, Lisha; Wang, Weiqiang; Xue, Ting; Yu, Juan; Sun, Zhina; Zhao, Chunxiao; Liao, Fang

    2016-09-01

    Objective To explore the mechanism modulating the function of dendritic cells (DCs) and promoting the T cell response by lipopolysaccharide (LPS). Methods Splenic DCs were purified with anti-CD11c immunomagnetic beads. After DCs were stimulated with LPS, the expressions of co-stimulatory molecules CD80 and CD86 on the DCs were detected by flow cytometry. The protein levels of pro-inflammatory cytokines interleukin 4 (IL-4), IL-5, IL-6, IL-12p40, IL-12p70 and tumor necrosis factor alpha (TNF-α) in the culture supernatant were measured by ELISA. The apoptotic levels of DCs which were labeled with annexinV-FITC/PI were determined by flow cytometry. The phosphorylation level of nuclear factor κB P65 (NF-κB P65) was assessed by phos-flow. The mRNA levels of variable genes in microarray were determined by real-time PCR. The proliferation of CD4(+) T cells which were co-cultured with OVA323-329-treated DCs was analyzed by flow cytometry. Results The purity of DCs reached over 93% after isolation. LPS up-regulated the expressions of CD80 and CD86 and enhanced DCs-mediated proliferation of CD4(+) T cells. In addition, LPS increased the protein levels of IL-12p40, TNF-α and IL-6, and inhibited the apoptosis of DCs through the NF-κB signaling pathway. Conclusion LPS could enhance DC-mediated proliferation of CD4(+) T cells by modulating the DCs survival and cytokine secretion. PMID:27609566

  5. Nude mice produce a T cell-derived antigen-binding factor that mediates the early component of delayed-type hypersensitivity.

    PubMed

    Herzog, W R; Meade, R; Pettinicchi, A; Ptak, W; Askenase, P W

    1989-03-15

    The elicitation of delayed-type hypersensitivity (DTH) reactions in mice is caused by the sequential action of two different T cells. An early-acting, DTH-initiating T cell produces an Ag-specific T cell factor, that is analogous to IgE antibody and initiates DTH by sensitizing the local tissues for release of the vasoactive amine serotonin. In picryl chloride or oxazolone contact sensitivity, this T cell factor is Ag-specific, but MHC unrestricted. We, therefore, hypothesized that DTH-initiating T cells are primitive T cells with Ag receptors that can bind Ag without MHC restriction. In order to characterize the origin of this DTH-initiating T cell and the conditions that are necessary for its development, we contact-sensitized various strains of immunodeficient mice. Surprisingly, we found that the early phase of DTH was present in athymic nude mice. In contrast, the early component of DTH was absent in mice with severe combined immunodeficiency. These mice lack T and B cells, but have NK cells. These findings suggested that the early component of DTH was not caused by NK cells, and was caused by cells belonging to a lineage from a rearranging gene family. The early component of DTH in nude mice was Ag specific, was caused by MHC unrestricted Thy-1+ T cells, and was mediated by Ag-binding, Ag-specific T cell factors. We found that DTH-initiating, T cell-derived, Ag-binding molecules from nude mice and normal CBA/J mice had the same functional properties. The early component of DTH was elicited in two different systems (contact sensitivity and SRBC-specific DTH) in two strains of nude mice (BALB/c athymic nudes and CByB6F1/J-nu) from two different suppliers, but not in BALB/c and athymic nudes from a third supplier. From these findings we concluded that DTH-initiating T cells, which produce IgE-like Ag-specific T cell factors, are present in some strains of athymic nude mice and thus are relatively thymic independent T cells. PMID:2466077

  6. An Engineered Switch in T Cell Receptor Specificity Leads to an Unusual but Functional Binding Geometry.

    PubMed

    Harris, Daniel T; Singh, Nishant K; Cai, Qi; Smith, Sheena N; Vander Kooi, Craig W; Procko, Erik; Kranz, David M; Baker, Brian M

    2016-07-01

    Utilizing a diverse binding site, T cell receptors (TCRs) specifically recognize a composite ligand comprised of a foreign peptide and a major histocompatibility complex protein (MHC). To help understand the determinants of TCR specificity, we studied a parental and engineered receptor whose peptide specificity had been switched via molecular evolution. Altered specificity was associated with a significant change in TCR-binding geometry, but this did not impact the ability of the TCR to signal in an antigen-specific manner. The determinants of binding and specificity were distributed among contact and non-contact residues in germline and hypervariable loops, and included disruption of key TCR-MHC interactions that bias αβ TCRs toward particular binding modes. Sequence-fitness landscapes identified additional mutations that further enhanced specificity. Our results demonstrate that TCR specificity arises from the distributed action of numerous sites throughout the interface, with significant implications for engineering therapeutic TCRs with novel and functional recognition properties. PMID:27238970

  7. Preselection Thymocytes Are More Sensitive to T Cell Receptor Stimulation Than Mature T Cells

    PubMed Central

    Davey, Gayle M.; Schober, Sonya L.; Endrizzi, Bart T.; Dutcher, Angela K.; Jameson, Stephen C.; Hogquist, Kristin A.

    1998-01-01

    During T cell development, thymocytes which are tolerant to self-peptides but reactive to foreign peptides are selected. The current model for thymocyte selection proposes that self-peptide–major histocompatibility complex (MHC) complexes that bind the T cell receptor with low affinity will promote positive selection while those with high affinity will result in negative selection. Upon thymocyte maturation, such low affinity self-peptide–MHC ligands no longer provoke a response, but foreign peptides can incidentally be high affinity ligands and can therefore stimulate T cells. For this model to work, thymocytes must be more sensitive to ligand than mature T cells. Contrary to this expectation, several groups have shown that thymocytes are less responsive than mature T cells to anti-T cell receptor for antigen (TCR)/CD3 mAb stimulation. Additionally, the lower TCR levels on thymocytes, compared with T cells, would potentially correlate with decreased thymocyte sensitivity. Here we compared preselection thymocytes and mature T cells for early activation events in response to peptide–MHC ligands. Remarkably, the preselection thymocytes were more responsive than mature T cells when stimulated with low affinity peptide variants, while both populations responded equally well to the antigenic peptide. This directly demonstrates the increased sensitivity of thymocytes compared with T cells for TCR engagement by peptide–MHC complexes. PMID:9815264

  8. Restricting Glutamine or Glutamine-Dependent Purine and Pyrimidine Syntheses Promotes Human T Cells with High FOXP3 Expression and Regulatory Properties.

    PubMed

    Metzler, Barbara; Gfeller, Patrick; Guinet, Elisabeth

    2016-05-01

    T cell subsets differ in their metabolic requirements, and further insight into such differences might be harnessed to selectively promote regulatory T cells (Tregs) for therapies in autoimmunity and transplantation. We found that Gln restriction during human T cell activation favored CD4 T cells with high expression of the Treg transcription factor FOXP3. This resulted from shrinking numbers and reduced proliferation of activated FOXP3(lo/-)CD4 T cells while FOXP3(hi)CD4 T cell numbers increased. This gain was abolished by blocking Gln synthetase, an enzyme that responds to Gln and purine/pyrimidine deficiencies. The shift toward FOXP3(hi)CD4 T cells under Gln restriction was recapitulated with inhibitors of Gln-dependent pyrimidine and purine syntheses that together closely mimicked declining cell numbers and cell cycles, and by small interfering RNA knockdown of the respective rate-limiting Gln-consuming enzymes CAD and PPAT. FOXP3(hi)-enriched CD25(hi)CD4 T cells from these cultures inhibited proliferation, but they also produced effector cytokines, including IL-17A. The latter was largely confined to CTLA-4(hi)-expressing FOXP3(hi)-enriched CD25(hi)CD4 T cells that suppressed proliferation more weakly than did CTLA-4(lo/-)CD25(hi)FOXP3(hi)-enriched T cells. A causal link between high IL-17A production and impaired suppression of proliferation could not be demonstrated, however. Collectively, these results reveal a Gln synthetase-dependent increase and resilience of FOXP3(hi) cells under Gln restriction, and they demonstrate that impaired Gln-dependent nucleotide synthesis promotes FOXP3(hi) cells with regulator properties. It remains to be investigated to what extent the concomitant retention of IL-17A-producing CD4 T cells may limit the therapeutic potential of Tregs enriched through targeting these pathways in vivo. PMID:27022197

  9. Leptin directly promotes T-cell glycolytic metabolism to drive effector T-cell differentiation in a mouse model of autoimmunity.

    PubMed

    Gerriets, Valerie A; Danzaki, Keiko; Kishton, Rigel J; Eisner, William; Nichols, Amanda G; Saucillo, Donte C; Shinohara, Mari L; MacIver, Nancie J

    2016-08-01

    Upon activation, T cells require energy for growth, proliferation, and function. Effector T (Teff) cells, such as Th1 and Th17 cells, utilize high levels of glycolytic metabolism to fuel proliferation and function. In contrast, Treg cells require oxidative metabolism to fuel suppressive function. It remains unknown how Teff/Treg-cell metabolism is altered when nutrients are limited and leptin levels are low. We therefore examined the role of malnutrition and associated hypoleptinemia on Teff versus Treg cells. We found that both malnutrition-associated hypoleptinemia and T cell-specific leptin receptor knockout suppressed Teff-cell number, function, and glucose metabolism, but did not alter Treg-cell metabolism or suppressive function. Using the autoimmune mouse model EAE, we confirmed that fasting-induced hypoleptinemia altered Teff-cell, but not Treg-cell, glucose metabolism, and function in vivo, leading to decreased disease severity. To explore potential mechanisms, we examined HIF-1α, a key regulator of Th17 differentiation and Teff-cell glucose metabolism, and found HIF-1α expression was decreased in T cell-specific leptin receptor knockout Th17 cells, and in Teff cells from fasted EAE mice, but was unchanged in Treg cells. Altogether, these data demonstrate a selective, cell-intrinsic requirement for leptin to upregulate glucose metabolism and maintain function in Teff, but not Treg cells. PMID:27222115

  10. Pleural mesothelial cells promote expansion of IL-17-producing CD8+ T cells in tuberculous pleural effusion.

    PubMed

    Li, X; Zhou, Q; Yang, W B; Xiong, X Z; Du, R H; Zhang, J C

    2013-05-01

    IL-17-producing CD8(+) T lymphocytes (Tc17 cells) have recently been detected in many cancers and autoimmune diseases. However, the possible implication of Tc17 cells in tuberculous pleural effusion remains unclarified. In this study, distribution and phenotypic features of Tc17 cells in both tuberculous pleural effusion (TPE) and peripheral blood from patients with tuberculosis were determined. The effects of proinflammatory cytokines and local accessory cells (pleural mesothelial cells) on Tc17 cell expansion were also explored. We found that TPE contained more Tc17 cells than the blood. Compared with IFN-γ-producing CD8(+) T cells, Tc17 cells displayed higher expression of chemokine receptors (CCRs) and lower expression of cytotoxic molecules. In particularly, Tc17 cells in TPE exhibited high expression levels of CCR6, which could migrate in response to CCL20. Furthermore, IL-1β, IL-6, IL-23, or their various combinations could promote Tc17 cell expansion from CD8(+) T cells, whereas the proliferative response of Tc17 cells to above cytokines was lower than that of Th17 cells. Pleural mesothelial cells (PMCs) were able to stimulate Tc17 cell expansion via cell contact in an IL-1β/IL-6/IL-23 independent fashion. Thus this study demonstrates that Tc17 cells marks a subset of non-cytotoxic, CCR6(+) CD8(+) T lymphocytes with low proliferative capacity. The overrepresentation of Tc17 cells in TPE may be due to Tc17 cell expansion stimulated by pleural proinflammatory cytokines and to recruitment of Tc17 cells from peripheral blood. Additionally, PMCs may promote the production of IL-17 by CD8(+) T cells at sites of TPE via cell-cell interactions. PMID:23299924

  11. A DNA-binding mutant of TAL1 cooperates with LMO2 to cause T cell leukemia in mice.

    PubMed

    Draheim, K M; Hermance, N; Yang, Y; Arous, E; Calvo, J; Kelliher, M A

    2011-03-10

    The most common translocation in childhood T-cell acute lymphoblastic leukemia (T-ALL) involves the LMO2 locus, resulting in ectopic expression of the LMO2 gene in human thymocytes. The LMO2 gene was also activated in patients with X-linked Severe Combined Immune Deficiency treated with gene therapy because of retroviral insertion in the LMO2 locus. The LMO2 insertions predisposed these children to T-ALL, yet how LMO2 contributes to T cell transformation remains unclear. The LIM (Lin 11, Isl-1, Mec-3) domain containing LMO2 protein regulates erythropoiesis as part of a large transcriptional complex consisting of LMO2, TAL1, E47, GATA1 and LDB1 that recognizes bipartite E-box-GATA1 sites on target genes. Similarly, a TAL1/E47/LMO2/LDB1 complex is observed in human T-ALL and Tal1 and Lmo2 expression in mice results in disease acceleration. To address the mechanism(s) of Tal1/Lmo2 synergy in leukemia, we generated Lmo2 transgenic mice and mated them with mice that express wild-type Tal1 or a DNA-binding mutant of TAL1. Tal1/Lmo2 and MutTAL1/Lmo2 bitransgenic mice exhibit perturbations in thymocyte development due to reduced E47/HEB transcriptional activity and develop leukemia with identical kinetics. These data demonstrate that the DNA-binding activity of Tal1 is not required to cooperate with Lmo2 to cause leukemia in mice and suggest that Lmo2 may cooperate with Tal1 to interfere with E47/HEB function(s). PMID:21057528

  12. CD226 ligation protects against EAE by promoting IL-10 expression via regulation of CD4+ T cell differentiation

    PubMed Central

    Chen, Kun; Zhang, Chunmei; Song, Chaojun; Fang, Liang; Xu, Zhuwei; Yang, Kun; Jin, Boquan; Wang, Qintao; Chen, Lihua

    2016-01-01

    Treatment targeting CD226 can ameliorate experimental autoimmune encephalomyelitis (EAE), the widely accepted model of MS. However, the mechanisms still need to be elucidated. Here we showed that CD226 blockage by anti-CD226 blocking mAb LeoA1 efficiently promoted IL-10 production in human peripheral blood monocytes (PBMC) or in mixed lymphocyte culture (MLC) system, significantly induced the CD4+IL-10+ T cell differentiation while suppressing the generation of Th1 and Th17. Furthermore, CD226 pAb administration in vivo reduced the onset of EAE in mice by promoting IL-10 production and regulating T cell differentiation. Concomitantly, the onset and severity of EAE were reduced and the serum IL-10 expression levels were increased in CD226 knockout mice than that in control mice when both received EAE induction. These novel findings confirmed that CD226 played a pivotal role in mediating autoimmune diseases such as EAE. Furthermore, to our knowledge, we show for the first time that IL-10 is an important contributor in the inhibitory effects of CD226 ligation on EAE. PMID:26942885

  13. Disparate Degrees of Hypervariable Loop Flexibility Control T-Cell Receptor Cross-Reactivity, Specificity, and Binding Mechanism

    SciTech Connect

    Scott, Daniel R.; Borbulevych, Oleg Y.; Piepenbrink, Kurt H.; Corcelli, Steven A.; Baker, Brian M.

    2012-06-19

    {alpha}{beta} T-cell receptors (TCRs) recognize multiple antigenic peptides bound and presented by major histocompatibility complex molecules. TCR cross-reactivity has been attributed in part to the flexibility of TCR complementarity-determining region (CDR) loops, yet there have been limited direct studies of loop dynamics to determine the extent of its role. Here we studied the flexibility of the binding loops of the {alpha}{beta} TCR A6 using crystallographic, spectroscopic, and computational methods. A significant role for flexibility in binding and cross-reactivity was indicated only for the CDR3{alpha} and CDR3{beta} hypervariable loops. Examination of the energy landscapes of these two loops indicated that CDR3{beta} possesses a broad, smooth energy landscape, leading to rapid sampling in the free TCR of a range of conformations compatible with different ligands. The landscape for CDR3{alpha} is more rugged, resulting in more limited conformational sampling that leads to specificity for a reduced set of peptides as well as the major histocompatibility complex protein. In addition to informing on the mechanisms of cross-reactivity and specificity, the energy landscapes of the two loops indicate a complex mechanism for TCR binding, incorporating elements of both conformational selection and induced fit in a manner that blends features of popular models for TCR recognition.

  14. Plasticity in the contribution of T cell receptor variable region residues to binding of peptide-HLA-A2 complexes

    PubMed Central

    Smith, Sheena N.; Sommermeyer, Daniel; Piepenbrink, Kurt H.; Blevins, Sydney J.; Bernhard, Helga; Uckert, Wolfgang; Baker, Brian M.; Kranz, David M.

    2013-01-01

    One hypothesis to account for MHC-restriction by T cell receptors (TCRs) holds that there are several evolutionary-conserved residues in TCR variable regions that contact MHC. While this ‘germline-codon’ hypothesis is supported by various lines of evidence, it has been difficult to test. The difficulty stems in part from the fact that TCRs exhibit low affinities for pep/MHC, thus limiting the range of binding energies that can be assigned to these key interactions using mutational analyses. To measure the magnitude of binding energies involved, here we used high-affinity TCRs engineered by mutagenesis of CDR3. The TCRs included a high-affinity, MART-1/HLA-A2-specific single-chain TCR and two other high-affinity TCRs that all contain the same Vα (HLA-A2), with different peptides and Vβ regions. Mutational analysis of residues in CDR1 and CDR2 of the three Vα2 regions showed the importance of the key ‘germline codon” residue Y51. However, two other proposed key residues showed significant differences among the TCRs in their relative contributions to binding. Using single-position, yeast-display libraries in two of the key residues, MART-1/HLA-A2 selections also revealed strong preferences for wild-type ‘germline codon’ residues, but several alternative residues could also accommodate binding and hence, MHC-restriction. Thus, although a single residue (Y51) could account for a proportion of the energy associated with positive selection (i.e. MHC-restriction), there is significant plasticity in requirements for particular side-chains in CDR1 and CDR2 and in their relative binding contributions among different TCRs. PMID:23954306

  15. T Cells Going Innate.

    PubMed

    Seyda, Midas; Elkhal, Abdallah; Quante, Markus; Falk, Christine S; Tullius, Stefan G

    2016-08-01

    Natural killer (NK) cell receptors (NKRs) play a crucial role in the homeostasis of antigen-experienced T cells. Indeed, prolonged antigen stimulation may induce changes in the receptor repertoire of T cells to a profile that features NKRs. Chronic antigen exposure, at the same time, has been shown to trigger the loss of costimulatory CD28 molecules with recently reported intensified antigen thresholds of antigen-experienced CD8(+) T cells. In transplantation, NKRs have been shown to assist allograft rejection in a CD28-independent fashion. We discuss here a role for CD28-negative T cells that have acquired the competency of the NKR machinery, potentially promoting allorecognition either through T cell receptor (TCR) crossreactivity or independently from TCR recognition. Collectively, NKRs can bring about innate-like T cells by providing alternative costimulatory pathways that gain relevance in chronic inflammation, potentially leading to resistance to CD28-targeting immunosuppressants. PMID:27402226

  16. Physical and functional interaction between the human T-cell lymphotropic virus type 1 Tax1 protein and the CCAAT binding protein NF-Y.

    PubMed Central

    Pise-Masison, C A; Dittmer, J; Clemens, K E; Brady, J N

    1997-01-01

    Tax1, a potent activator of human T-cell lymphotropic virus type 1 (HTLV-1) transcription, has been shown to modulate expression of many cellular genes. Tax1 does not bind DNA directly but regulates transcription through protein-protein interactions with sequence-specific transcription factors. Using the yeast two-hybrid system to screen for proteins which interact with Tax1, we isolated the B subunit of the CCAAT binding protein NF-Y from a HeLa cDNA library. The interaction of Tax1 with NF-YB was specific in that NF-YB did not interact with a variety of other transcription factors, including human immunodeficiency virus Tat, human papillomavirus E6, and Bicoid, or with the M7 (amino acids 29CP-AS) Tax1 mutant. However, NF-YB did interact with the C-terminal Tax1 mutants M22 (130TL-AS) and M47 (319LL-RS). We also show that in vitro-translated NF-YB specifically bound to a glutathione S-transferase-Tax1 fusion protein. Further, Tax1 coimmunoprecipitated with NF-Y from nuclear extracts of HTLV-1-transformed cells, providing evidence for in vivo interaction of Tax1 and NF-YB. We further demonstrate that Tax1 specifically activated the NF-Y-responsive DQbeta promoter, as well as a minimal promoter which contains only the Y-box element. In addition, mutation of the Y-box element alone abrogated Tax1-mediated activation. Taken together, these data indicate that Tax1 interacts with NF-Y through the B subunit and that this interaction results in activation of the major histocompatibility complex class II promoter. Through activation of this and other NF-Y driven promoters, the Tax1-NF-Y interaction may play a critical role in causing cellular transformation and HTLV-1 pathogenesis. PMID:9032250

  17. Relevance of regulatory T cell promotion of donor-specific tolerance in solid organ transplantation

    PubMed Central

    Sagoo, Pervinder; Lombardi, Giovanna; Lechler, Robert I.

    2012-01-01

    Current clinical strategies to control the alloimmune response after transplantation do not fully prevent induction of the immunological processes which lead to acute and chronic immune-mediated graft rejection, and as such the survival of a solid organ allograft is limited. Experimental research on naturally occurring CD4+CD25highFoxP3+ Regulatory T cells (Tregs) has indicated their potential to establish stable long-term graft acceptance, with the promise of providing a more effective therapy for transplant recipients. Current approaches for clinical use are based on the infusion of freshly isolated or ex vivo polyclonally expanded Tregs into graft recipients with an aim to redress the in vivo balance of T effector cells to Tregs. However mounting evidence suggests that regulation of donor-specific immunity may be central to achieving immunological tolerance. Therefore, the next stages in optimizing translation of Tregs to organ transplantation will be through the refinement and development of donor alloantigen-specific Treg therapy. The altering kinetics and intensity of alloantigen presentation pathways and alloimmune priming following transplantation may indeed influence the specificity of the Treg required and the timing or frequency at which it needs to be administered. Here we review and discuss the relevance of antigen-specific regulation of alloreactivity by Tregs in experimental and clinical studies of tolerance and explore the concept of delivering an optimal Treg for the induction and maintenance phases of achieving transplantation tolerance. PMID:22811678

  18. PSGL-1 Is an Immune Checkpoint Regulator that Promotes T Cell Exhaustion.

    PubMed

    Tinoco, Roberto; Carrette, Florent; Barraza, Monique L; Otero, Dennis C; Magaña, Jonathan; Bosenberg, Marcus W; Swain, Susan L; Bradley, Linda M

    2016-05-17

    Chronic viruses and cancers thwart immune responses in humans by inducing T cell dysfunction. Using a murine chronic virus that models human infections, we investigated the function of the adhesion molecule, P-selectin glycoprotein ligand-1 (PSGL-1), that is upregulated on responding T cells. PSGL-1-deficient mice cleared the virus due to increased intrinsic survival of multifunctional effector T cells that had downregulated PD-1 as well as other inhibitory receptors. Notably, this response resulted in CD4(+)-T-cell-dependent immunopathology. Mechanistically, PSGL-1 ligation on exhausted CD8(+) T cells inhibited T cell receptor (TCR) and interleukin-2 (IL-2) signaling and upregulated PD-1, leading to diminished survival with TCR stimulation. In models of melanoma cancer in which T cell dysfunction occurs, PSGL-1 deficiency led to PD-1 downregulation, improved T cell responses, and tumor control. Thus, PSGL-1 plays a fundamental role in balancing viral control and immunopathology and also functions to regulate T cell responses in the tumor microenvironment. PMID:27192578

  19. Interleukin-6 (IL-6) and IL-17 Synergistically Promote Viral Persistence by Inhibiting Cellular Apoptosis and Cytotoxic T Cell Function

    PubMed Central

    Hou, Wanqiu; Jin, Young-Hee; Kang, Hyun Seok

    2014-01-01

    ABSTRACT Interleukin-6 (IL-6) plays an important role in the development and progression of inflammatory responses, autoimmune diseases, and cancers. Many viral infections, including Theiler's murine encephalomyelitis virus (TMEV), result in the vigorous production of IL-6. However, the role of IL-6 in the development of virus-induced inflammatory responses is unclear. The infection of susceptible mice with TMEV induces the development of chronic demyelinating disease, which is considered a relevant infectious model for multiple sclerosis. In this study, we demonstrate that resistant C57BL/6 mice carrying an IL-6 transgene (IL-6 Tg) develop a TMEV-induced demyelinating disease accompanied by an increase in viral persistence and an elevated Th17 cell response in the central nervous system. Either IL-6 or IL-17 induced the expression of Bcl-2 and Bcl-xL at a high concentration. The upregulated expression of prosurvival molecules in turn inhibited target cell destruction by virus-specific CD8+ T cells. More interestingly, IL-6 and IL-17 synergistically promoted the expression of these prosurvival molecules, preventing cellular apoptosis at a much lower (<5-fold) concentration. The signals involved in the synergy appear to include the activation of both STAT3 and NF-κB via distinct cytokine-dependent pathways. Thus, the excessive IL-6 promotes the generation of Th17 cells, and the resulting IL-6 and IL-17 synergistically promote viral persistence by protecting virus-infected cells from apoptosis and CD8+ T cell-mediated target destruction. These results suggest that blocking both IL-6 and IL-17 functions are important considerations for therapies of chronic viral diseases, autoimmune diseases, and cancers. IMPORTANCE This study indicates that an excessive level of IL-6 cytokine produced following viral infection promotes the development of IL-17-producing pathogenic helper T cells. We demonstrate here for the first time that IL-6 together with IL-17 synergistically

  20. T-Cell Memory Responses Elicited by Yellow Fever Vaccine are Targeted to Overlapping Epitopes Containing Multiple HLA-I and -II Binding Motifs

    PubMed Central

    de Melo, Andréa Barbosa; Nascimento, Eduardo J. M.; Braga-Neto, Ulisses; Dhalia, Rafael; Silva, Ana Maria; Oelke, Mathias; Schneck, Jonathan P.; Sidney, John; Sette, Alessandro; Montenegro, Silvia M. L.; Marques, Ernesto T. A.

    2013-01-01

    The yellow fever vaccines (YF-17D-204 and 17DD) are considered to be among the safest vaccines and the presence of neutralizing antibodies is correlated with protection, although other immune effector mechanisms are known to be involved. T-cell responses are known to play an important role modulating antibody production and the killing of infected cells. However, little is known about the repertoire of T-cell responses elicited by the YF-17DD vaccine in humans. In this report, a library of 653 partially overlapping 15-mer peptides covering the envelope (Env) and nonstructural (NS) proteins 1 to 5 of the vaccine was utilized to perform a comprehensive analysis of the virus-specific CD4+ and CD8+ T-cell responses. The T-cell responses were screened ex-vivo by IFN-γ ELISPOT assays using blood samples from 220 YF-17DD vaccinees collected two months to four years after immunization. Each peptide was tested in 75 to 208 separate individuals of the cohort. The screening identified sixteen immunodominant antigens that elicited activation of circulating memory T-cells in 10% to 33% of the individuals. Biochemical in-vitro binding assays and immunogenetic and immunogenicity studies indicated that each of the sixteen immunogenic 15-mer peptides contained two or more partially overlapping epitopes that could bind with high affinity to molecules of different HLAs. The prevalence of the immunogenicity of a peptide in the cohort was correlated with the diversity of HLA-II alleles that they could bind. These findings suggest that overlapping of HLA binding motifs within a peptide enhances its T-cell immunogenicity and the prevalence of the response in the population. In summary, the results suggests that in addition to factors of the innate immunity, “promiscuous” T-cell antigens might contribute to the high efficacy of the yellow fever vaccines. PMID:23383350

  1. Tax abolishes histone H1 repression of p300 acetyltransferase activity at the human T-cell leukemia virus type 1 promoter.

    PubMed

    Konesky, Kasey L; Nyborg, Jennifer K; Laybourn, Paul J

    2006-11-01

    Upon infection of human T-cell leukemia virus type 1 (HTLV-1), the provirus is integrated into the host cell genome and subsequently packaged into chromatin that contains histone H1. Consequently, transcriptional activation of the virus requires overcoming the environment of chromatin and H1. To efficiently activate transcription, HTLV-1 requires the virally encoded protein Tax and cellular transcription factor CREB. Together Tax and CREB interact with three cis-acting promoter elements called viral cyclic-AMP response elements (vCREs). Binding of Tax and CREB to the vCREs promotes association of p300/CBP into the complex and leads to transcriptional activation. Therefore, to fully understand the mechanism of Tax transactivation, it is necessary to examine transcriptional activation from chromatin assembled with H1. Using a DNA template harboring the complete HTLV-1 promoter sequence and a highly defined recombinant assembly system, we demonstrate proper incorporation of histone H1 into chromatin. Addition of H1 to the chromatin template reduces HTLV-1 transcriptional activation through a novel mechanism. Specifically, H1 does not inhibit CREB or Tax binding to the vCREs or p300 recruitment to the promoter. Rather, H1 directly targets p300 acetyltransferase activity. Interestingly, in determining the mechanism of H1 repression, we have discovered a previously undefined function of Tax, overcoming the repressive effects of H1-chromatin. Tax specifically abrogates the H1 repression of p300 enzymatic activity in a manner independent of p300 recruitment and without displacement of H1 from the promoter. PMID:16943293

  2. MicroRNA223 promotes pathogenic T-cell development and autoimmune inflammation in central nervous system in mice.

    PubMed

    Satoorian, Tiffany; Li, Bo; Tang, Xiaolei; Xiao, Jidong; Xing, Weirong; Shi, Weixing; Lau, Kin-Hing William; Baylink, David J; Qin, Xuezhong

    2016-08-01

    Multiple sclerosis (MS) is an incurable central nervous system autoimmune disease. Understanding MS pathogenesis is essential for the development of new MS therapies. In the present study, we identified a novel microRNA (miR) that regulates experimental autoimmune encephalomyelitis (EAE), an animal model of MS. Expression of miR223 was up-regulated specifically in spinal cords and lymphoid organs but not in other examined tissues. A global miR223 knockout (miR223(-/-) ) in mice led to a significant delay in EAE onset, reduction in spinal cord lesion, and lessening of neurological symptoms. These protective effects could be reproduced in bone marrow chimeras reconstituted with miR223(-/-) haematopoietic stem cells. We also found that miR223 deficiency reduced T helper type 1 (Th1) and Th17 infiltration into spinal cords. To address underlying mechanisms, we investigated the role of miR223 in regulating the function, development and interaction of the major immune cells. Expression of the genes associated with dendritic cell (DC) activation (CD86 and MHC II) and Th1 and Th17 differentiation [interleukin-12 (IL-12) and IL-23, respectively] was significantly decreased in the spleens of miR223(-/-) mice bearing EAE. The miR223(-/-) DCs expressed significantly lower levels of basal and lipopolysaccharide-induced IL-12 and IL-23 compared with the wild-type DCs. These data are consistent with the observed lower efficiency of miR223(-/-) DCs to support Th1 and Th17 differentiation from naive T cells over-expressing an EAE antigen-specific T-cell receptor. Our data suggest that miR223 promotes EAE, probably through enhancing DC activation and subsequently the differentiation of naive T cells toward Th1 and Th17 effector cells. PMID:27083389

  3. Enhancing the Tumor Selectivity of a Picornavirus Virotherapy Promotes Tumor Regression and the Accumulation of Infiltrating CD8+ T Cells.

    PubMed

    Bell, Michael P; Pavelko, Kevin D

    2016-03-01

    Picornaviruses have emerged as promising cancer therapies due to their ability to drive cytotoxic cellular immune responses and for promoting oncolysis. These properties include preferential replication in tumor cells, the induction of strong innate and adaptive immune responses, and the ease with which their genomes can be manipulated. We have developed Theiler's murine encephalomyelitis virus (TMEV) as an immunotherapy vector that promotes strong adaptive immune responses to tumor antigens embedded within its genome. To further explore its usefulness as cancer therapy, we investigated whether direct intratumoral delivery of TMEV could promote tumor regression. We generated several picornavirus hybrids using substrains of TMEV that have unique immunopathologic characteristics, despite their extensive sequence homology. These hybrids exhibit a unique propensity to infect and replicate in melanoma. We have identified GD7-KS1, a virus that is particularly effective at replicating and infecting B16 melanoma in vitro and provides benefit as an oncolytic therapy in vivo after intratumoral injection. In addition, this virus promotes the mobilization and accumulation of CD8(+) T cells within treated tumors. Altogether, these findings demonstrate that picornavirus substrains can be used to rationally design virus hybrids that promote antitumor responses and add to the known strategies identified by us and others to further enhance the therapeutic potential of vectors used to treat cancer. PMID:26823492

  4. Chitin Recognition via Chitotriosidase Promotes Pathologic Type-2 Helper T Cell Responses to Cryptococcal Infection

    PubMed Central

    Wiesner, Darin L.; Specht, Charles A.; Lee, Chrono K.; Smith, Kyle D.; Mukaremera, Liliane; Lee, S. Thera; Lee, Chun G.; Elias, Jack A.; Nielsen, Judith N.; Boulware, David R.; Bohjanen, Paul R.; Jenkins, Marc K.; Levitz, Stuart M.; Nielsen, Kirsten

    2015-01-01

    Pulmonary mycoses are often associated with type-2 helper T (Th2) cell responses. However, mechanisms of Th2 cell accumulation are multifactorial and incompletely known. To investigate Th2 cell responses to pulmonary fungal infection, we developed a peptide-MHCII tetramer to track antigen-specific CD4+ T cells produced in response to infection with the fungal pathogen Cryptococcus neoformans. We noted massive accruement of pathologic cryptococcal antigen-specific Th2 cells in the lungs following infection that was coordinated by lung-resident CD11b+ IRF4-dependent conventional dendritic cells. Other researchers have demonstrated that this dendritic cell subset is also capable of priming protective Th17 cell responses to another pulmonary fungal infection, Aspergillus fumigatus. Thus, higher order detection of specific features of fungal infection by these dendritic cells must direct Th2 cell lineage commitment. Since chitin-containing parasites commonly elicit Th2 responses, we hypothesized that recognition of fungal chitin is an important determinant of Th2 cell-mediated mycosis. Using C. neoformans mutants or purified chitin, we found that chitin abundance impacted Th2 cell accumulation and disease. Importantly, we determined Th2 cell induction depended on cleavage of chitin via the mammalian chitinase, chitotriosidase, an enzyme that was also prevalent in humans experiencing overt cryptococcosis. The data presented herein offers a new perspective on fungal disease susceptibility, whereby chitin recognition via chitotriosidase leads to the initiation of harmful Th2 cell differentiation by CD11b+ conventional dendritic cells in response to pulmonary fungal infection. PMID:25764512

  5. Decidual vascular endothelial cells promote maternal-fetal immune tolerance by inducing regulatory T cells through canonical Notch1 signaling.

    PubMed

    Yao, Yanyi; Song, Jieping; Wang, Weipeng; Liu, Nian

    2016-05-01

    Adaptation of the maternal immune response to accommodate the semiallogeneic fetus is necessary for pregnancy success. However, the mechanisms by which the fetus avoids rejection despite expression of paternal alloantigens remain incompletely understood. Regulatory T cells (Treg cells) are pivotal for maintaining immune homeostasis, preventing autoimmune disease and fetus rejection. In this study, we found that maternal decidual vascular endothelial cells (DVECs) sustained Foxp3 expression in resting Treg cells in vitro. Moreover, under in vitro Treg cell induction condition with agonistic antibodies and transforming growth factor (TGF)-β, DVECs promoted Treg cell differentiation from non-Treg conventional T cells. Consistent with the promotion of Treg cell maintenance and differentiation, Treg cell-associated gene expression such as TGF-β, Epstein-Barr-induced gene-3, CD39 and glucocorticoid-induced tumor necrosis factor receptor was also increased in the presence of DVECs. Further study revealed that DVECs expressed Notch ligands such as Jagged-1, Delta-like protein 1 (DLL-1) and DLL-4, while Treg cells expressed Notch1 on their surface. The effects of DVECs on Treg cells was inhibited by siRNA-induced knockdown of expression of Jagged-1 and DLL-1 in DVECs. Downregulation of Notch1 in Treg cells using lentiviral shRNA transduction decreased Foxp3 expression in Treg cells. Adoptive transfer of Notch1-deficient Treg cells increased abortion rate in a murine semiallogeneic pregnancy model. Taken together, our study suggests that maternal DVECs are able to maintain decidual Treg cell identity and promote Treg cell differentiation through activation of Notch1 signal pathway in Treg cells and subsequently inhibit the immune response against semiallogeneic fetuses and preventing spontaneous abortion. PMID:26714886

  6. MAR binding protein SMAR1 favors IL-10 mediated regulatory T cell function in acute colitis.

    PubMed

    Mirlekar, Bhalchandra; Patil, Sachin; Bopanna, Ramanamurthy; Chattopadhyay, Samit

    2015-08-21

    Treg cells are not only crucial for controlling immune responses to autoantigens but also prevent those directed towards commensal pathogens. Control of effector immune responses by Treg cells depend on their capacity to accumulate at inflammatory site and accordingly accommodate to inflammatory environment. Till date, the factors associated with maintaining these aspects of Treg phenotype is not understood properly. Here we have shown that a known nuclear matrix binding protein SMAR1 is selectively expressed more in colonic Treg cells and is required for their ability to accumulate at inflammatory site and to sustain high levels of Foxp3 and IL-10 expression during acute colitis. Elimination of anti-inflammatory subsets revealed a protective role for IL-10 producing Treg cells in SMAR1(-/-) mice. Moreover, a combined action of Foxp3 and SMAR1 restricts effector cytokine production and enhance the production of IL-10 by colonic Treg cells that controls acute colitis. This data highlights a critical role of SMAR1 in maintaining Treg physiology during inflammatory disorders. PMID:26168735

  7. MAR binding protein SMAR1 favors IL-10 mediated regulatory T cell function in acute colitis

    SciTech Connect

    Mirlekar, Bhalchandra; Patil, Sachin; Bopanna, Ramanamurthy; Chattopadhyay, Samit

    2015-08-21

    T{sub reg} cells are not only crucial for controlling immune responses to autoantigens but also prevent those directed towards commensal pathogens. Control of effector immune responses by T{sub reg} cells depend on their capacity to accumulate at inflammatory site and accordingly accommodate to inflammatory environment. Till date, the factors associated with maintaining these aspects of T{sub reg} phenotype is not understood properly. Here we have shown that a known nuclear matrix binding protein SMAR1 is selectively expressed more in colonic T{sub reg} cells and is required for their ability to accumulate at inflammatory site and to sustain high levels of Foxp3 and IL-10 expression during acute colitis. Elimination of anti-inflammatory subsets revealed a protective role for IL-10 producing T{sub reg} cells in SMAR1{sup −/−} mice. Moreover, a combined action of Foxp3 and SMAR1 restricts effector cytokine production and enhance the production of IL-10 by colonic T{sub reg} cells that controls acute colitis. This data highlights a critical role of SMAR1 in maintaining T{sub reg} physiology during inflammatory disorders. - Highlights: • SMAR1 is essential to sustain high level of Foxp3 and IL-10 in T{sub reg} cells. • SMAR1{sup −/−} T{sub reg} cells produce pro-inflammatory cytokine IL-17 leads to inflammation. • IL-10 administration can control the inflammation in SMAR1{sup −/−} mice. • Both Foxp3 and SMAR1 maintain T{sub reg} phenotype that controls colitis.

  8. The intracellular B30.2 domain of Butyrophilin 3A1 binds phosphoantigens to mediate activation of human Vγ9Vδ2 T cells

    PubMed Central

    Sandstrom, Andrew; Peigné, Cassie-Marie; Léger, Alexandra; Crooks, James E.; Konczak, Fabienne; Gesnel, Marie-Claude; Breathnach, Richard; Bonneville, Marc; Scotet, Emmanuel; Adams, Erin J.

    2014-01-01

    Summary In humans, Vγ9Vδ2 T cells detect tumor cells and microbial infections including Mycobacterium tuberculosis through recognition of small pyrophosphate containing organic molecules known as phosphoantigens (pAgs). Key to pAg-mediated activation of Vγ9Vδ2 T cells is the butyrophilin 3A1 (BTN3A1) protein that contains an intracellular B30.2 domain critical to pAg reactivity. Here, we have demonstrated through structural, biophysical and functional approaches that the intracellular B30.2 domain of BTN3A1 directly binds pAg through a positively-charged surface pocket. Charge-reversal of pocket residues abrogates binding and Vγ9Vδ2 T cell activation. We have also identified a gain-of-function mutation within this pocket that when introduced into B30.2 domain of the non-stimulatory BTN3A3 isoform, transfers pAg binding ability and Vγ9Vδ2 T cell activation. These studies demonstrate that internal sensing of changes in pAg metabolite concentrations by BTN3A1 molecules is a critical step in Vγ9Vδ2 T cell detection of infection and tumorigenesis. PMID:24703779

  9. The intracellular B30.2 domain of butyrophilin 3A1 binds phosphoantigens to mediate activation of human Vγ9Vδ2 T cells.

    PubMed

    Sandstrom, Andrew; Peigné, Cassie-Marie; Léger, Alexandra; Crooks, James E; Konczak, Fabienne; Gesnel, Marie-Claude; Breathnach, Richard; Bonneville, Marc; Scotet, Emmanuel; Adams, Erin J

    2014-04-17

    In humans, Vγ9Vδ2 T cells detect tumor cells and microbial infections, including Mycobacterium tuberculosis, through recognition of small pyrophosphate containing organic molecules known as phosphoantigens (pAgs). Key to pAg-mediated activation of Vγ9Vδ2 T cells is the butyrophilin 3A1 (BTN3A1) protein that contains an intracellular B30.2 domain critical to pAg reactivity. Here, we have demonstrated through structural, biophysical, and functional approaches that the intracellular B30.2 domain of BTN3A1 directly binds pAg through a positively charged surface pocket. Charge reversal of pocket residues abrogates binding and Vγ9Vδ2 T cell activation. We have also identified a gain-of-function mutation within this pocket that, when introduced into the B30.2 domain of the nonstimulatory BTN3A3 isoform, transfers pAg binding ability and Vγ9Vδ2 T cell activation. These studies demonstrate that internal sensing of changes in pAg metabolite concentrations by BTN3A1 molecules is a critical step in Vγ9Vδ2 T cell detection of infection and tumorigenesis. PMID:24703779

  10. Snail and Slug Promote Epithelial-Mesenchymal Transition through β-Catenin–T-Cell Factor-4-dependent Expression of Transforming Growth Factor-β3

    PubMed Central

    Medici, Damian; Hay, Elizabeth D.

    2008-01-01

    Members of the Snail family of transcription factors have been shown to induce epithelial-mesenchymal transition (EMT), a fundamental mechanism of embryogenesis and progressive disease. Here, we show that Snail and Slug promote formation of β-catenin–T-cell factor (TCF)-4 transcription complexes that bind to the promoter of the TGF-β3 gene to increase its transcription. Subsequent transforming growth factor (TGF)-β3 signaling increases LEF-1 gene expression causing formation of β-catenin–lymphoid enhancer factor (LEF)-1 complexes that initiate EMT. TGF-β1 or TGF-β2 stimulates this signaling mechanism by up-regulating synthesis of Snail and Slug. TGF-β1- and TGF-β2-induced EMT were found to be TGF-β3 dependent, establishing essential roles for multiple TGF-β isoforms. Finally, we determined that β-catenin–LEF-1 complexes can promote EMT without upstream signaling pathways. These findings provide evidence for a unified signaling mechanism driven by convergence of multiple TGF-β and TCF signaling molecules that confers loss of cell–cell adhesion and acquisition of the mesenchymal phenotype. PMID:18799618

  11. Analyzing the effect of peptide-HLA-binding ability on the immunogenicity of potential CD8+ and CD4+ T cell epitopes in a large dataset.

    PubMed

    Wang, Shufeng; Li, Jintao; Chen, Xiaoling; Wang, Li; Liu, Wei; Wu, Yuzhang

    2016-08-01

    Immunogenicity is a key factor that influences whether a peptide presented by major histocompatibility complex (MHC) can be a T cell epitope. However, peptide immunization experiments have shown that approximately half of MHC class I-binding peptides cannot elicit a T cell response, indicating the importance of analyzing the variables affecting the immunogenicity of MHC-binding peptides. In this study, we hierarchically investigated the contribution of the binding stability and affinity of peptide-MHC complexes to immunogenicity based on the available quantitative data. We found that the immunogenicity of peptides presented by human leukocyte antigen (HLA) class I molecules was still predictable using the experimental binding affinity, although approximately one-third of the peptides with a binding affinity stronger than 500 nM were non-immunogenic, whereas the immunogenicity of HLA-II-presented peptides was predicted well using the experimental affinity and even the predicted affinity. The positive correlation between the binding affinity and stability was only observed in peptide-HLA-I complexes with a binding affinity stronger than 500 nM, which suggested that the stability alone could not be used for the prediction of immunogenicity. A characterization and comparison of the 'holes' in the CD8+ and CD4+ T cell repertoire provided an explanation for the observed differences between the immunogenicity of peptides presented by HLA class I and II molecules. We also provided the optimal affinity threshold for the potential CD4+ and CD8+ T cell epitopes. Our results provide important insights into the cellular immune response and the accurate prediction of T cell epitopes. PMID:27094547

  12. Lectin interactions with the Jurkat leukemic T-cell line: quantitative binding studies and interleukin-2 production

    SciTech Connect

    Dupuis, G.; Bastin, B.

    1988-03-01

    Phytohemagglutinin (PHA), concanavalin A (Con A), pea lectin, and wheat germ agglutinin (WGA) have been used to investigate their binding properties to Jurkat 77 6.8 leukemic human T cells and their ability to induce these cells to produce interleukin-2 (IL-2). Binding studies showed that the Jurkat cells fixed 0.82 +/- 0.11 microgram pea lectin, 2.02 +/- 0.17 micrograms Con A, 1.85 +/- 0.07 micrograms PHA and 8.88 +/- 0.61 micrograms WGA. Scatchard plots were linear, indicating that the binding process was homogeneous with respect to the binding constant. PHA and Con A bound with the highest affinity (Kass (apparent) approximately equal to 9 x 10(9) M-1), followed by pea lectin and WGA (Kass (apparent) approximately equal to 3 x 10(9) M-1). The number of lectin binding sites was in agreement with the results of saturation experiments. We also evaluated the effect of the presence of 12-O-tetradecanoylphorbol-13-acetate (TPA) on the binding process. Results show that there were no gross alterations in the value of (apparent) Kass in the case of PHA and WGA. In contrast, the presence of TPA decreased the affinity of Con A and modified the Scatchard profile for pea lectin, which was curvilinear with a concavity turned upward. In this case, data were (apparent) K1 = 17.7 x 10(9) M-1 (high-affinity sites) and (apparent) K2 = 2.6 x 10(9) M-1 (low-affinity sites). The four lectins shared the ability to stimulate Jurkat 77 6.8 cells to secrete IL-2. Optimal lectin concentrations were 20 micrograms/ml (PHA) and 50 micrograms/ml (WGA and Con A). Pea lectin failed to display a dose-response relationship, and IL-2 production increased proportionally with lectin concentration. Con A was the most efficient stimulator (250 U/ml), followed by WGA (160 U/ml) and PHA (108 U/ml).

  13. T cell intrinsic USP15 deficiency promotes excessive IFN-γ production and an immunosuppressive tumor microenvironment in MCA-induced fibrosarcoma

    PubMed Central

    Zou, Qiang; Jin, Jin; Xiao, Yichuan; Zhou, Xiaofei; Hu, Hongbo; Cheng, Xuhong; Kazimi, Nasser; Ullrich, Stephen E; Sun, Shao-Cong

    2015-01-01

    USP15 is a deubiquitinase that negatively regulates activation of naïve CD4+ T cells and generation of IFN-γ-producing T helper 1 (Th1) cells. USP15 deficiency in mice promotes antitumor T cell responses in a transplantable cancer model; however, it has remained unclear how deregulated T cell activation impacts primary tumor development during the prolonged interplay between tumors and the immune system. Here, we find that the USP15-deficient mice are hypersensitive to methylcholantrene (MCA)-induced fibrosarcomas. Excessive IFN-γ production in USP15-deficient mice promotes expression of the immunosuppressive molecule PD-L1 and the chemokine CXCL12, causing accumulation of T-bet+ regulatory T cells and CD11b+Gr-1+ myeloid-derived suppressor cells at tumor site. Mixed bone marrow adoptive transfer studies further reveals a T cell-intrinsic role for USP15 in regulating IFN-γ production and tumor development. These findings suggest that T cell intrinsic USP15 deficiency causes excessive production of IFN-γ, which promotes an immunosuppressive tumor microenvironment, during MCA-induced primary tumorigenesis. PMID:26686633

  14. Regulatory CD4+CD25+ T Cells Dampen Inflammatory Disease in Murine Mycoplasma Pneumonia and Promote IL-17 and IFN-γ Responses

    PubMed Central

    Odeh, Adam N.; Simecka, Jerry W.

    2016-01-01

    Mycoplasmas cause respiratory diseases characterized by persistent infection and chronic airway inflammation. Mycoplasma lung disease is immunopathologic, with CD4+ Th cells determining both disease severity and resistance to infection. Th2 cell responses promote immunopathology, while Th1 cells confer resistance to infection. However, regulatory CD4+ T cells may also have a role in the pathogenesis of mycoplasma respiratory diseases. We hypothesized Treg cells control the severity of the inflammatory lesions and may also promote persistence of infection. To examine this, BALB/c mice were depleted of CD25+ cells, and had increased disease severity due to Mycoplasma pulmonis infection. Increases in mycoplasma antibody responses and lymphocyte infiltration into lungs also occurred after CD25+ cell depletion. CD4+CD25+ regulatory T cells promoted IFN-γ and IL-17 mycoplasma-specific CD4+ T cell responses in vitro and in vivo, while dampening IL-13+ Th responses. Neither IL-10 nor TGF-ß expression was detected in CD4+CD25+ T cells from lymph nodes. Thus, a regulatory T cell population plays an important role in controlling damaging immune responses in mycoplasma respiratory disease but does not contribute to persistence of infection. It appears that a regulatory T cell population preferentially dampens Th2 cell-mediated inflammatory responses to mycoplasma through a mechanism independent of IL-10 or TGF-ß characteristic of “classic” Treg cells. PMID:27175511

  15. [miR-126 knockdown enhances the activity of murine CD4(+);T cells in vivo and promotes their differentiation into Th1 cells].

    PubMed

    Cui, Panpan; Hu, Yan; Tao, Yijing; Chen, Chao; Zhao, Juanjuan; Guo, Mengmeng; Zhou, Ya; Xu, Lin

    2016-03-01

    Objective To investigate the change of CD4(+);T cell activity in microRNA-126 (miR-126) knockdown (KD) mice and explore its significance. Methods The expression level of mature miR-126 in CD4(+);CD62L(+);T cells purified by magnetic-activated cell sorting (MACS) was analyzed by real-time PCR using specific probe. Furthermore, the expression levels of CD69, CD62L and CD44 molecules, as well as intracellular proliferating nuclear antigen Ki-67, in CD4(+);T cells in miR-126 KD mice were detected by fluorescence-activated cell sorting (FACS). Moreover, the apoptosis of CD4(+);T cells was analyzed by annexin V/PI staining assay combined with flow cytometry. Finally, the relative expressions of function-related cytokines including interleukine 4 (IL-4), IL-10, IL-12, transforming growth factor (TGF-β), interferon (IFN-γ) and tumor necrosis factor (TNF-α) in CD4(+);T cells were determined by real-time PCR. Results Compared with wild-type (WT) mice, the expression level of mature miR-126 in CD4(+);T cells in miR-126 KD mice was dramatically reduced. Furthermore, the proportion of CD62L(+); in CD4(+);T cells also decreased significantly, while the proportions of CD69(+);, CD44(+); and Ki-67(+); cells were remarkably elevated. Meanwhile, the apoptosis proportion of CD4(+);T cells in vivo dropped dramatically in miR-126 KD mice. Finally, the mRNA expressions of IL-4 and IL-10 in CD4(+);T cells were significantly downregulated, but IL-12, TGF-β, TNF-α and IFN-γ mRNAs were obviously up-regulated. Conclusion miR-126 knockdown could significantly enhance the functional activity of CD4(+); T cells in vivo and promote cell differentiation into Th1 cells. PMID:26927555

  16. Polyelectrolyte Multilayers Assembled Entirely from Immune Signals on Gold Nanoparticle Templates Promote Antigen-Specific T Cell Response.

    PubMed

    Zhang, Peipei; Chiu, Yu-Chieh; Tostanoski, Lisa H; Jewell, Christopher M

    2015-06-23

    Materials that allow modular, defined assembly of immune signals could support a new generation of rationally designed vaccines that promote tunable immune responses. Toward this goal, we have developed the first polyelectrolyte multilayer (PEM) coatings built entirely from immune signals. These immune-PEMs (iPEMs) are self-assembled on gold nanoparticle templates through stepwise electrostatic interactions between peptide antigen and polyanionic toll-like receptor (TLR) agonists that serve as molecular adjuvants. iPEMs do not require solvents or mixing, offer direct control over the composition and loading of vaccine components, and can be coated on substrates at any scale. These films also do not require other structural components, eliminating the potentially confounding effects caused by the inherent immune-stimulatory characteristics of many synthetic polymers. iPEM loading on gold nanoparticle substrates is tunable, and cryoTEM reveals iPEM shells coated on gold cores. These nanoparticles are efficiently internalized by primary dendritic cells (DCs), resulting in activation, selective triggering of TLR signaling, and presentation of the antigens used to assemble iPEMs. In coculture, iPEMs drive antigen-specific T cell proliferation and effector cytokines but not cytokines associated with more generalized inflammation. Compared to mice treated with soluble antigen and adjuvant, iPEM immunization promotes high levels of antigen-specific CD8(+) T cells in peripheral blood after 1 week. These enhancements result from increased DC activation and antigen presentation in draining lymph nodes. iPEM-immunized mice also exhibit a potent recall response after boosting, supporting the potential of iPEMs for designing well-defined vaccine coatings that provide high cargo density and eliminate synthetic film components. PMID:26035231

  17. Transactivation of human osteopontin promoter by human T-cell leukemia virus type 1-encoded Tax protein.

    PubMed

    Zhang, Jing; Yamada, Osamu; Matsushita, Yoshihisa; Chagan-Yasutan, Haorile; Hattori, Toshio

    2010-06-01

    Osteopontin (OPN) is a cytokine that contributes substantially to the growth and metastasis in a wide spectrum of malignancies. We report here that OPN gene is transactivated by Tax protein of human T-cell leukemia virus type 1 (HTLV-1). Northern blot showed enhanced OPN gene expression in cells stably expressing Tax. Co-expression of Tax increased the reporter gene expression directed by OPN promoter. Tax-induced OPN activation was abrogated by treatment with LY294002 (PI3K inhibitor) or co-transfection with AKT siRNA, suggesting PI3K/AKT pathway is involved in Tax-mediated transactivation. Reporter assay with deletion mutants showed that the 5'-partial sequence between -765 and -660 of the OPN promoter is the region responsive to Tax, and further, disrupting the AP-1 site within this region abolished the OPN induction by Tax, indicating that Tax activation of OPN promoter is likely mediated by AP-1 site. This study suggests that OPN is one of the downstream mediators of aberrantly activated PI3K/AKT signaling by Tax, which may partially contribute to HTLV-1-associated leukemogenesis. PMID:19767100

  18. The Length Distribution of Class I-Restricted T Cell Epitopes Is Determined by Both Peptide Supply and MHC Allele-Specific Binding Preference.

    PubMed

    Trolle, Thomas; McMurtrey, Curtis P; Sidney, John; Bardet, Wilfried; Osborn, Sean C; Kaever, Thomas; Sette, Alessandro; Hildebrand, William H; Nielsen, Morten; Peters, Bjoern

    2016-02-15

    HLA class I-binding predictions are widely used to identify candidate peptide targets of human CD8(+) T cell responses. Many such approaches focus exclusively on a limited range of peptide lengths, typically 9 aa and sometimes 9-10 aa, despite multiple examples of dominant epitopes of other lengths. In this study, we examined whether epitope predictions can be improved by incorporating the natural length distribution of HLA class I ligands. We found that, although different HLA alleles have diverse length-binding preferences, the length profiles of ligands that are naturally presented by these alleles are much more homogeneous. We hypothesized that this is due to a defined length profile of peptides available for HLA binding in the endoplasmic reticulum. Based on this, we created a model of HLA allele-specific ligand length profiles and demonstrate how this model, in combination with HLA-binding predictions, greatly improves comprehensive identification of CD8(+) T cell epitopes. PMID:26783342

  19. A Cleavable N-Terminal Signal Peptide Promotes Widespread Olfactory Receptor Surface Expression in HEK293T Cells

    PubMed Central

    Shepard, Blythe D.; Natarajan, Niranjana; Protzko, Ryan J.; Acres, Omar W.; Pluznick, Jennifer L.

    2013-01-01

    Olfactory receptors (ORs) are G protein-coupled receptors that detect odorants in the olfactory epithelium, and comprise the largest gene family in the genome. Identification of OR ligands typically requires OR surface expression in heterologous cells; however, ORs rarely traffic to the cell surface when exogenously expressed. Therefore, most ORs are orphan receptors with no known ligands. To date, studies have utilized non-cleavable rhodopsin (Rho) tags and/or chaperones (i.e. Receptor Transporting Protein, RTP1S, Ric8b and Gαolf) to improve surface expression. However, even with these tools, many ORs still fail to reach the cell surface. We used a test set of fifteen ORs to examine the effect of a cleavable leucine-rich signal peptide sequence (Lucy tag) on OR surface expression in HEK293T cells. We report here that the addition of the Lucy tag to the N-terminus increases the number of ORs reaching the cell surface to 7 of the 15 ORs (as compared to 3/15 without Rho or Lucy tags). Moreover, when ORs tagged with both Lucy and Rho were co-expressed with previously reported chaperones (RTP1S, Ric8b and Gαolf), we observed surface expression for all 15 receptors examined. In fact, two-thirds of Lucy-tagged ORs are able to reach the cell surface synergistically with chaperones even when the Rho tag is removed (10/15 ORs), allowing for the potential assessment of OR function with only an 8-amino acid Flag tag on the mature protein. As expected for a signal peptide, the Lucy tag was cleaved from the mature protein and did not alter OR-ligand binding and signaling. Our studies demonstrate that widespread surface expression of ORs can be achieved in HEK293T cells, providing promise for future large-scale deorphanization studies. PMID:23840901

  20. Thymic B cells promote thymus-derived regulatory T cell development and proliferation.

    PubMed

    Lu, Fang-Ting; Yang, Wei; Wang, Yin-Hu; Ma, Hong-Di; Tang, Wei; Yang, Jing-Bo; Li, Liang; Ansari, Aftab A; Lian, Zhe-Xiong

    2015-07-01

    Thymic CD4(+) FoxP3(+) regulatory T (Treg) cells are critical for the development of immunological tolerance and immune homeostasis and requires contributions of both thymic dendritic and epithelial cells. Although B cells have been reported to be present within the thymus, there has not hitherto been a definition of their role in immune cell development and, in particular, whether or how they contribute to the Treg cellular thymic compartment. Herein, using both phenotypic and functional approaches, we demonstrate that thymic B cells contribute to the maintenance of thymic Treg cells and, using an in vitro culture system, demonstrate that thymic B cells contribute to the size of the thymic Treg compartment via cell-cell MHC II contact and the involvement of two independent co-stimulatory pathways that include interactions between the CD40/CD80/CD86 co-stimulatory molecules. Our data also suggest that thymic B cells promote the generation of thymic Treg cell precursors (pre-Treg cells), but not the conversion of FoxP3(+) Treg cells from pre-Treg cells. In addition, thymic B cells directly promote the proliferation of thymic Treg cells that is MHC II contact dependent with a minimal if any role for co-stimulatory molecules including CD40/CD80/CD86. Both pathways are independent of TGFβ. In conclusion, we rigorously define the critical role of thymic B cells in the development of thymic Treg cells from non-Treg to precursor stage and in the proliferation of mature thymic Treg cells. PMID:26071985

  1. HTLV-1 bZIP factor protein targets the Rb/E2F-1 pathway to promote proliferation and apoptosis of primary CD4(+) T cells.

    PubMed

    Kawatsuki, A; Yasunaga, J-I; Mitobe, Y; Green, P L; Matsuoka, M

    2016-08-25

    Human T-cell leukemia virus type 1 (HTLV-1) is an oncogenic retrovirus that induces a fatal T-cell malignancy, adult T-cell leukemia (ATL). Among several regulatory/accessory genes in HTLV-1, HTLV-1 bZIP factor (HBZ) is the only viral gene constitutively expressed in infected cells. Our previous study showed that HBZ functions in two different molecular forms, HBZ protein and HBZ RNA. In this study, we show that HBZ protein targets retinoblastoma protein (Rb), which is a critical tumor suppressor in many types of cancers. HBZ protein interacts with the Rb/E2F-1 complex and activates the transcription of E2F-target genes associated with cell cycle progression and apoptosis. Mouse primary CD4(+) T cells transduced with HBZ show accelerated G1/S transition and apoptosis, and importantly, T cells from HBZ transgenic (HBZ-Tg) mice also demonstrate enhanced cell proliferation and apoptosis. To evaluate the functions of HBZ protein alone in vivo, we generated a new transgenic mouse strain that expresses HBZ mRNA altered by silent mutations but encoding intact protein. In these mice, the numbers of effector/memory and Foxp3(+) T cells were increased, and genes associated with proliferation and apoptosis were upregulated. This study shows that HBZ protein promotes cell proliferation and apoptosis in primary CD4(+) T cells through activation of the Rb/E2F pathway, and that HBZ protein also confers onto CD4(+) T-cell immunophenotype similar to those of ATL cells, suggesting that HBZ protein has important roles in dysregulation of CD4(+) T cells infected with HTLV-1. PMID:26804169

  2. IFNα/βR Signaling Promotes Regulatory T Cell Development and Function Under Stress Conditions

    PubMed Central

    Metidji, Amina; Rieder, Sadiye Amcaoglu; Glass, Deborah Dacek; Cremer, Isabelle; Punkosdy, George A.; Shevach, Ethan M.

    2015-01-01

    Type I IFNs are a family of cytokines with antiviral and immunomodulatory properties. While the antiviral effects of IFNs are well characterized, their immunomodulatory properties are less clear. To specifically address the effects of type I IFNs on Treg, we studied mixed bone morrow (BM) chimeras between wild-type (WT) and IFNα/βR (IFNAR) knockout (KO) mice, and heterozygous female mice expressing a Treg-specific deletion of the IFNAR. In these two models, IFNAR signaling promotes the development of the Treg lineage in the thymus and their survival in the periphery. IFNAR KO Treg had a higher expression of the pro-apoptotic gene Bim and higher frequency of active caspase positive cells. IFNAR KO Treg from chimeric mice displayed a more naïve phenotype, accompanied by lower levels of CD25 and phosphorylated STAT5. Therefore, in Treg IFNAR signaling may directly or indirectly affect phosphorylation of STAT5. In mixed chimeras with Scurfy fetal liver, Treg derived from IFNAR KO BM were unable to control T effector cell activation and tissue inflammation. Under stress conditions or in a competitive environment, IFNAR signaling may be required to maintain Treg homeostasis and function. PMID:25795758

  3. Two Different Regulatory T Cell Populations That Promote Corneal Allograft Survival

    PubMed Central

    Cunnusamy, Khrishen; Paunicka, Kathryn; Reyes, Nancy; Yang, Wanhua; Chen, Peter W.

    2010-01-01

    Purpose. To compare and contrast the T regulatory cells (Tregs) induced by anterior chamber (AC) injection of antigen with those induced by orthotopic corneal allografts. Methods. Anterior chamber–associated immune deviation (ACAID) Tregs were induced by injecting C57BL/6 spleen cells into the AC of BALB/c mice. Delayed-type hypersensitivity responses to C57BL/6 alloantigens were evaluated by a conventional ear swelling assay. Corneal allograft Tregs were induced by applying orthotopic C57BL/6 corneal allografts onto BALB/c hosts. The effects of anti-CD25, anti-CD8, anti-interferon-γ (IFN-γ), anti-IL-17A, or cyclophosphamide treatments on corneal allograft survival and ACAID were evaluated. Results. Administration of either anti-CD25 or anti-IFN-γ antibodies prevented the expression of ACAID and abolished the immune privilege of corneal allografts. By contrast, in vivo treatment with anti-CD8 antibody abrogated ACAID but had no effect on corneal allograft survival. Further discordance between ACAID and corneal allograft survival emerged in experiments in which the induction of allergic conjunctivitis or the administration of anti-IL-17A abolished the immune privilege of corneal allografts but had no effect on the induction or expression of ACAID. Conclusions. Although orthotopic corneal allografts are strategically located for the induction of ACAID by the sloughing of corneal cells into the AC, the results reported here indicate that the Tregs induced by orthotopic corneal allografts are remarkably different from the Tregs that are induced by AC injection of alloantigen. Although both of these Treg populations promote corneal allograft survival, they display distinctly different phenotypes. PMID:20702818

  4. XCR1+ dendritic cells promote memory CD8+ T cell recall upon secondary infections with Listeria monocytogenes or certain viruses.

    PubMed

    Alexandre, Yannick O; Ghilas, Sonia; Sanchez, Cindy; Le Bon, Agnès; Crozat, Karine; Dalod, Marc

    2016-01-11

    Naive CD8(+) T cell priming during tumor development or many primary infections requires cross-presentation by XCR1(+) dendritic cells (DCs). Memory CD8(+) T lymphocytes (mCTLs) harbor a lower activation threshold as compared with naive cells. However, whether their recall responses depend on XCR1(+) DCs is unknown. By using a new mouse model allowing fluorescent tracking and conditional depletion of XCR1(+) DCs, we demonstrate a differential requirement of these cells for mCTL recall during secondary infections by different pathogens. XCR1(+) DCs were instrumental to promote this function upon secondary challenges with Listeria monocytogenes, vesicular stomatitis virus, or Vaccinia virus, but dispensable in the case of mouse cytomegalovirus. We deciphered how XCR1(+) DCs promote mCTL recall upon secondary infections with Listeria. By visualizing for the first time the in vivo choreography of XCR1(+) DCs, NK cells and mCTLs during secondary immune responses, and by neutralizing in vivo candidate molecules, we demonstrate that, very early after infection, mCTLs are activated, and attracted in a CXCR3-dependent manner, by NK cell-boosted, IL-12-, and CXCL9-producing XCR1(+) DCs. Hence, depending on the infectious agent, strong recall of mCTLs during secondary challenges can require cytokine- and chemokine-dependent cross-talk with XCR1(+) DCs and NK cells. PMID:26694969

  5. Eosinophilia of dystrophin-deficient muscle is promoted by perforin-mediated cytotoxicity by T cell effectors

    NASA Technical Reports Server (NTRS)

    Cai, B.; Spencer, M. J.; Nakamura, G.; Tseng-Ong, L.; Tidball, J. G.

    2000-01-01

    Previous investigations have shown that cytotoxic T lymphocytes (CTLs) contribute to muscle pathology in the dystrophin-null mutant mouse (mdx) model of Duchenne muscular dystrophy through perforin-dependent and perforin-independent mechanisms. We have assessed whether the CTL-mediated pathology includes the promotion of eosinophilia in dystrophic muscle, and thereby provides a secondary mechanism through which CTLs contribute to muscular dystrophy. Quantitative immunohistochemistry confirmed that eosinophilia is a component of the mdx dystrophy. In addition, electron microscopic observations show that eosinophils traverse the basement membrane of mdx muscle fibers and display sites of close apposition of eosinophil and muscle membranes. The close membrane apposition is characterized by impingement of eosinophilic rods of major basic protein into the muscle cell membrane. Transfer of mdx splenocytes and mdx muscle extracts to irradiated C57 mice by intraperitoneal injection resulted in muscle eosinophilia in the recipient mice. Double-mutant mice lacking dystrophin and perforin showed less eosinophilia than was displayed by mdx mice that expressed perforin. Finally, administration of prednisolone, which has been shown previously to reduce the concentration of CTLs in dystrophic muscle, produced a significant reduction in eosinophilia. These findings indicate that eosinophilia is a component of the mdx pathology that is promoted by perforin-dependent cytotoxicity of effector T cells. However, some eosinophilia of mdx muscle is independent of perforin-mediated processes.

  6. XCR1+ dendritic cells promote memory CD8+ T cell recall upon secondary infections with Listeria monocytogenes or certain viruses

    PubMed Central

    Alexandre, Yannick O.; Ghilas, Sonia; Sanchez, Cindy; Le Bon, Agnès

    2016-01-01

    Naive CD8+ T cell priming during tumor development or many primary infections requires cross-presentation by XCR1+ dendritic cells (DCs). Memory CD8+ T lymphocytes (mCTLs) harbor a lower activation threshold as compared with naive cells. However, whether their recall responses depend on XCR1+ DCs is unknown. By using a new mouse model allowing fluorescent tracking and conditional depletion of XCR1+ DCs, we demonstrate a differential requirement of these cells for mCTL recall during secondary infections by different pathogens. XCR1+ DCs were instrumental to promote this function upon secondary challenges with Listeria monocytogenes, vesicular stomatitis virus, or Vaccinia virus, but dispensable in the case of mouse cytomegalovirus. We deciphered how XCR1+ DCs promote mCTL recall upon secondary infections with Listeria. By visualizing for the first time the in vivo choreography of XCR1+ DCs, NK cells and mCTLs during secondary immune responses, and by neutralizing in vivo candidate molecules, we demonstrate that, very early after infection, mCTLs are activated, and attracted in a CXCR3-dependent manner, by NK cell–boosted, IL-12–, and CXCL9-producing XCR1+ DCs. Hence, depending on the infectious agent, strong recall of mCTLs during secondary challenges can require cytokine- and chemokine-dependent cross-talk with XCR1+ DCs and NK cells. PMID:26694969

  7. Identification of a complex that binds to the CD154 3' untranslated region: implications for a role in message stability during T cell activation.

    PubMed

    Barnhart, B; Kosinski, P A; Wang, Z; Ford, G S; Kiledjian, M; Covey, L R

    2000-10-15

    CD154 expression is regulated throughout a time course of CD3-dependent T cell activation by differential mRNA decay. To understand the molecular basis of the "stability" phase of this pathway, experiments were conducted to identify sequences and specific complexes important in this regulation. Gel retardation assays using extracts from both Jurkat T cells and CD3-activated CD4(+) T cells revealed a major complex (complex I) that bound a 65-bp highly CU-rich region of the CD154 3' untranslated region. The specificity of the CU-rich element for complex-I formation was confirmed by disruption of this complex by oligo(dCT) competition. Formation of complex I strongly correlated with CD154 mRNA stability across a time course of T cell activation. UV cross-linking identified a major oligo(dCT)-sensitive species at approximately 90 kDa that showed induced and increased expression in extracts from 24- and 48-hr anti-CD3-activated T cells, respectively. This protein was absent in equivalent extracts from resting or 2-h-activated T cells. Using an in vitro decay assay, we found that a CD154-specific transcript was more rapidly degraded in 2-h-activated extract and stabilized in the 24- and 48-h extracts compared to extracts from resting T cells. Disruption of complex I resulted in the rapid decay of a CD154-specific transcript demonstrating a functional role for complex I in mRNA stabilization in vitro. These studies support a model of posttranscriptional regulation of CD154 expression being controlled in part by the interaction of a poly(CU)-binding complex with a specific sequence in the 3' untranslated region. PMID:11035087

  8. Synergy between IL-15 and Id2 promotes the expansion of human NK progenitor cells, which can be counteracted by the E protein HEB required to drive T cell development.

    PubMed

    Schotte, Remko; Dontje, Wendy; Nagasawa, Maho; Yasuda, Yuko; Bakker, Arjen Q; Spits, Hergen; Blom, Bianca

    2010-06-15

    The cytokine IL-15 and the inhibitor of DNA binding (Id)2, which negatively regulates the activity of basic helix-loop-helix transcription factors, have been shown to play key roles in NK cell development. Consistent with this, exogenous IL-15 added to human thymic progenitor cells stimulated their development into NK cells at the expense of T cells both in fetal thymic organ culture and in coculture with stromal cells expressing the Notch ligand Delta-like 1. Overexpression of Id2 in thymic progenitor cells stimulated NK cell development and blocked T cell development. This, in part, is attributed to inhibition of the transcriptional activity of the E protein HEB, which we show in this study is the only E protein that enhanced T cell development. Notably, Id2 increased a pool of lineage CD1a-CD5+ progenitor cells that in synergy with IL-15 furthered expansion and differentiation into NK cells. Taken together, our findings point to a dualistic function of Id2 in controlling T/NK cell lineage decisions; T cell development is impaired by Id2, most likely by sequestering HEB, whereas NK cell development is promoted by increasing a pool of CD1a-CD5+ NK cell progenitors, which together with IL-15 differentiate into mature NK cells. PMID:20483740

  9. Regulatory T Cells Promote β-Catenin–Mediated Epithelium-to-Mesenchyme Transition During Radiation-Induced Pulmonary Fibrosis

    SciTech Connect

    Xiong, Shanshan; Pan, Xiujie; Xu, Long; Yang, Zhihua; Guo, Renfeng; Gu, Yongqing; Li, Ruoxi; Wang, Qianjun; Xiao, Fengjun; Du, Li; Zhou, Pingkun; Zhu, Maoxiang

    2015-10-01

    Purpose: Radiation-induced pulmonary fibrosis results from thoracic radiation therapy and severely limits radiation therapy approaches. CD4{sup +}CD25{sup +}FoxP3{sup +} regulatory T cells (Tregs) as well as epithelium-to-mesenchyme transition (EMT) cells are involved in pulmonary fibrosis induced by multiple factors. However, the mechanisms of Tregs and EMT cells in irradiation-induced pulmonary fibrosis remain unclear. In the present study, we investigated the influence of Tregs on EMT in radiation-induced pulmonary fibrosis. Methods and Materials: Mice thoraxes were irradiated (20 Gy), and Tregs were depleted by intraperitoneal injection of a monoclonal anti-CD25 antibody 2 hours after irradiation and every 7 days thereafter. Mice were treated on days 3, 7, and 14 and 1, 3, and 6 months post irradiation. The effectiveness of Treg depletion was assayed via flow cytometry. EMT and β-catenin in lung tissues were detected by immunohistochemistry. Tregs isolated from murine spleens were cultured with mouse lung epithelial (MLE) 12 cells, and short interfering RNA (siRNA) knockdown of β-catenin in MLE 12 cells was used to explore the effects of Tregs on EMT and β-catenin via flow cytometry and Western blotting. Results: Anti-CD25 antibody treatment depleted Tregs efficiently, attenuated the process of radiation-induced pulmonary fibrosis, hindered EMT, and reduced β-catenin accumulation in lung epithelial cells in vivo. The coculture of Tregs with irradiated MLE 12 cells showed that Tregs could promote EMT in MLE 12 cells and that the effect of Tregs on EMT was partially abrogated by β-catenin knockdown in vitro. Conclusions: Tregs can promote EMT in accelerating radiation-induced pulmonary fibrosis. This process is partially mediated through β-catenin. Our study suggests a new mechanism for EMT, promoted by Tregs, that accelerates radiation-induced pulmonary fibrosis.

  10. B cells promote inflammation in obesity and type 2 diabetes through regulation of T-cell function and an inflammatory cytokine profile.

    PubMed

    DeFuria, Jason; Belkina, Anna C; Jagannathan-Bogdan, Madhumita; Snyder-Cappione, Jennifer; Carr, Jordan David; Nersesova, Yanina R; Markham, Douglas; Strissel, Katherine J; Watkins, Amanda A; Zhu, Min; Allen, Jessica; Bouchard, Jacqueline; Toraldo, Gianluca; Jasuja, Ravi; Obin, Martin S; McDonnell, Marie E; Apovian, Caroline; Denis, Gerald V; Nikolajczyk, Barbara S

    2013-03-26

    Patients with type 2 diabetes (T2D) have disease-associated changes in B-cell function, but the role these changes play in disease pathogenesis is not well established. Data herein show B cells from obese mice produce a proinflammatory cytokine profile compared with B cells from lean mice. Complementary in vivo studies show that obese B cell-null mice have decreased systemic inflammation, inflammatory B- and T-cell cytokines, adipose tissue inflammation, and insulin resistance (IR) compared with obese WT mice. Reduced inflammation in obese/insulin resistant B cell-null mice associates with an increased percentage of anti-inflammatory regulatory T cells (Tregs). This increase contrasts with the sharply decreased percentage of Tregs in obese compared with lean WT mice and suggests that B cells may be critical regulators of T-cell functions previously shown to play important roles in IR. We demonstrate that B cells from T2D (but not non-T2D) subjects support proinflammatory T-cell function in obesity/T2D through contact-dependent mechanisms. In contrast, human monocytes increase proinflammatory T-cell cytokines in both T2D and non-T2D analyses. These data support the conclusion that B cells are critical regulators of inflammation in T2D due to their direct ability to promote proinflammatory T-cell function and secrete a proinflammatory cytokine profile. Thus, B cells are potential therapeutic targets for T2D. PMID:23479618

  11. Donor treatment with pegylated G-CSF augments the generation of IL-10-producing regulatory T cells and promotes transplantation tolerance.

    PubMed

    Morris, Edward S; MacDonald, Kelli P A; Rowe, Vanessa; Johnson, Diana H; Banovic, Tatjana; Clouston, Andrew D; Hill, Geoffrey R

    2004-05-01

    We investigated whether the protection from graft-versus-host disease (GVHD) afforded by donor treatment with granulocyte colony-stimulating factor (G-CSF) could be enhanced by dose escalation. Donor treatment with human G-CSF prevented GVHD in the B6 --> B6D2F1 murine model in a dose-dependent fashion, and murine G-CSF provided equivalent protection from GVHD at 10-fold lower doses. Donor pretreatment with a single dose of pegylated G-CSF (peg-G-CSF) prevented GVHD to a significantly greater extent than standard G-CSF (survival, 75% versus 11%, P <.001). Donor T cells from peg-G-CSF-treated donors failed to proliferate to alloantigen and inhibited the responses of control T cells in an interleukin 10 (IL-10)-dependent fashion in vitro. T cells from peg-G-CSF-treated IL-10(-/-) donors induced lethal GVHD; T cells from peg-G-CSF-treated wild-type (wt) donors promoted long-term survival. Whereas T cells from peg-G-CSF wt donors were able to regulate GVHD induced by T cells from control-treated donors, T cells from G-CSF-treated wt donors and peg-G-CSF-treated IL-10(-/-) donors did not prevent mortality. Thus, peg-G-CSF is markedly superior to standard G-CSF for the prevention of GVHD following allogeneic stem cell transplantation (SCT), due to the generation of IL-10-producing regulatory T cells. These data support prospective clinical trials of peg-G-CSF-mobilized allogeneic blood SCT. PMID:14726406

  12. Infiltrating T Cells Promote Bladder Cancer Progression via Increasing IL1→Androgen Receptor→HIF1α→VEGFa Signals.

    PubMed

    Tao, Le; Qiu, Jianxin; Jiang, Ming; Song, Wenbin; Yeh, Shuyuan; Yu, Hong; Zang, Lijuan; Xia, Shujie; Chang, Chawnshang

    2016-08-01

    The tumor microenvironment impacts tumor progression and individual cells, including CD4(+) T cells, which have been detected in bladder cancer tissues. The detailed mechanism of how these T cells were recruited to the bladder cancer tumor and their impact on bladder cancer progression, however, remains unclear. Using a human clinical bladder cancer sample survey and in vitro coculture system, we found that bladder cancer has a greater capacity to recruit T cells than surrounding normal bladder tissues. The consequences of higher levels of recruited T cells in bladder cancer included increased bladder cancer metastasis. Mechanism dissection revealed that infiltrating T cells might function through secreting the cytokine IL1, which increases the recruitment of T cells to bladder cancer and enhances the bladder cancer androgen receptor (AR) signaling that results in increased bladder cancer cell invasion via upregulation of hypoxia-inducible factor-1α (HIF1α)/VEGFa expression. Interruption of the IL1→AR→HIF1α→VEGFa signals with inhibitors of HIF1α or VEGFa partially reversed the enhanced bladder cancer cell invasion. Finally, in vivo mouse models of xenografted bladder cancer T24 cells with CD4(+) T cells confirmed in vitro coculture studies and concluded that infiltrating CD4(+) T cells can promote bladder cancer metastasis via modulation of the IL1→AR→HIF1α→VEGFa signaling. Future clinical trials using small molecules to target this newly identified signaling pathway may facilitate the development of new therapeutic approaches to better suppress bladder cancer metastasis. Mol Cancer Ther; 15(8); 1943-51. ©2016 AACR. PMID:27196763

  13. Combination OX40 agonism/CTLA-4 blockade with HER2 vaccination reverses T-cell anergy and promotes survival in tumor-bearing mice

    PubMed Central

    Linch, Stefanie N.; Kasiewicz, Melissa J.; McNamara, Michael J.; Hilgart-Martiszus, Ian F.; Farhad, Mohammad

    2016-01-01

    Immunotherapy is gathering momentum as a primary therapy for cancer patients. However, monotherapies have limited efficacy in improving outcomes and benefit only a subset of patients. Combination therapies targeting multiple pathways can augment an immune response to improve survival further. Here, we demonstrate that dual aOX40 (anti-CD134)/aCTLA-4 (anti–cytotoxic T-lymphocyte–associated protein 4) immunotherapy generated a potent antigen-specific CD8 T-cell response, enhancing expansion, effector function, and memory T-cell persistence. Importantly, OX40 and CTLA-4 expression on CD8 T cells was critical for promoting their maximal expansion following combination therapy. Animals treated with combination therapy and vaccination using anti–DEC-205 (dendritic and epithelial cells, 205 kDa)–HER2 (human epidermal growth factor receptor 2) had significantly improved survival in a mammary carcinoma model. Vaccination with combination therapy uniquely restricted Th2-cytokine production by CD4 cells, relative to combination therapy alone, and enhanced IFNγ production by CD8 and CD4 cells. We observed an increase in MIP-1α (macrophage inflammatory protein-1α)/CCL3 [chemokine (C-C motif) ligand 3], MIP-1β/CCL4, RANTES (regulated on activation, normal T-cell expressed and excreted)/CCL5, and GM-CSF production by CD8 and CD4 T cells following treatment. Furthermore, this therapy was associated with extensive tumor destruction and T-cell infiltration into the tumor. Notably, in a spontaneous model of prostate adenocarcinoma, vaccination with combination therapy reversed anergy and enhanced the expansion and function of CD8 T cells recognizing a tumor-associated antigen. Collectively, these data demonstrate that the addition of a vaccine with combined aOX40/aCTLA-4 immunotherapy augmented antitumor CD8 T-cell function while limiting Th2 polarization in CD4 cells and improved overall survival. PMID:26729864

  14. Human T-cell leukemia virus type 1 oncoprotein tax represses ZNF268 expression through the cAMP-responsive element-binding protein/activating transcription factor pathway.

    PubMed

    Wang, Di; Guo, Ming-Xiong; Hu, Hai-Ming; Zhao, Zhou-Zhou; Qiu, Hong-Ling; Shao, Huan-Jie; Zhu, Chen-Gang; Xue, Lu; Shi, Yun-Bo; Li, Wen-Xin

    2008-06-13

    Expression of the human T-cell leukemia virus type 1 (HTLV-1) oncoprotein Tax is correlated with cellular transformation, contributing to the development of adult T-cell leukemia. In this study, we investigated the role of Tax in the regulation of the ZNF268 gene, which plays a role in the differentiation of blood cells and the pathogenesis of leukemia. We demonstrated that ZNF268 mRNA was repressed in HTLV-1-infected cells. We also showed that stable and transient expression of HTLV-1 Tax led to repression of ZNF268. In addition, by using reporter constructs that bear the human ZNF268 promoter and its mutants, we showed that Tax repressed ZNF268 promoter in a process dependent on a functional cAMP-responsive element. By using Tax, cAMP-responsive element-binding protein (CREB)-1, CREB-2, and their mutants, we further showed that Tax repressed ZNF268 through the CREB/activating transcription factor pathway. Electrophoretic mobility shift assays and chromatin immunoprecipitation demonstrated the formation of the complex of Tax.CREB-1 directly at the cAMP-responsive element both in vitro and in vivo. These findings suggest a role for ZNF268 in aberrant T-cell proliferation observed in HTLV-1-associated diseases. PMID:18375384

  15. Infiltrating T cells promote prostate cancer metastasis via modulation of FGF11→miRNA-541→androgen receptor (AR)→MMP9 signaling.

    PubMed

    Hu, Shuai; Li, Lei; Yeh, Shuyuan; Cui, Yun; Li, Xin; Chang, Hong-Chiang; Jin, Jie; Chang, Chawnshang

    2015-01-01

    Early clinical studies suggested infiltrating T cells might be associated with poor outcomes in prostate cancer (PCa) patients. The detailed mechanisms how T cells contribute to PCa progression, however, remained unclear. Here, we found PCa cells have a better capacity to recruit more CD4(+) T cells than the surrounding normal prostate cells via secreting more chemokines-CXCL9. The consequences of more recruited CD4(+) T cells to PCa might then lead to enhance PCa cell invasion. Mechanism dissection revealed that infiltrating CD4(+) T cells might function through the modulation of FGF11→miRNA-541 signals to suppress PCa androgen receptor (AR) signals. The suppressed AR signals might then alter the MMP9 signals to promote the PCa cell invasion. Importantly, suppressed AR signals via AR-siRNA or anti-androgen Enzalutamide in PCa cells also enhanced the recruitment of T cells and the consequences of this positive feed back regulation could then enhance the PCa cell invasion. Targeting these newly identified signals via FGF11-siRNA, miRNA-541 inhibitor or MMP9 inhibitor all led to partially reverse the enhanced PCa cell invasion. Results from in vivo mouse models also confirmed the in vitro cell lines in co-culture studies. Together, these results concluded that infiltrating CD4(+) T cells could promote PCa metastasis via modulation of FGF11→miRNA-541→AR→MMP9 signaling. Targeting these newly identified signals may provide us a new potential therapeutic approach to better battle PCa metastasis. PMID:25135278

  16. Chimeric Antigen Receptors With Mutated IgG4 Fc Spacer Avoid Fc Receptor Binding and Improve T Cell Persistence and Antitumor Efficacy

    PubMed Central

    Jonnalagadda, Mahesh; Mardiros, Armen; Urak, Ryan; Wang, Xiuli; Hoffman, Lauren J; Bernanke, Alyssa; Chang, Wen-Chung; Bretzlaff, William; Starr, Renate; Priceman, Saul; Ostberg, Julie R; Forman, Stephen J; Brown, Christine E

    2015-01-01

    The success of adoptive therapy using chimeric antigen receptor (CAR)–expressing T cells partly depends on optimal CAR design. CARs frequently incorporate a spacer/linker region based on the constant region of either IgG1 or IgG4 to connect extracellular ligand-binding with intracellular signaling domains. Here, we evaluated the potential for the IgG4-Fc linker to result in off-target interactions with Fc gamma receptors (FcγRs). As proof-of-principle, we focused on a CD19-specific scFv-IgG4-CD28-zeta CAR and found that, in contrast to CAR-negative cells, CAR+ T cells bound soluble FcγRs in vitro and did not engraft in NSG mice. We hypothesized that mutations to avoid FcγR binding would improve CAR+ T cell engraftment and antitumor efficacy. Thus, we generated CD19-specific CARs with IgG4-Fc spacers that had either been mutated at two sites (L235E; N297Q) within the CH2 region (CD19R(EQ)) or incorporated a CH2 deletion (CD19Rch2Δ). These mutations reduced binding to soluble FcγRs without altering the ability of the CAR to mediate antigen-specific lysis. Importantly, CD19R(EQ) and CD19Rch2Δ T cells exhibited improved persistence and more potent CD19-specific antilymphoma efficacy in NSG mice. Together, these studies suggest that optimal CAR function may require the elimination of cellular FcγR interactions to improve T cell persistence and antitumor responses. PMID:25366031

  17. Nuclear lymphocyte-specific protein tyrosine kinase and its interaction with CR6-interacting factor 1 promote the survival of human leukemic T cells

    PubMed Central

    VAHEDI, SHAHROOZ; CHUEH, FU-YU; DUTTA, SUJOY; CHANDRAN, BALA; YU, CHAO-LAN

    2015-01-01

    Overexpression and hyperactivation of lymphocyte-specific protein tyrosine kinase (Lck) have been associated with leukemia development. We previously showed that, other than its known function as a cytoplasmic signal transducer, Lck also acts as a nuclear transcription factor in mouse leukemic cells. In the present study, we demonstrated the presence of nuclear Lck in human leukemic T cells and in primary cells. We further established a positive correlation between Lck nuclear localization and its kinase activity. Proteomic analysis identified CR6-interacting factor 1 (CRIF1) as one of the Lck-interacting proteins. CRIF1 and Lck association in the nucleus was confirmed both by immunofluorescence microscopy and co-immunoprecipitation in human leukemic T cells. Close-range interaction between Lck and CRIF1 was validated by in situ proximity ligation assay (PLA). Consistent with the role of nuclear CRIF1 as a tumor suppressor, CRIF1 silencing promotes leukemic T cell survival in the absence of growth factors. This protective effect can be recapitulated by endogenous Lck or reconstituted Lck in leukemic T cells. All together, our results support a novel function of nuclear Lck in promoting human leukemic T cell survival through interaction with a tumor suppressor. It has important implications in defining a paradigm shift of non-canonical protein tyrosine kinase signaling. PMID:25997448

  18. Kaposi's-sarcoma-associated-herpesvirus-activated dendritic cells promote HIV-1 trans-infection and suppress CD4{sup +} T cell proliferation

    SciTech Connect

    Liu, Wan; Qin, Yan; Bai, Lei; Lan, Ke; Wang, Jian-Hua

    2013-06-05

    Infection of Kaposi's sarcoma-associated herpesvirus (KSHV) is commonly occurred in AIDS patients. KSHV and HIV-1 act cooperatively in regulating infection with each other and in human carcinogenesis. Dendritic cells (DCs), as the pivotal cells in host immunity, may be modulated by both viruses, for immunoevasion and dissemination, therefore, the interaction between DCs and each virus has been a prior focus for pathogenesis elucidation. Here, we assessed the potential effect of KSHV on DC–HIV-1 interaction. We found that KSHV stimulation could promote maturation of monocyte-derived DCs (MDDCs) and impaired the ability of MDDCs to drive proliferation of resting CD4{sup +} T cells, demonstrating the immunosuppression induced by KSHV. More importantly, KSHV-stimulated MDDCs could capture more HIV-1 and efficiently transferred these infectious viruses to Hut/CCR5 T cell line. Our results reveal the novel modulation of DC-mediated HIV-1 dissemination by KSHV, and highlight the importance of studying DC–HIV-1 interaction to elucidate HIV/AIDS pathogenesis. - Highlights: ► KSHV impaired the ability of MDDCs to drive proliferation of resting CD4{sup +} T cells. ► KSHV stimulation matured MDDCs and enhanced HIV-1 endocytosis. ► KSHV stimulated MDDCs increased ICAM-1 expression and tighten contact with T cells. ► KSHV-stimulated MDDCs promoted HIV-1 trans-infection of CD4{sup +} T cells.

  19. Neisseria gonorrhoeae triggers the PGE2/IL-23 pathway and promotes IL-17 production by human memory T cells.

    PubMed

    Stefanelli, Paola; Teloni, Raffaela; Carannante, Anna; Mariotti, Sabrina; Nisini, Roberto; Gagliardi, Maria Cristina

    2012-10-01

    PGE2 is a potent modulator of the T helper (Th)17 immune response that plays a critical role in the host defense against bacterial, fungal and viral infections. We recently showed high serum levels of interleukin (IL)-17 in patients with gonococcal infection and we hypothesized that Neisseria gonorrhoeae could exploit a PGE2 mediated mechanism to promote IL-17 production. Here we show that N. gonorrhoeae induces human dendritic cell (DC) maturation, secretion of prostaglandin E2 and proinflammatory cytokines, including the pro-Th17 cytokine IL-23. Blocking PGE2 endogenous synthesis selectively reduces IL-23 production by DC in response to gonococcal stimulation, confirming recent data on PGE2/IL-23 crosstalk. N. gonorrhoeae stimulated DC induce a robust IL-17 production by memory CD4(+) T cells and this function correlates with PGE2 production. Our findings delineate a previously unknown role for PGE2 in the immune response to N. gonorrhoeae, suggesting its contribute via Th17 cell expansion. PMID:22542425

  20. MAP Kinase Inhibition Promotes T Cell and Anti-tumor Activity in Combination with PD-L1 Checkpoint Blockade.

    PubMed

    Ebert, Peter J R; Cheung, Jeanne; Yang, Yagai; McNamara, Erin; Hong, Rebecca; Moskalenko, Marina; Gould, Stephen E; Maecker, Heather; Irving, Bryan A; Kim, Jeong M; Belvin, Marcia; Mellman, Ira

    2016-03-15

    Targeted inhibition of mitogen-activated protein kinase (MAPK) kinase (MEK) can induce regression of tumors bearing activating mutations in the Ras pathway but rarely leads to tumor eradication. Although combining MEK inhibition with T-cell-directed immunotherapy might lead to more durable efficacy, T cell responses are themselves at least partially dependent on MEK activity. We show here that MEK inhibition did profoundly block naive CD8(+) T cell priming in tumor-bearing mice, but actually increased the number of effector-phenotype antigen-specific CD8(+) T cells within the tumor. MEK inhibition protected tumor-infiltrating CD8(+) T cells from death driven by chronic TCR stimulation while sparing cytotoxic activity. Combining MEK inhibition with anti-programmed death-ligand 1 (PD-L1) resulted in synergistic and durable tumor regression even where either agent alone was only modestly effective. Thus, despite the central importance of the MAP kinase pathway in some aspects of T cell function, MEK-targeted agents can be compatible with T-cell-dependent immunotherapy. PMID:26944201

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

    PubMed Central

    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

  2. Antigen targeting to CD11b+ dendritic cells in association with TLR4/TRIF signaling promotes strong CD8+ T cell responses.

    PubMed

    Dadaglio, Gilles; Fayolle, Catherine; Zhang, Xiaoming; Ryffel, Bernard; Oberkampf, Marine; Felix, Tristan; Hervas-Stubbs, Sandra; Osicka, Radim; Sebo, Peter; Ladant, Daniel; Leclerc, Claude

    2014-08-15

    Deciphering the mechanisms that allow the induction of strong immune responses is crucial to developing efficient vaccines against infectious diseases and cancer. Based on the discovery that the adenylate cyclase from Bordetella pertussis binds to the CD11b/CD18 integrin, we developed a highly efficient detoxified adenylate cyclase-based vector (CyaA) capable of delivering a large variety of Ags to the APC. This vector allows the induction of protective and therapeutic immunity against viral and tumoral challenges as well as against transplanted tumors in the absence of any added adjuvant. Two therapeutic vaccine candidates against human papilloma viruses and melanoma have been developed recently, based on the CyaA vector, and are currently in clinical trials. We took advantage of one of these highly purified vaccines, produced under good manufacturing practice-like conditions, to decipher the mechanisms by which CyaA induces immune responses. In this study, we demonstrate that CyaA binds both human and mouse CD11b(+) dendritic cells (DCs) and induces their maturation, as shown by the upregulation of costimulatory and MHC molecules and the production of proinflammatory cytokines. Importantly, we show that DCs sense CyaA through the TLR4/Toll/IL-1R domain-containing adapter-inducing IFN-β pathway, independent of the presence of LPS. These findings show that CyaA possesses the intrinsic ability to not only target DCs but also to activate them, leading to the induction of strong immune responses. Overall, this study demonstrates that Ag delivery to CD11b(+) DCs in association with TLR4/Toll/IL-1R domain-containing adapter-inducing IFN-β activation is an efficient strategy to promote strong specific CD8(+) T cell responses. PMID:25024388

  3. CCR10 regulates balanced maintenance and function of resident regulatory and effector T cells to promote immune homeostasis in skin

    PubMed Central

    Xia, Mingcan; Hu, Shaomin; Fu, Yaoyao; Jin, Wensen; Yi, Qiyi; Matsui, Yurika; Yang, Jie; McDowell, Mary Ann; Sarkar, Surojit; Kalia, Vandana; Xiong, Na

    2014-01-01

    Background CCR10 and CCL27 are the most skin-specific chemokine receptor/ligand pair implicated in skin allergy and inflammatory diseases including atopic dermatitis and psoriasis. This pair is thought to regulate migration and/or maintenance of skin T cells and suggested as therapeutic targets for treatment of skin diseases. However, the functional importance of CCR10/CCL27 in vivo remains elusive. Objective We sought to determine expression and function of CCR10 in different subsets of skin T cells under both homeostatic and inflammatory conditions to gain a mechanistic insight into potential roles of CCR10 during skin inflammation. Methods Using heterozygous and homozygous CCR10-knockout/EGFP-knockin mice, we assessed expression of CCR10 on regulatory and effector T cells of healthy and inflamed skin induced by chemicals, pathogens and auto-reactive T cells. In addition, we assessed the effect of CCR10-knockout on the maintenance and functions of different T cells and inflammatory status in the skin during different phases of the immune response. Results CCR10 expression is preferentially induced on memory-like skin-resident T cells and their progenitors for their maintenance in homeostatic skin but not expressed on most skin-infiltrating effector T cells during inflammation. In CCR10-knockout mice, the imbalanced presence and dysregulated function of resident regulatory and effector T cells result in over-reactive and prolonged innate and memory responses in the skin, leading to increased clearance of Leishmamia infection in the skin. Conclusion CCR10 is a critical regulator of skin immune homeostasis. PMID:24767879

  4. Neuroinvasive West Nile Infection Elicits Elevated and Atypically Polarized T Cell Responses That Promote a Pathogenic Outcome

    PubMed Central

    James, Eddie A.; Gates, Theresa J.; LaFond, Rebecca E.; Yamamoto, Shinobu; Ni, Chester; Mai, Duy; Gersuk, Vivian H.; O’Brien, Kimberly; Nguyen, Quynh-Anh; Zeitner, Brad; Lanteri, Marion C.; Norris, Philip J.; Chaussabel, Damien; Malhotra, Uma; Kwok, William W.

    2016-01-01

    Most West Nile virus (WNV) infections are asymptomatic, but some lead to neuroinvasive disease with symptoms ranging from disorientation to paralysis and death. Evidence from animal models suggests that neuroinvasive infections may arise as a consequence of impaired immune protection. However, other data suggest that neurologic symptoms may arise as a consequence of immune mediated damage. We demonstrate that elevated immune responses are present in neuroinvasive disease by directly characterizing WNV-specific T cells in subjects with laboratory documented infections using human histocompatibility leukocyte antigen (HLA) class II tetramers. Subjects with neuroinvasive infections had higher overall numbers of WNV-specific T cells than those with asymptomatic infections. Independent of this, we also observed age related increases in WNV-specific T cell responses. Further analysis revealed that WNV-specific T cell responses included a population of atypically polarized CXCR3+CCR4+CCR6- T cells, whose presence was highly correlated with neuroinvasive disease. Moreover, a higher proportion of WNV-specific T cells in these subjects co-produced interferon-γ and interleukin 4 than those from asymptomatic subjects. More globally, subjects with neuroinvasive infections had reduced numbers of CD4+FoxP3+ Tregs that were CTLA4 positive and exhibited a distinct upregulated transcript profile that was absent in subjects with asymptomatic infections. Thus, subjects with neuroinvasive WNV infections exhibited elevated, dysregulated, and atypically polarized responses, suggesting that immune mediated damage may indeed contribute to pathogenic outcomes. PMID:26795118

  5. DHEA promotes osteoblast differentiation by regulating the expression of osteoblast-related genes and Foxp3(+) regulatory T cells.

    PubMed

    Qiu, Xuemin; Gui, Yuyan; Xu, Yingping; Li, Dajin; Wang, Ling

    2015-10-01

    Several studies have reported that dehydroepiandrosterone (DHEA) promotes osteoblast proliferation and inhibits osteoblast apoptosis and that DHEA inhibits osteoclast maturation. However, whether DHEA regulates osteoblast differentiation remains unclear. The present study first examined the effect of DHEA on bone morphology in vivo. DHEA was found to increase bone volume (BV), bone mineral density (BMD), and the number of trabeculae in bone (Th.N) and it was found to decrease trabecular spacing in bone (Th.sp) in ovariectomized (OVX) mice. Next, the effect of DHEA on osteoblast differentiation was examined in vitro and osteoblastogenesis-related marker genes, such as Runx2, Osterix, Collagen1, and Osteocalcin, were also detected. DHEA increased osteoblast production in mesenchymal stem cells (MSCs) cultured in osteoblastogenic medium, and DHEA increased the expression of Runx2 and osterix, thereby increasing the expression of osteocalcin and collagen1. Immune cells and bone interact, so changes in immune cells were detected in vivo. DHEA increased the number of Foxp3(+) regulatory T cells (Tregs) in the spleen but it did not affect CTLA-4 or IL-10. When MSCs were treated with DHEA in the presence of Tregs, alkaline phosphatase (ALP) activity increased. Osteoblasts and adipocytes are both generated by MSCs. If osteoblast differentiation increases, adipocyte differentiation will decrease, and the reverse also holds true. DHEA was found to increase the number of adipocytes in osteoblastogenic medium but it had no effect on the number of adipocytes and expression of PPARγ mRNA in adipogenic medium. This finding suggests that osteoblasts may be involved in adipocyte production. In conclusion, the current results suggest that DHEA can improve postmenopausal osteoporosis (PMO) by up-regulating osteoblast differentiation via the up-regulation of the expression of osteoblastogenesis-related genes and via an increase in Foxp3(+) Tregs. PMID:26559023

  6. Autoreactive natural killer T cells: promoting immune protection and immune tolerance through varied interactions with myeloid antigen-presenting cells

    PubMed Central

    Hegde, Subramanya; Fox, Lisa; Wang, Xiaohua; Gumperz, Jenny E

    2010-01-01

    Natural killer T (NKT) cells are innate T lymphocytes that are restricted by CD1d antigen-presenting molecules and recognize lipids and glycolipids as antigens. NKT cells have attracted attention for their potent immunoregulatory effects. Like other types of regulatory lymphocytes, a high proportion of NKT cells appear to be autoreactive to self antigens. Thus, as myeloid antigen-presenting cells (APCs) such as monocytes, dendritic cells (DCs) and myeloid-derived suppressor cells (MDSCs) constitutively express CD1d, NKT cells are able to interact with these APCs not only during times of immune activation but also in immunologically quiescent periods. The interactions of NKT cells with myeloid APCs can have either pro-inflammatory or tolerizing outcomes, and a central question is how the ensuing response is determined. Here we bring together published results from a variety of model systems to highlight three critical factors that influence the outcome of the NKT–APC interaction: (i) the strength of the antigenic signal delivered to the NKT cell, as determined by antigen abundance and/or T-cell receptor (TCR) affinity; (ii) the presence or absence of cytokines that costimulate NKT cells [e.g. interleukin (IL)-12, IL-18 and interferon (IFN)-α]; (iii) APC intrinsic factors such as differentiation state (e.g. monocyte versus DC) and Toll-like receptor (TLR) stimulation. Together with recent findings that demonstrate new links between NKT cell activation and endogenous lipid metabolism, these results outline a picture in which the functions of NKT cells are closely attuned to the existing biological context. Thus, NKT cells may actively promote tolerance until a critical level of danger signals arises, at which point they switch to activating pro-inflammatory immune responses. PMID:20465577

  7. The human fetal placenta promotes tolerance against the semiallogeneic fetus by inducing regulatory T cells and homeostatic M2 macrophages.

    PubMed

    Svensson-Arvelund, Judit; Mehta, Ratnesh B; Lindau, Robert; Mirrasekhian, Elahe; Rodriguez-Martinez, Heriberto; Berg, Göran; Lash, Gendie E; Jenmalm, Maria C; Ernerudh, Jan

    2015-02-15

    A successful pregnancy requires that the maternal immune system is instructed to a state of tolerance to avoid rejection of the semiallogeneic fetal-placental unit. Although increasing evidence supports that decidual (uterine) macrophages and regulatory T cells (Tregs) are key regulators of fetal tolerance, it is not known how these tolerogenic leukocytes are induced. In this article, we show that the human fetal placenta itself, mainly through trophoblast cells, is able to induce homeostatic M2 macrophages and Tregs. Placental-derived M-CSF and IL-10 induced macrophages that shared the CD14(+)CD163(+)CD206(+)CD209(+) phenotype of decidual macrophages and produced IL-10 and CCL18 but not IL-12 or IL-23. Placental tissue also induced the expansion of CD25(high)CD127(low)Foxp3(+) Tregs in parallel with increased IL-10 production, whereas production of IFN-γ (Th1), IL-13 (Th2), and IL-17 (Th17) was not induced. Tregs expressed the suppressive markers CTLA-4 and CD39, were functionally suppressive, and were induced, in part, by IL-10, TGF-β, and TRAIL. Placental-derived factors also limited excessive Th cell activation, as shown by decreased HLA-DR expression and reduced secretion of Th1-, Th2-, and Th17-associated cytokines. Thus, our data indicate that the fetal placenta has a central role in promoting the homeostatic environment necessary for successful pregnancy. These findings have implications for immune-mediated pregnancy complications, as well as for our general understanding of tissue-induced tolerance. PMID:25560409

  8. Reduced Epithelial Na+/H+ Exchange Drives Gut Microbial Dysbiosis and Promotes Inflammatory Response in T Cell-Mediated Murine Colitis.

    PubMed

    Laubitz, Daniel; Harrison, Christy A; Midura-Kiela, Monica T; Ramalingam, Rajalakshmy; Larmonier, Claire B; Chase, John H; Caporaso, J Gregory; Besselsen, David G; Ghishan, Fayez K; Kiela, Pawel R

    2016-01-01

    Inflammatory bowel diseases (IBD) are associated with functional inhibition of epithelial Na+/H+ exchange. In mice, a selective disruption of NHE3 (Slc9a3), a major apical Na+/H+ exchanger, also promotes IBD-like symptoms and gut microbial dysbiosis. We hypothesized that disruption of Na+/H+ exchange is necessary for the development of dysbiosis, which promotes an exacerbated mucosal inflammatory response. Therefore, we performed a temporal analysis of gut microbiota composition, and mucosal immune response to adoptive T cell transfer was evaluated in Rag2-/- and NHE3-/-/Rag2-/- (DKO) mice with and without broad-spectrum antibiotics. Microbiome (16S profiling), colonic histology, T cell and neutrophil infiltration, mucosal inflammatory tone, and epithelial permeability were analyzed. In adoptive T cell transfer colitis model, Slc9a3 status was the most significant determinant of gut microbial community. In DKO mice, NHE3-deficiency and dysbiosis were associated with dramatically accelerated and exacerbated disease, with rapid body weight loss, increased mucosal T cell and neutrophil influx, increased mucosal cytokine expression, increased permeability, and expansion of CD25-FoxP3+ Tregs; this enhanced susceptibility was alleviated by oral broad-spectrum antibiotics. Based on these results and our previous work, we postulate that epithelial electrolyte homeostasis is an important modulator in the progression of colitis, acting through remodeling of the gut microbial community. PMID:27050757

  9. Reduced Epithelial Na+/H+ Exchange Drives Gut Microbial Dysbiosis and Promotes Inflammatory Response in T Cell-Mediated Murine Colitis

    PubMed Central

    Midura-Kiela, Monica T.; Ramalingam, Rajalakshmy; Larmonier, Claire B.; Chase, John H.; Caporaso, J. Gregory; Besselsen, David G.; Ghishan, Fayez K.; Kiela, Pawel R.

    2016-01-01

    Inflammatory bowel diseases (IBD) are associated with functional inhibition of epithelial Na+/H+ exchange. In mice, a selective disruption of NHE3 (Slc9a3), a major apical Na+/H+ exchanger, also promotes IBD-like symptoms and gut microbial dysbiosis. We hypothesized that disruption of Na+/H+ exchange is necessary for the development of dysbiosis, which promotes an exacerbated mucosal inflammatory response. Therefore, we performed a temporal analysis of gut microbiota composition, and mucosal immune response to adoptive T cell transfer was evaluated in Rag2-/- and NHE3-/-/Rag2-/- (DKO) mice with and without broad-spectrum antibiotics. Microbiome (16S profiling), colonic histology, T cell and neutrophil infiltration, mucosal inflammatory tone, and epithelial permeability were analyzed. In adoptive T cell transfer colitis model, Slc9a3 status was the most significant determinant of gut microbial community. In DKO mice, NHE3-deficiency and dysbiosis were associated with dramatically accelerated and exacerbated disease, with rapid body weight loss, increased mucosal T cell and neutrophil influx, increased mucosal cytokine expression, increased permeability, and expansion of CD25-FoxP3+ Tregs; this enhanced susceptibility was alleviated by oral broad-spectrum antibiotics. Based on these results and our previous work, we postulate that epithelial electrolyte homeostasis is an important modulator in the progression of colitis, acting through remodeling of the gut microbial community. PMID:27050757

  10. RNA Binding of T-cell Intracellular Antigen-1 (TIA-1) C-terminal RNA Recognition Motif Is Modified by pH Conditions*

    PubMed Central

    Cruz-Gallardo, Isabel; Aroca, Ángeles; Persson, Cecilia; Karlsson, B. Göran; Díaz-Moreno, Irene

    2013-01-01

    T-cell intracellular antigen-1 (TIA-1) is a DNA/RNA-binding protein that regulates critical events in cell physiology by the regulation of pre-mRNA splicing and mRNA translation. TIA-1 is composed of three RNA recognition motifs (RRMs) and a glutamine-rich domain and binds to uridine-rich RNA sequences through its C-terminal RRM2 and RRM3 domains. Here, we show that RNA binding mediated by either isolated RRM3 or the RRM23 construct is controlled by slight environmental pH changes due to the protonation/deprotonation of TIA-1 RRM3 histidine residues. The auxiliary role of the C-terminal RRM3 domain in TIA-1 RNA recognition is poorly understood, and this work provides insight into its binding mechanisms. PMID:23902765

  11. MicroRNA-184 Promotes Proliferation and Inhibits Apoptosis in HaCaT Cells: An In Vitro Study.

    PubMed

    Bi, Xiaodong; Cao, Yu; Chen, Rixin; Liu, Chengyin; Chen, Jinghong; Min, Dongfang

    2016-01-01

    BACKGROUND This study aimed to investigate the role of miR-184 in the proliferation and apoptosis of keratinocyte (HaCaT cells). MATERIAL AND METHODS HaCaT cells were cultured in a growth medium. The miR-184 was transfected with siRNA, then cell viability and apoptosis were assayed by MTT and flow cytometry, respectively. The colony-forming efficacy of HaCaT cells were detected as well. mRNA expressions of basic fibroblast growth factor (bFGF) and transforming growth factor (TGF)-β1 were measured with RT-PCR. The expressions of apoptosis-related proteins caspase-3 and Bcl-x in HaCaT cells were determined by Western blot. RESULTS After miR-184 was transfected with siRNA, cell viability and colony forming ability decreased significantly, and apoptosis was significantly increased. The expressions of growth factors TGF-β1 and bFGF mRNAs, as well as apoptosis-related proteins Bcl-x, in HaCaT cells declined significantly after miR-184 was transfected with siRNA. In addition, the expression of pro-apoptotic protein caspase-3 increased significantly. CONCLUSIONS Our results suggest distinct roles of miR-184 during the growth, proliferation, and apoptosis of keratinocytes. PMID:27571235

  12. Binding immunoglobulin protein-treated peripheral blood monocyte-derived dendritic cells are refractory to maturation and induce regulatory T-cell development.

    PubMed

    Corrigall, Valerie M; Vittecoq, Olivier; Panayi, Gabriel S

    2009-10-01

    Binding immunoglobulin protein (BiP) has been shown previously to have immunomodulatory functions. Herein we investigated whether BiP could affect the differentiation of monocytes into dendritic cells (DCs) and thence the development of regulatory T cells. Peripheral blood monocyte-derived DCs were matured with lipopolysaccharide in the presence or absence of BiP. DC development and T-cell changes were monitored by flow cytometry and regulatory T-cell function was measured by uptake of tritiated thymidine. More BiP-treated DCs (DC((BiP))s) expressed amounts of intracellular indoleamine 2,3-dioxygenase (IDO) and cell surface leucocyte immunoglobulin-like receptor subfamily B member 1 (LILRB1), retained CD14 expression but down-regulated expression of human leucocyte antigen (HLA)-DR and CD86, and produced copious amounts of interleukin (IL)-10, when compared with control DCs. T cells co-cultured with DC((BiP))s developed regulatory function with increased surface expression of CD4(+) CD25(hi) CD27(hi) but with no concomitant increase in forkhead box P3 (Foxp3). These T cells also showed significantly higher levels of intracellular cytotoxic T-lymphocyte antigen (CTLA)-4. The latter could be inhibited by the presence of the IDO inhibitor 1 methyl tryptophan. The addition of neutralizing anti-IL-10 antibody or the specific mitogen-activated protein kinase (MAPK) p38 inhibitor SB203580 reversed the inhibition of DC differentiation by BiP. In conclusion, BiP is an immunomodulator able to arrest inflammation through induction of tolerogenic DCs and subsequent generation of T regulatory cells. PMID:19740378

  13. The NF-kappa B binding site is necessary for efficient replication of simian immunodeficiency virus of macaques in primary macrophages but not in T cells in vitro.

    PubMed Central

    Bellas, R E; Hopkins, N; Li, Y

    1993-01-01

    We demonstrate here that the nuclear factor-kappa B (NF-kappa B) binding site in the simian immunodeficiency virus (SIVmac) long terminal repeat is essential for efficient virus replication in primary alveolar macrophages but dispensable for efficient replication in primary T cells. Mutation of the NF-kappa B site does not seriously impair replication of a T-cell-tropic SIVmac239 or a macrophagetropic SIVmacEm* in peripheral blood lymphocytes or established CD4+ cell lines; however, mutation of the NF-kappa B site prevents efficient SIVmacEm* replication in primary alveolar macrophages. These data suggest that efficient replication in primary macrophages requires both envelope and long terminal repeat determinants. Images PMID:8474179

  14. Triple synergism of human T-lymphotropic virus type 1-encoded tax, GATA-binding protein, and AP-1 is required for constitutive expression of the interleukin-5 gene in adult T-cell leukemia cells.

    PubMed Central

    Yamagata, T; Mitani, K; Ueno, H; Kanda, Y; Yazaki, Y; Hirai, H

    1997-01-01

    Accumulated evidence demonstrates that adult T-cell leukemia (ATL) is frequently associated with eosinophilia, and human T-lymphotropic virus type 1 (HTLV-1)-infected cells frequently express interleukin-5 (IL-5). However, the molecular mechanism of constitutive IL-5 expression in HTLV-1-infected cells remains unclear. To clarify the mechanism of aberrant IL-5 expression in HTLV-1-infected cells, we investigated the response of the human IL-5 promoter to the HTLV-1-encoded protein Tax. Cotransfection experiments using Jurkat cells revealed that Tax is incapable of activating the IL-5 promoter by itself but that it synergistically transactivates the promoter with GATA-binding protein (GATA-4) and 12-O-tetradecanoylphorbol-13-acetate (TPA) stimulation. By introducing a series of mutations within the IL-5 promoter, we found that conserved lymphokine element 0 (CLE0) is responsible for mediating the signal induced by Tax-TPA. A deletion construct of the promoter indicated that the -75 GATA element and CLE0 are sufficient to mediate synergistic activation of the IL-5 promoter. Electrophoretic mobility shift assays using Jurkat cell nuclear extracts demonstrated that TPA induces a transcription factor to bind CLE0, and an experiment using JPX-9 cell nuclear extracts showed that Tax enhances this binding activity. An antibody supershift experiment revealed that this band consists of c-Jun and JunD. However, among the Jun family members, only c-Jun is able to cooperate with Tax and GATA-4 to activate the IL-5 promoter. We have determined the minimum factors required for IL-5 gene activation by reconstituting the IL-5 promoter activity in F9 cells. This is the first report to demonstrate the functional involvement of Tax protein in IL-5 gene regulation and to suggest the functional triple synergism among Tax, GATA-4, and AP-1, which disrupts regulated control of the gene and leads to constitutive expression of the IL-5 gene. PMID:9234684

  15. SLAM-SAP signaling promotes differentiation of IL-17-producing T cells and progression of experimental autoimmune encephalomyelitis.

    PubMed

    Huang, Yu-Hsuan; Tsai, Kevin; Ma, Caixia; Vallance, Bruce A; Priatel, John J; Tan, Rusung

    2014-12-15

    IL-17 plays critical roles in host defenses, combating bacterial and fungal infections, as well as the pathogenesis of autoimmune diseases such as experimental autoimmune encephalomyelitis (EAE). The signaling adaptor SAP is essential for normal immune homeostasis and mutations within SH2D1A, the locus encoding this protein, result in serious and sometimes fatal syndromes, including X-linked lymphoproliferative disease and severe cases of common variable immunodeficiency. However, the precise cellular basis of how SAP deficiency contributes to immune dysfunction remains incompletely understood. In this study, we found that CD4 and CD8 T cells lacking SAP had a diminished capacity to differentiate into IL-17-producing Th17 and T cytotoxic (Tc17) cells relative to wild-type lymphocytes. The use of costimulating SLAM Abs was found to augment the differentiation of IL-17-secreting effectors in wild-type but not Sh2d1a(-/-) splenic T cells under IL-17-polarizing conditions. In addition, SAP's regulation of IL-17-secreting T cells was shown to be a T cell-intrinsic role, as purified naive Sh2d1a(-/-) CD4 and CD8 T cells were inherently defective at converting into Th17 and Tc17 cells in vitro and in vivo. Furthermore, Sh2d1a(-/-) mice were protected from EAE and exhibited greatly decreased numbers of CNS-infiltrating Th17 and Tc17 effector T cells and reduced disease severity. Collectively, these results suggest that SLAM-SAP signaling drives the differentiation and function of Th17 and Tc17 cells in vitro and in vivo and contributes to the pathogenesis of autoimmunity in EAE. PMID:25362182

  16. The Extracellular Domain of CD83 Inhibits Dendritic Cell–mediated T Cell Stimulation and Binds to a Ligand on Dendritic Cells

    PubMed Central

    Lechmann, Matthias; Krooshoop, Daniëlle J.E.B.; Dudziak, Diana; Kremmer, Elisabeth; Kuhnt, Christine; Figdor, Carl G.; Schuler, Gerold; Steinkasserer, Alexander

    2001-01-01

    CD83 is an immunoglobulin (Ig) superfamily member that is upregulated during the maturation of dendritic cells (DCs). It has been widely used as a marker for mature DCs, but its function is still unknown. To approach its potential functional role, we have expressed the extracellular Ig domain of human CD83 (hCD83ext) as a soluble protein. Using this tool we could show that immature as well as mature DCs bind to CD83. Since CD83 binds a ligand also expressed on immature DCs, which do not express CD83, indicates that binding is not a homophilic interaction. In addition we demonstrate that hCD83ext interferes with DC maturation downmodulating the expression of CD80 and CD83, while no phenotypical effects were observed on T cells. Finally, we show that hCD83ext inhibits DC-dependent allogeneic and peptide-specific T cell proliferation in a concentration dependent manner in vitro. This is the first report regarding functional aspects of CD83 and the binding of CD83 to DCs. PMID:11748282

  17. Structure and Function of a Fungal Adhesin that Binds Heparin and Mimics Thrombospondin-1 by Blocking T Cell Activation and Effector Function

    PubMed Central

    Brandhorst, T. Tristan; Roy, René; Wüthrich, Marcel; Nanjappa, Som; Filutowicz, Hanna; Galles, Kevin; Tonelli, Marco; McCaslin, Darrell R.; Satyshur, Kenneth; Klein, Bruce

    2013-01-01

    Blastomyces adhesin-1 (BAD-1) is a 120-kD surface protein on B. dermatitidis yeast. We show here that BAD-1 contains 41 tandem repeats and that deleting even half of them impairs fungal pathogenicity. According to NMR, the repeats form tightly folded 17-amino acid loops constrained by a disulfide bond linking conserved cysteines. Each loop contains a highly conserved WxxWxxW motif found in thrombospondin-1 (TSP-1) type 1 heparin-binding repeats. BAD-1 binds heparin specifically and saturably, and is competitively inhibited by soluble heparin, but not related glycosaminoglycans. According to SPR analysis, the affinity of BAD-1 for heparin is 33 nM±14 nM. Putative heparin-binding motifs are found both at the N-terminus and within each tandem repeat loop. Like TSP-1, BAD-1 blocks activation of T cells in a manner requiring the heparan sulfate-modified surface molecule CD47, and impairs effector functions. The tandem repeats of BAD-1 thus confer pathogenicity, harbor motifs that bind heparin, and suppress T-cell activation via a CD47-dependent mechanism, mimicking mammalian TSP-1. PMID:23853587

  18. IFNγ Signaling Endows DCs with the Capacity to Control Type I Inflammation during Parasitic Infection through Promoting T-bet+ Regulatory T Cells

    PubMed Central

    Lee, Hyang-Mi; Fleige, Anne; Forman, Ruth; Cho, Sunglim; Khan, Aly Azeem; Lin, Ling-Li; Nguyen, Duc T.; O'Hara-Hall, Aisling; Yin, Zhinan; Hunter, Christopher A.; Muller, Werner; Lu, Li-Fan

    2015-01-01

    IFNγ signaling drives dendritic cells (DCs) to promote type I T cell (Th1) immunity. Here, we show that activation of DCs by IFNγ is equally crucial for the differentiation of a population of T-bet+ regulatory T (Treg) cells specialized to inhibit Th1 immune responses. Conditional deletion of IFNγ receptor in DCs but not in Treg cells resulted in a severe defect in this specific Treg cell subset, leading to exacerbated immune pathology during parasitic infections. Mechanistically, IFNγ-unresponsive DCs failed to produce sufficient amount of IL-27, a cytokine required for optimal T-bet induction in Treg cells. Thus, IFNγ signalling endows DCs with the ability to efficiently control a specific type of T cell immunity through promoting a corresponding Treg cell population. PMID:25658840

  19. Interleukin-21 is associated with the severity of psoriasis vulgaris through promoting CD4+ T cells to differentiate into Th17 cells

    PubMed Central

    Wang, Ying; Wang, Li-Li; Yang, Hao-Yu; Wang, Fei-Fei; Zhang, Xue-Xiu; Bai, Yan-Ping

    2016-01-01

    Interleukin-21 (IL-21) and T helper 17 (Th17) cells are known to be involved in the pathogenesis of psoriasis, but little is known about their relationship in psoriasis. Herein, we investigated whether IL-21 could regulate Th17 cell induction in patients with psoriasis vulgaris. 32 patients with psoriasis vulgaris and 13 healthy controls were recruited. Flow cytometry was used to detect the frequencies of cells mainly secreting IL-21 (including IL-21+CD4+ T and IL-21+ Th17 cells) and Th17 cells. An enzyme-linked immunosorbent assay (ELISA) was used to determine the serum content of IL-21. Severity of the psoriasis was evaluated by a Psoriasis Area and Severity Index (PASI) score. In addition, the differentiation of CD4+ T cells with IL-21 and the different frequencies of IL-21+CD4+ T cells, IL-21+ Th17 cells and Th17 cells were assessed, as were serum levels of IL-21 in patients with moderate to severe psoriasis before and after treatment. Our results showed that the levels of IL-21, IL-21+CD4+ T cells, IL-21+ Th17 cells and Th17 cells were significantly increased in patients and positively associated with PASI score (P < 0.01). Moreover, the levels of IL-21, IL-21+CD4+ T cells and IL-21+ Th17 cells were positively correlated with the frequency of Th17 cells (P < 0.01). In vitro experiments demonstrated that IL-21 could promote CD4+ T cells to differentiate into Th17 cells. After a 4-week treatment of acitretin and a topical therapy, all the immune markers observed in patients decreased significantly (P < 0.01), but the levels remained higher than those in healthy controls (P < 0.01). These findings indicate that IL-21 might promote Th17 cell induction in psoriasis and might be a potential immune marker for targeting this disease. PMID:27508040

  20. Biphasic function of TLR3 adjuvant on tumor and spleen dendritic cells promotes tumor T cell infiltration and regression in a vaccine therapy.

    PubMed

    Azuma, Masahiro; Takeda, Yohei; Nakajima, Hiroko; Sugiyama, Haruo; Ebihara, Takashi; Oshiumi, Hiroyuki; Matsumoto, Misako; Seya, Tsukasa

    2016-08-01

    Successful cancer immunotherapy necessitates T cell proliferation and infiltration into tumor without exhaustion, a process closely links optimal maturation of dendritic cells (DC), and adjuvant promotes this process as an essential prerequisite. Poly(I:C) has contributed to adjuvant immunotherapy that evokes an antitumor response through the Toll-loke receptor 3 (TLR3)/TICAM-1 pathway in DC. However, the mechanism whereby Poly(I:C) acts on DC for T cell proliferation and migration remains undetermined. Subcutaneous injection of Poly(I:C) regressed implant tumors (WT1-C1498 or OVA-EG7) in C57BL/6 mice, which coincided with tumor-infiltration of CD8(+) T cells. Epitope-specific cytotoxic T lymphocytes (CTLs) were increased in spleen by challenge with Poly(I:C)+Db126 WT-1 peptide but not Poly(I:C) alone, suggesting the need of an exogenous Ag density for cross-priming. In tumor, CXCR3 ligands were upregulated by Poly(I:C), which facilitated recruitment of CTL to the tumor. Thus, Poly(I:C) acts on splenic CD8α(+) DC to cross-prime T cells and on intratumor cells to attract CTLs. Besides CD8(+) T cell cross-priming, T cell recruitment into tumor was significantly dampened in Batf3 (-/-) mice, reflecting the importance of tumor Batf3-dependent DC rather than macrophages in T cell recruitment. Poly(I:C)-induced XCR1(hi) CD8α(+) DC with high TLR3 levels were markedly decreased in Batf3 (-/-) mice, which hampered the production of IL-12 and IL-12-mediated CD4(+)/CD8(+) T cell proliferation. Subcutaneous administration of Poly(I:C) and adoptive transfer of wild-type CD8α(+) DC largely recovered antitumor response in those Batf3 (-/-) mice. Collectively, Poly(I:C) tunes up proper maturation of CD8α(+) DC to establish TLR3-mediated IL-12 function and cross-presentation in spleen and lymphocyte-attractive antitumor microenvironment in tumor. PMID:27622060

  1. Increased expression of TLR2 in CD4(+) T cells from SLE patients enhances immune reactivity and promotes IL-17 expression through histone modifications.

    PubMed

    Liu, Yu; Liao, Jieyue; Zhao, Ming; Wu, Haijing; Yung, Susan; Chan, Tak Mao; Yoshimura, Akihiko; Lu, Qianjin

    2015-09-01

    The innate immune system has been shown to play an important pathologic role in systemic lupus erythematosus (SLE). TLR2, a PRR, recognizes exogenous PAMPs, and endogenous damage-associated molecular patterns and has been implicated in the initiation and maintenance of the perpetuated inflammatory reactions in autoimmune diseases. Here, we report increased expression of TLR2 in CD4(+) and CD8(+) T cells, CD19(+) B cells, and CD14(+) monocytes from SLE patients. Conventional treatment, such as hydroxychloroquine and corticosteroids, showed no effect on TLR2 expression in CD4(+) T cells from SLE patients. In vitro stimulation of TLR2 in CD4(+) T cells from SLE patients increased CD40L and CD70 expression, as well as secretion of IL-6, IL-17A, IL-17F, and TNF-α, while Foxp3 transcription decreased. This effect was reversed by TLR2 siRNA. Moreover, TLR2 activation upregulated H3K4 tri-methylation and H4 acetylation levels while downregulated H3K9 tri-methylation level in the IL-17A promoter region. In addition, it also increased H4 acetylation levels and decreased H3K9 tri-methylation levels in the IL-17F promoter region. In summary, our findings demonstrate that increased expression of TLR2 contributes to immune reactivity and promotes IL-17A and IL-17F expression through histone modifications in SLE. PMID:26079624

  2. Gene Expression Driven by a Strong Viral Promoter in MVA Increases Vaccination Efficiency by Enhancing Antibody Responses and Unmasking CD8+ T Cell Epitope

    PubMed Central

    Becker, Pablo D.; Nörder, Miriam; Weissmann, Sebastian; Ljapoci, Ronny; Erfle, Volker; Drexler, Ingo; Guzmán, Carlos A.

    2014-01-01

    Viral vectors are promising tools for vaccination strategies and immunotherapies. However, CD8+ T cell responses against pathogen-derived epitopes are usually limited to dominant epitopes and antibody responses to recombinant encoded antigens (Ags) are mostly weak. We have previously demonstrated that the timing of viral Ag expression in infected professional Ag-presenting cells strongly shapes the epitope immunodominance hierarchy. T cells recognizing determinants derived from late viral proteins have a clear disadvantage to proliferate during secondary responses. In this work we evaluate the effect of overexpressing the recombinant Ag using the modified vaccinia virus early/late promoter H5 (mPH5). Although the Ag-expression from the natural promoter 7.5 (P7.5) and the mPH5 seemed similar, detailed analysis showed that mPH5 not only induces higher expression levels than P7.5 during early phase of infection, but also Ag turnover is enhanced. The strong overexpression during the early phase leads to broader CD8 T cell responses, while preserving the priming efficiency of stable Ags. Moreover, the increase in Ag-secretion favors the induction of strong antibody responses. Our findings provide the rationale to develop new strategies for fine-tuning the responses elicited by recombinant modified vaccinia virus Ankara by using selected promoters to improve the performance of this viral vector. PMID:26344747

  3. Up-regulation of Store-operated Ca2+ Entry and Nuclear Factor of Activated T Cells Promote the Acinar Phenotype of the Primary Human Salivary Gland Cells.

    PubMed

    Jang, Shyh-Ing; Ong, Hwei Ling; Liu, Xibao; Alevizos, Ilias; Ambudkar, Indu S

    2016-04-15

    The signaling pathways involved in the generation and maintenance of exocrine gland acinar cells have not yet been established. Primary human salivary gland epithelial cells, derived from salivary gland biopsies, acquired an acinar-like phenotype when the [Ca(2+)] in the serum-free medium (keratinocyte growth medium, KGM) was increased from 0.05 mm (KGM-L) to 1.2 mm (KGM-H). Here we examined the mechanism underlying this Ca(2+)-dependent generation of the acinar cell phenotype. Compared with cells in KGM-L, those in KGM-H display enhancement of Orai1, STIM1, STIM2, and nuclear factor of activated T cells 1 (NFAT1) expression together with an increase in store-operated Ca(2+) entry (SOCE), SOCE-dependent nuclear translocation of pGFP-NFAT1, and NFAT-dependent but not NFκB-dependent gene expression. Importantly, AQP5, an acinar-specific protein critical for function, is up-regulated in KGM-H via SOCE/NFAT-dependent gene expression. We identified critical NFAT binding motifs in the AQP5 promoter that are involved in Ca(2+)-dependent up-regulation of AQP5. These important findings reveal that the Ca(2+)-induced switch of salivary epithelial cells to an acinar-like phenotype involves remodeling of SOCE and NFAT signaling, which together control the expression of proteins critically relevant for acinar cell function. Our data provide a novel strategy for generating and maintaining acinar cells in culture. PMID:26903518

  4. Gamma delta T cells promote inflammation and insulin resistance during high fat diet-induced obesity in mice

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Gamma delta T cells are resident in adipose tissue and increase during diet-induced obesity. Their possible contribution to the inflammatory response that accompanies diet-induced obesity was investigated in mice after a 5-10 week high milk fat diet. The high milk fat diet resulted in significant in...

  5. LFA-1/ICAM-1 Ligation in Human T Cells Promotes Th1 Polarization through a GSK3β Signaling-Dependent Notch Pathway.

    PubMed

    Verma, Navin K; Fazil, M H U Turabe; Ong, Seow Theng; Chalasani, Madhavi Latha S; Low, Jian Hui; Kottaiswamy, Amuthavalli; P, Praseetha; Kizhakeyil, Atish; Kumar, Sunil; Panda, Aditya K; Freeley, Michael; Smith, Sinead M; Boehm, Bernhard O; Kelleher, Dermot

    2016-07-01

    In this study, we report that the integrin LFA-1 cross-linking with its ligand ICAM-1 in human PBMCs or CD4(+) T cells promotes Th1 polarization by upregulating IFN-γ secretion and T-bet expression. LFA-1 stimulation in PBMCs, CD4(+) T cells, or the T cell line HuT78 activates the Notch pathway by nuclear translocation of cleaved Notch1 intracellular domain (NICD) and upregulation of target molecules Hey1 and Hes1. Blocking LFA-1 by a neutralizing Ab or specific inhibition of Notch1 by a γ-secretase inhibitor substantially inhibits LFA-1/ICAM-1-mediated activation of Notch signaling. We further demonstrate that the Notch pathway activation is dependent on LFA-1/ICAM-1-induced inactivation of glycogen synthase kinase 3β (GSK3β), which is mediated via Akt and ERK. Furthermore, in silico analysis in combination with coimmunoprecipitation assays show an interaction between NICD and GSK3β. Thus, there exists a molecular cross-talk between LFA-1 and Notch1 through the Akt/ERK-GSK3β signaling axis that ultimately enhances T cell differentiation toward Th1. Although clinical use of LFA-1 antagonists is limited by toxicity related to immunosuppression, these findings support the concept that Notch inhibitors could be attractive for prevention or treatment of Th1-related immunologic disorders and have implications at the level of local inflammatory responses. PMID:27206767

  6. The Receptor Complex Associated with Human T-Cell Lymphotropic Virus Type 3 (HTLV-3) Env-Mediated Binding and Entry Is Distinct from, but Overlaps with, the Receptor Complexes of HTLV-1 and HTLV-2▿

    PubMed Central

    Jones, Kathryn S.; Huang, Ying K.; Chevalier, Sébastien A.; Afonso, Philippe V.; Petrow-Sadowski, Cari; Bertolette, Daniel C.; Gessain, Antoine; Ruscetti, Francis W.; Mahieux, Renaud

    2009-01-01

    Little is known about the transmission or tropism of the newly discovered human retrovirus, human T-cell lymphotropic virus type 3 (HTLV-3). Here, we examine the entry requirements of HTLV-3 using independently expressed Env proteins. We observed that HTLV-3 surface glycoprotein (SU) binds efficiently to both activated CD4+ and CD8+ T cells. This contrasts with both HTLV-1 SU, which primarily binds to activated CD4+ T cells, and HTLV-2 SU, which primarily binds to activated CD8+ T cells. Binding studies with heparan sulfate proteoglycans (HSPGs) and neuropilin-1 (NRP-1), two molecules important for HTLV-1 entry, revealed that these molecules also enhance HTLV-3 SU binding. However, unlike HTLV-1 SU, HTLV-3 SU can bind efficiently in the absence of both HSPGs and NRP-1. Studies of entry performed with HTLV-3 Env-pseudotyped viruses together with SU binding studies revealed that, for HTLV-1, glucose transporter 1 (GLUT-1) functions at a postbinding step during HTLV-3 Env-mediated entry. Further studies revealed that HTLV-3 SU binds efficiently to naïve CD4+ T cells, which do not bind either HTLV-1 or HTLV-2 SU and do not express detectable levels of HSPGs, NRP-1, and GLUT-1. These results indicate that the complex of receptor molecules used by HTLV-3 to bind to primary T lymphocytes differs from that of both HTLV-1 and HTLV-2. PMID:19279090

  7. The distal upstream promoter in Ly49 genes, Pro1, is active in mature NK cells and T cells, does not require TATA boxes, and displays enhancer activity1

    PubMed Central

    Gays, Frances; Taha, Sally; Brooks, Colin G.

    2016-01-01

    Missing self recognition of MHC class I molecules is mediated in murine species through the stochastic expression of CD94/NKG2 and Ly49 receptors on NK cells. Previous studies have suggested that the stochastic expression of Ly49 receptors is achieved through the use of an alternate upstream promoter, designated Pro1, that is active only in immature NK cells, and operates via the mutually exclusive binding of transcription initiation complexes to closely opposed forward and reverse TATA boxes, forward transcription being transiently required to activate the downstream promoters, Pro2/Pro3, that are subsequently responsible for transcription in mature NK cells. Here we report that Pro1 transcripts are not restricted to immature NK cells but are also found in mature NK cells and T cells, and that Pro1-fragments display strong promoter activity in mature NK cell and T cell lines as well as in immature NK cells. However, the strength of promoter activity in vitro does not correlate well with Ly49 expression in vivo and forward promoter activity is generally weak or undetectable, suggesting that components outside of Pro1 are required for efficient forward transcription. Indeed, conserved sequences immediately upstream and downstream of the core Pro1 region were found to inhibit or enhance promoter activity. Most surprisingly, promoter activity does not require either the forward or reverse TATA boxes, but is instead dependent on residues in the largely invariant central region of Pro1. Importantly, Pro1 displays strong enhancer activity suggesting that this may be its principal function in vivo. PMID:25926675

  8. Splenic Long-Lived Plasma Cells Promote the Development of Follicular Helper T Cells during Autoimmune Responses.

    PubMed

    Jang, Eunkyeong; Cho, Wang Sik; Oh, Yeon-Kyung; Cho, Mi-La; Kim, Jung Mogg; Paik, Doo-Jin; Youn, Jeehee

    2016-02-01

    Long-lived plasma cells (LLPCs) develop under the help of follicular helper T (Tfh) cells and reside mainly in the bone marrow. However, these cells are unusually abundant in the spleen of several autoimmune models including K/BxNsf mice, yet their pathogenic impact remains unknown. To investigate a previously unappreciated role of splenic LLPCs, we sorted splenic plasma cells (PCs) from K/BxNsf and K/BxN mice, corresponding to LLPCs and conventional short-lived PCs, respectively, and compared their phenotypes and ability to prime and induce the differentiation of naive CD4(+) T cells into effector cells in vitro and in vivo. We found that K/BxNsf PCs had lower levels of the Ag presentation machinery and costimulators than K/BxN PCs, and also a lower CD4(+) T cell priming capacity. Autoantigen-pulsed K/BxNsf PCs selectively polarized cognate CD4(+) T cells toward the expression of molecules necessary for Tfh development and function. As a result, the K/BxNsf PC-primed CD4(+) T cells were more effective in stimulating B cells to produce autoantigen-specific IgGs than K/BxN PCs or even dendritic cells. Adoptive transfer of K/BxNsf PCs, but not K/BxN PCs, to K/BxN mice increased numbers of Tfh cells in draining lymph nodes. These results propose that abnormal accumulation of LLPCs in the spleen of autoimmune models drives the differentiation of autoantigen-primed CD4(+) T cells to Tfh cells. This positive feedback loop between splenic LLPCs and Tfh cells may contribute to the persistence of humoral autoimmunity. PMID:26729802

  9. A Novel Copper Chelate Modulates Tumor Associated Macrophages to Promote Anti-Tumor Response of T Cells

    PubMed Central

    Chakraborty, Paramita; Ganguly, Avishek; Adhikary, Arghya; Mukhopadhyay, Debanjan; Ganguli, Sudipta; Banerjee, Rajdeep; Ashraf, Mohammad; Biswas, Jaydip; Das, Pradeep K.; Sa, Gourisankar; Chatterjee, Mitali; Das, Tanya; Choudhuri, Soumitra Kumar

    2009-01-01

    Background At the early stages of carcinogenesis, the induction of tumor specific T cell mediated immunity seems to block the tumor growth and give protective anti-tumor immune response. However, tumor associated macrophages (TAMs) might play an immunosuppressive role and subvert this anti tumor immunity leading to tumor progression and metastasis. Methodology/Principal Findings The Cu (II) complex, (chelate), copper N-(2-hydroxy acetophenone) glycinate (CuNG), synthesized by us, has previously been shown to have a potential usefulness in immunotherapy of multiple drug resistant cancers. The current study demonstrates that CuNG treatment of TAMs modulates their status from immunosuppressive to proimmunogenic nature. Interestingly, these activated TAMs produced high levels of IL-12 along with low levels of IL-10 that not only allowed strong Th1 response marked by generation of high levels of IFN-γ but also reduced activation induced T cell death. Similarly, CuNG treatment of peripheral blood monocytes from chemotherapy and/or radiotherapy refractory cancer patients also modulated their cytokine status. Most intriguingly, CuNG treated TAMs could influence reprogramming of TGF-β producing CD4+CD25+ T cells toward IFN-γ producing T cells. Conclusion/Significance Our results show the potential usefulness of CuNG in immunotherapy of drug-resistant cancers through reprogramming of TAMs that in turn reprogram the T cells and reeducate the T helper function to elicit proper anti-tumorogenic Th1 response leading to effective reduction in tumor growth. PMID:19756150

  10. Endothelial CD47 promotes Vascular Endothelial-cadherin tyrosine phosphorylation and participates in T-cell recruitment at sites of inflammation in vivo

    PubMed Central

    Azcutia, Veronica; Stefanidakis, Michael; Tsuboi, Naotake; Mayadas, Tanya; Croce, Kevin J.; Fukuda, Daiju; Aikawa, Masanori; Newton, Gail; Luscinskas, Francis W.

    2012-01-01

    At sites of inflammation, endothelial adhesion molecules bind leukocytes and transmit signals required for transendothelial migration (TEM). We previously reported that adhesive interactions between endothelial cell CD47 and leukocyte Signal Regulatory Proteinγ (SIRPγ) regulate human T-cell TEM. The role of endothelial CD47 in T-cell TEM in vivo, however, has not been explored. Here, CD47−/− mice showed reduced recruitment of blood T-cells as well as neutrophils and monocytes in a dermal air pouch model of TNF-α induced inflammation. Reconstitution of CD47−/− mice with wild type bone marrow (BM) cells did not restore leukocyte recruitment to the air pouch, indicating a role for endothelial CD47. The defect in leukocyte TEM in the CD47−/− endothelium was corroborated by intravital microscopy of inflamed cremaster muscle microcirculation in BM chimera mice. In an in vitro human system, CD47 on both HUVEC and T-cells were required for TEM. Although previous studies showed CD47-dependent signaling required Gαi coupled pathways, this was not the case for endothelial CD47 because pertussis toxin (PTX), which inactivates Gαi, had no inhibitory effect, whereas Gαi was required by the T-cell for TEM. We next investigated the endothelial CD47-dependent signaling events that accompany leukocyte TEM. Antibody-induced crosslinking of CD47 revealed robust actin cytoskeleton reorganization and Src and Pyk-2 kinase dependent tyrosine phosphorylation of the VE-cadherin cytoplasmic tail. This signaling was PTX insensitive suggesting that endothelial CD47 signaling is independent of Gαi. These findings suggest that engagement of endothelial CD47 by its ligands triggers “outside-in” signals in endothelium that facilitate leukocyte TEM. PMID:22815286

  11. The PD-1/PD-L1 complex resembles the antigen-binding Fv domains of antibodies and T cell receptors

    SciTech Connect

    Lin, David Yin-wei; Tanaka, Yoshimasa; Iwasaki, Masashi; Gittis, Apostolos G.; Su, Hua-Poo; Mikami, Bunzo; Okazaki, Taku; Honjo, Tasuku; Minato, Nagahiro; Garboczi, David N.

    2008-07-29

    Signaling through the programmed death 1 (PD-1) inhibitory receptor upon binding its ligand, PD-L1, suppresses immune responses against autoantigens and tumors and plays an important role in the maintenance of peripheral immune tolerance. Release from PD-1 inhibitory signaling revives 'exhausted' virus-specific T cells in chronic viral infections. Here we present the crystal structure of murine PD-1 in complex with human PD-L1. PD-1 and PD-L1 interact through the conserved front and side of their Ig variable (IgV) domains, as do the IgV domains of antibodies and T cell receptors. This places the loops at the ends of the IgV domains on the same side of the PD-1/PD-L1 complex, forming a surface that is similar to the antigen-binding surface of antibodies and T cell receptors. Mapping conserved residues allowed the identification of residues that are important in forming the PD-1/PD-L1 interface. Based on the structure, we show that some reported loss-of-binding mutations involve the PD-1/PD-L1 interaction but that others compromise protein folding. The PD-1/PD-L1 interaction described here may be blocked by antibodies or by designed small-molecule drugs to lower inhibitory signaling that results in a stronger immune response. The immune receptor-like loops offer a new surface for further study and potentially the design of molecules that would affect PD-1/PD-L1 complex formation and thereby modulate the immune response.

  12. A highly tilted binding mode by a self-reactive T cell receptor results in altered engagement of peptide and MHC

    SciTech Connect

    Sethi, D.K.; Heroux, A.; Schubert, D. A.; Anders, A.-K.; Bonsor, D. A.; Thomas, C. P.; Sundberg, E. J.; Pyrdol, J.; Wucherpfennig, K. W.

    2011-01-17

    Self-reactive T cells that escape elimination in the thymus can cause autoimmune pathology, and it is therefore important to understand the structural mechanisms of self-antigen recognition. We report the crystal structure of a T cell receptor (TCR) from a patient with relapsing-remitting multiple sclerosis that engages its self-peptide-major histocompatibility complex (pMHC) ligand in an unusual manner. The TCR is bound in a highly tilted orientation that prevents interaction of the TCR-{alpha} chain with the MHC class II {beta} chain helix. In this structure, only a single germline-encoded TCR loop engages the MHC protein, whereas in most other TCR-pMHC structures all four germline-encoded TCR loops bind to the MHC helices. The tilted binding mode also prevents peptide contacts by the short complementarity-determining region (CDR) 3{beta} loop, and interactions that contribute to peptide side chain specificity are focused on the CDR3{alpha} loop. This structure is the first example in which only a single germline-encoded TCR loop contacts the MHC helices. Furthermore, the reduced interaction surface with the peptide may facilitate TCR cross-reactivity. The structural alterations in the trimolecular complex are distinct from previously characterized self-reactive TCRs, indicating that there are multiple unusual ways for self-reactive TCRs to bind their pMHC ligand.

  13. A Highly Tilted Binding Mode by a Self-Reactive T Cell Receptor Results in Altered Engagement of Peptide and MHC

    SciTech Connect

    D Sethi; D Schubert; A Anders; A Heroux; D Bonsor; C Thomas; E Sundberg; J Pyrdol; K Wucherpfennig

    2011-12-31

    Self-reactive T cells that escape elimination in the thymus can cause autoimmune pathology, and it is therefore important to understand the structural mechanisms of self-antigen recognition. We report the crystal structure of a T cell receptor (TCR) from a patient with relapsing-remitting multiple sclerosis that engages its self-peptide-major histocompatibility complex (pMHC) ligand in an unusual manner. The TCR is bound in a highly tilted orientation that prevents interaction of the TCR-{alpha} chain with the MHC class II {beta} chain helix. In this structure, only a single germline-encoded TCR loop engages the MHC protein, whereas in most other TCR-pMHC structures all four germline-encoded TCR loops bind to the MHC helices. The tilted binding mode also prevents peptide contacts by the short complementarity-determining region (CDR) 3{beta} loop, and interactions that contribute to peptide side chain specificity are focused on the CDR3{alpha} loop. This structure is the first example in which only a single germline-encoded TCR loop contacts the MHC helices. Furthermore, the reduced interaction surface with the peptide may facilitate TCR cross-reactivity. The structural alterations in the trimolecular complex are distinct from previously characterized self-reactive TCRs, indicating that there are multiple unusual ways for self-reactive TCRs to bind their pMHC ligand.

  14. Strategic mutations in the class I major histocompatibility complex HLA-A2 independently affect both peptide binding and T cell receptor recognition.

    PubMed

    Baxter, Tiffany K; Gagnon, Susan J; Davis-Harrison, Rebecca L; Beck, John C; Binz, Anne-Kathrin; Turner, Richard V; Biddison, William E; Baker, Brian M

    2004-07-01

    Mutational studies of T cell receptor (TCR) contact residues on the surface of the human class I major histocompatibility complex (MHC) molecule HLA-A2 have identified a "functional hot spot" that comprises Arg(65) and Lys(66) and is involved in recognition by most peptide-specific HLA-A2-restricted TCRs. Although there is a significant amount of functional data on the effects of mutations at these positions, there is comparatively little biochemical information that could illuminate their mode of action. Here, we have used a combination of fluorescence anisotropy, functional assays, and Biacore binding experiments to examine the effects of mutations at these positions on the peptide-MHC interaction and TCR recognition. The results indicate that mutations at both position 65 and position 66 influence peptide binding by HLA-A2 to various extents. In particular, mutations at position 66 result in significantly increased peptide dissociation rates. However, these effects are independent of their effects on TCR recognition, and the Arg(65)-Lys(66) region thus represents a true "hot spot" for TCR recognition. We also made the observation that in vitro T cell reactivity does not scale with the half-life of the peptide-MHC complex, as is often assumed. Finally, position 66 is implicated in the "dual recognition" of both peptide and TCR, emphasizing the multiple roles of the class I MHC peptide-binding domain. PMID:15131131

  15. Developmental exposure to trichloroethylene promotes CD4{sup +} T cell differentiation and hyperactivity in association with oxidative stress and neurobehavioral deficits in MRL+/+ mice

    SciTech Connect

    Blossom, Sarah J. Doss, Jason C.; Hennings, Leah J.; Jernigan, Stefanie; Melnyk, Stepan; James, S. Jill

    2008-09-15

    The non adult immune system is particularly sensitive to perinatal and early life exposures to environmental toxicants. The common environmental toxicant, trichloroethylene (TCE), was shown to increase CD4+ T cell production of the proinflammatory cytokine IFN-{gamma} following a period of prenatal and lifetime exposure in autoimmune-prone MRL+/+ mice. In the current study, MRL+/+ mice were used to further examine the impact of TCE on the immune system in the thymus and periphery. Since there is considerable cross-talk between the immune system and the brain during development, the potential relationship between TCE and neurobehavioral endpoints were also examined. MRL+/+ mice were exposed to 0.1 mg/ml TCE ({approx} 31 mg/kg/day) via maternal drinking water or direct exposure via the drinking water from gestation day 1 until postnatal day (PD) 42. TCE exposure did not impact gross motor skills but instead significantly altered social behaviors and promoted aggression associated with indicators of oxidative stress in brain tissues in male mice. The immunoregulatory effects of TCE involved a redox-associated promotion of T cell differentiation in the thymus that preceded the production of proinflammatory cytokines, IL-2, TNF-{alpha}, and IFN-{gamma} by mature CD4+ T cells. The results demonstrated that developmental and early life TCE exposure modulated immune function and may have important implications for neurodevelopmental disorders.

  16. Induced miR-99a expression represses Mtor cooperatively with miR-150 to promote regulatory T-cell differentiation

    PubMed Central

    Warth, Sebastian C; Hoefig, Kai P; Hiekel, Anian; Schallenberg, Sonja; Jovanovic, Ksenija; Klein, Ludger; Kretschmer, Karsten; Ansel, K Mark; Heissmeyer, Vigo

    2015-01-01

    Peripheral induction of regulatory T (Treg) cells provides essential protection from inappropriate immune responses. CD4+ T cells that lack endogenous miRNAs are impaired to differentiate into Treg cells, but the relevant miRNAs are unknown. We performed an overexpression screen with T-cell-expressed miRNAs in naive mouse CD4+ T cells undergoing Treg differentiation. Among 130 candidates, the screen identified 29 miRNAs with a negative and 10 miRNAs with a positive effect. Testing reciprocal Th17 differentiation revealed specific functions for miR-100, miR-99a and miR-10b, since all of these promoted the Treg and inhibited the Th17 program without impacting on viability, proliferation and activation. miR-99a cooperated with miR-150 to repress the expression of the Th17-promoting factor mTOR. The comparably low expression of miR-99a was strongly increased by the Treg cell inducer “retinoic acid”, and the abundantly expressed miR-150 could only repress Mtor in the presence of miR-99a. Our data suggest that induction of Treg cell differentiation is regulated by a miRNA network, which involves cooperation of constitutively expressed as well as inducible miRNAs. PMID:25712478

  17. Cutting Edge: NFAT Transcription Factors Promote the Generation of Follicular Helper T Cells in Response to Acute Viral Infection.

    PubMed

    Martinez, Gustavo J; Hu, Joyce K; Pereira, Renata M; Crampton, Jordan S; Togher, Susan; Bild, Nicholas; Crotty, Shane; Rao, Anjana

    2016-03-01

    Follicular CD4(+) Th (Tfh) cells provide B cell help in germinal center reactions that support class switching, somatic hypermutation, and the generation of high-affinity Abs. In this article, we show that deficiency in NFAT1 and NFAT2 in CD4(+) T cells leads to impaired germinal center reactions upon viral infection because of reduced Tfh cell differentiation and defective expression of proteins involved in T/B interactions and B cell help, including ICOS, PD-1, and SLAM family receptors. Genome-wide chromatin immunoprecipitation data suggest that NFAT proteins likely directly participate in regulation of genes important for Tfh cell differentiation and function. NFAT proteins are important TCR and Ca(2+)-dependent regulators of T cell biology, and in this article we demonstrate a major positive role of NFAT family members in Tfh differentiation. PMID:26851216

  18. IL4RA on lymphatic endothelial cells promotes T cell egress during sclerodermatous graft versus host disease

    PubMed Central

    Urso, Katia; Alvarez, David; Cremasco, Viviana; Tsang, Kelly; Grauel, Angelo; Lafyatis, Robert; von Andrian, Ulrich H.; Ermann, Joerg; Aliprantis, Antonios O.

    2016-01-01

    Systemic sclerosis (SSc) is a potentially fatal autoimmune disorder with limited therapeutic options. Sclerodermatous graft versus host disease (sclGvHD), induced by transfer of B10.D2 splenocytes into BALB/c Rag2−/− mice, models an inflammatory subset of SSc characterized by a prominent IL13-induced gene expression signature in the skin. Host mice deficient in IL4RA, a subunit of the type II IL4/IL13 receptor, are protected from sclGvHD. While IL4RA has a well-established role in Th2 differentiation and alternative macrophage activation, we report here a previously unappreciated function for IL4RA in lymphatic endothelial cells (LECs): regulation of activated T cell egress. Seven days after splenocyte transfer, Il4ra−/− hosts had increased numbers of activated graft CD4+ T cells in skin draining lymph nodes (dLNs) but fewer T cells in efferent lymph, blood, and skin. Sphingosine-1 phosphate (S1P), master regulator of lymphocyte egress from LNs, was lower in dLNs of Il4ra−/− hosts with a corresponding decrease of S1P kinase 1 (Sphk1) expression in LECs. Bypassing the efferent lymphatics via i.v. injection of CD4+ T cells from dLNs of Il4ra−/− sclGvHD mice restored clinical GvHD in secondary Il4ra−/− recipients. These results identify a role for IL4RA and suggest that modulation of lymphocyte egress from LNs may be effective in SSc and GvHD. PMID:27547823

  19. Progesterone suppresses the mTOR pathway and promotes generation of induced regulatory T cells with increased stability

    PubMed Central

    Lee, Jee H.; Lydon, John P.; Kim, Chang H.

    2013-01-01

    Summary While induced FoxP3+ T cells (iTregs) are promising cellular therapeutics to treat inflammatory diseases, a limitation in utilizing iTregs prepared in vitro is their low stability in inflammatory conditions. Progesterone (P4) is an immune regulatory nuclear hormone with a potent Treg induction activity. We reasoned that this function of progesterone would be utilized to generate iTregs with highly suppressive activity and improved stability in vivo. We generated iTregs with progesterone in vitro and found that progesterone generates iTregs that are highly stable in inflammatory conditions. Moreover, P4-induced iTregs highly express latency-associated peptide TGFβ1 and are efficient in regulating inflammation in multiple tissues, whereas control iTregs induced with TGFβ1 alone are less stable and ineffective in suppressing inflammation. The function of progesterone in inducing iTregs with improved regulatory activity is associated with the function of P4 in suppressing the mTOR pathway. Moreover, the function of progesterone in inducing FoxP3+ T cells is decreased but not completely abolished on nuclear progesterone receptor-deficient T cells, suggesting that both nuclear and non-nuclear progesterone receptors are involved in mediating the function. We conclude that P4 can be utilized to generate iTregs with a high therapeutic potential in treatment of tissue inflammation. PMID:22740122

  20. 4.1R proteins associate with interphase microtubules in human T cells: a 4.1R constitutive region is involved in tubulin binding.

    PubMed

    Pérez-Ferreiro, C M; Luque, C M; Correas, I

    2001-11-30

    Red blood cell protein 4.1 (4.1R) is an 80-kDa protein that stabilizes the spectrin-actin network and anchors it to the plasma membrane. To contribute to the characterization of functional roles and partners of specific nonerythroid 4.1R isoforms, we analyzed 4.1R in human T cells and found that endogenous 4.1R was distributed to the microtubule network. Transfection experiments of T cell 4.1R cDNAs in conjunction with confocal microscopy analysis revealed the colocalization of exogenous 4.1R isoforms with the tubulin skeleton. Biochemical analyses using Taxol (paclitaxel)-polymerized microtubules from stably transfected T cells confirmed the association of the exogenous 4.1R proteins with microtubules. Consistent with this, endogenous 4.1R immunoreactive proteins were also detected in the microtubule-containing fraction. In vitro binding assays using glutathione S-transferase-4.1R fusion proteins showed that a constitutive domain of the 4.1R molecule, one that is therefore present in all 4.1R isoforms, is responsible for the association with tubulin. A 22-amino acid sequence comprised in this domain and containing heptad repeats of leucine residues was essential for tubulin binding. Furthermore, ectopic expression of 4.1R in COS-7 cells provoked microtubule disorganization. Our results suggest an involvement of 4.1R in interphase microtubule architecture and support the hypothesis that some 4.1R functional activities are cell type-regulated. PMID:11579097

  1. Dynamic regulation of permissive histone modifications and GATA3 binding underpin acquisition of granzyme A expression by virus-specific CD8(+) T cells.

    PubMed

    Nguyen, Michelle L T; Hatton, Lauren; Li, Jasmine; Olshansky, Moshe; Kelso, Anne; Russ, Brendan E; Turner, Stephen J

    2016-02-01

    Numerous studies have focused on the molecular regulation of perforin (PFP) and granzyme B (GZMB) expression by activated cytotoxic T lymphocytes (CTLs), but little is known about the molecular factors that underpin granzyme A (GZMA) expression. In vitro activation of naïve CD8(+) T cells, in the presence of IL-4, enhanced STAT6-dependent GZMA expression and was associated with GATA3 binding and enrichment of transcriptionally permissive histone posttranslational modifications (PTMs) across the Gzma gene locus. While GZMA expression by effector influenza A virus specific CTLs was also associated with a similar permissive epigenetic signature, memory CTL lacked enrichment of permissive histone PTMs at the Gzma locus, although this was restored within recalled secondary effector CTLs. Importantly, GZMA expression by virus-specific CTLs was associated with GATA3 binding at the Gzma locus, and independent of STAT6-mediated signaling. This suggests regulation of GZMA expression is underpinned by differentiation-dependent regulation of chromatin composition at the Gzma locus and that, given GATA3 is key for CTL differentiation in response to infection, GATA3 expression is regulated by a distinct, IL-4 independent, signaling pathway. Overall, this study provides insights into the molecular mechanisms that control transcription of Gzma during virus-induced CD8(+) T-cell differentiation. PMID:26519105

  2. B7-H3 Promotes Pathogenesis of Autoimmune Disease and Inflammation by Regulating the Activity of Different T Cell Subsets.

    PubMed

    Luo, Liqun; Zhu, Gefeng; Xu, Haiying; Yao, Sheng; Zhou, Gang; Zhu, Yuwen; Tamada, Koji; Huang, Lanqing; Flies, Andrew D; Broadwater, Megan; Ruff, William; van Deursen, Jan M A; Melero, Ignacio; Zhu, Zhou; Chen, Lieping

    2015-01-01

    B7-H3 is a cell surface molecule in the immunoglobulin superfamily that is frequently upregulated in response to autoantigens and pathogens during host T cell immune responses. However, B7-H3's role in the differential regulation of T cell subsets remains largely unknown. Therefore, we constructed a new B7-H3 deficient mouse strain (B7-H3 KO) and evaluated the functions of B7-H3 in the regulation of Th1, Th2, and Th17 subsets in experimental autoimmune encephalomyelitis (EAE), experimental asthma, and collagen-induced arthritis (CIA); these mouse models were used to predict human immune responses in multiple sclerosis, asthma, and rheumatoid arthritis, respectively. Here, we demonstrate that B7-H3 KO mice have significantly less inflammation, decreased pathogenesis, and limited disease progression in both EAE and CIA mouse models when compared with littermates; these results were accompanied by a decrease in IFN-γ and IL-17 production. In sharp contrast, B7-H3 KO mice developed severe ovalbumin (OVA)-induced asthma with characteristic infiltrations of eosinophils in the lung, increased IL-5 and IL-13 in lavage fluid, and elevated IgE anti-OVA antibodies in the blood. Our results suggest B7-H3 has a costimulatory function on Th1/Th17 but a coinhibitory function on Th2 responses. Our studies reveal that B7-H3 could affect different T cell subsets which have important implications for regulating pathogenesis and disease progression in human autoimmune disease. PMID:26065426

  3. Deletion of 5-Lipoxygenase in the Tumor Microenvironment Promotes Lung Cancer Progression and Metastasis through Regulating T Cell Recruitment.

    PubMed

    Poczobutt, Joanna M; Nguyen, Teresa T; Hanson, Dwight; Li, Howard; Sippel, Trisha R; Weiser-Evans, Mary C M; Gijon, Miguel; Murphy, Robert C; Nemenoff, Raphael A

    2016-01-15

    Eicosanoids, including PGs, produced by cyclooxygenases (COX), and leukotrienes, produced by 5-lipoxygenase (5-LO) have been implicated in cancer progression. These molecules are produced by both cancer cells and the tumor microenvironment (TME). We previously reported that both COX and 5-LO metabolites increase during progression in an orthotopic immunocompetent model of lung cancer. Although PGs in the TME have been well studied, less is known regarding 5-LO products produced by the TME. We examined the role of 5-LO in the TME using a model in which Lewis lung carcinoma cells are directly implanted into the lungs of syngeneic WT mice or mice globally deficient in 5-LO (5-LO-KO). Unexpectedly, primary tumor volume and liver metastases were increased in 5-LO-KO mice. This was associated with an ablation of leukotriene (LT) production, consistent with production mainly mediated by the microenvironment. Increased tumor progression was partially reproduced in global LTC4 synthase KO or mice transplanted with LTA4 hydrolase-deficient bone marrow. Tumor-bearing lungs of 5-LO-KO had decreased numbers of CD4 and CD8 T cells compared with WT controls, as well as fewer dendritic cells. This was associated with lower levels of CCL20 and CXL9, which have been implicated in dendritic and T cell recruitment. Depletion of CD8 cells increased tumor growth and eliminated the differences between WT and 5-LO mice. These data reveal an antitumorigenic role for 5-LO products in the microenvironment during lung cancer progression through regulation of T cells and suggest that caution should be used in targeting this pathway in lung cancer. PMID:26663781

  4. Deletion of 5-Lipoxygenase in the Tumor Microenvironment Promotes Lung Cancer Progression and Metastasis through Regulating T Cell Recruitment

    PubMed Central

    Poczobutt, Joanna M.; Nguyen, Teresa T.; Hanson, Dwight; Li, Howard; Sippel, Trisha R.; Weiser-Evans, Mary C. M.; Gijon, Miguel; Murphy, Robert C.

    2016-01-01

    Eicosanoids, including PGs, produced by cyclooxygenases (COX), and leukotrienes, produced by 5-lipoxygenase (5-LO) have been implicated in cancer progression. These molecules are produced by both cancer cells and the tumor microenvironment (TME). We previously reported that both COX and 5-LO metabolites increase during progression in an orthotopic immunocompetent model of lung cancer. Although PGs in the TME have been well studied, less is known regarding 5-LO products produced by the TME. We examined the role of 5-LO in the TME using a model in which Lewis lung carcinoma cells are directly implanted into the lungs of syngeneic WT mice or mice globally deficient in 5-LO (5-LO-KO). Unexpectedly, primary tumor volume and liver metastases were increased in 5-LO-KO mice. This was associated with an ablation of leukotriene (LT) production, consistent with production mainly mediated by the microenvironment. Increased tumor progression was partially reproduced in global LTC4 synthase KO or mice transplanted with LTA4 hydrolase-deficient bone marrow. Tumor-bearing lungs of 5-LO-KO had decreased numbers of CD4 and CD8 T cells compared with WT controls, as well as fewer dendritic cells. This was associated with lower levels of CCL20 and CXL9, which have been implicated in dendritic and T cell recruitment. Depletion of CD8 cells increased tumor growth and eliminated the differences between WT and 5-LO mice. These data reveal an antitumorigenic role for 5-LO products in the microenvironment during lung cancer progression through regulation of T cells and suggest that caution should be used in targeting this pathway in lung cancer. PMID:26663781

  5. Loss of CAR promotes migration and proliferation of HaCaT cells, and accelerates wound healing in rats via Src-p38 MAPK pathway

    PubMed Central

    Su, Linlin; Fu, Lanqing; Li, Xiaodong; Zhang, Yue; Li, Zhenzhen; Wu, Xue; Li, Yan; Bai, Xiaozhi; Hu, Dahai

    2016-01-01

    The coxsackie and adenovirus receptor (CAR) is a cell adhesion molecule mostly localized to cell-cell contacts in epithelial and endothelial cells. CAR is known to regulate tumor progression, however, its physiological role in keratinocyte migration and proliferation, two essential steps in re-epithelialization during wound healing, has less been investigated. Here we showed that CAR was predominantly expressed in the epidermis of human skin, CAR knockdown by RNAi significantly accelerated HaCaT cell migration and proliferation. In addition, knockdown of CAR in vitro increased p-Src, p-p38, and p-JNK protein levels; however, Src inhibitor PP2 prevented the increase of p-Src and p-p38 induced by CAR RNAi, but not p-JNK, and decelerated cell migration and proliferation. More intriguingly, in vivo CAR RNAi on the skin area surrounding the wounds on rat back visually accelerated wound healing and re-epithelialization process, while treatment with PP2 or p38 inhibitor SB203580 obviously inhibited these effects. By contrast, overexpressing CAR in HaCaT cells significantly decelerated cell migration and proliferation. Above results demonstrate that suppression of CAR could accelerate HaCaT cell migration and proliferation, and promote wound healing in rat skin, probably via Src-p38 MAPK pathway. CAR thus might serve as a novel therapeutic target for facilitating wound healing. PMID:26804208

  6. Genetic deficiency of Irgm1 (LRG-47) suppresses induction of experimental autoimmune encephalomyelitis by promoting apoptosis of activated CD4+ T cells

    PubMed Central

    Xu, Hongwei; Wu, Zhi-Ying; Fang, Fang; Guo, Lan; Chen, Doris; Chen, John Xi; Stern, David; Taylor, Gregory A.; Jiang, Hong; Yan, Shirley ShiDu

    2010-01-01

    The immunity-related GTPase Irgm1, also called LRG-47, is known to regulate host resistance to intracellular pathogens through multiple mechanisms that include controlling the survival of T lymphocytes. Here, we address whether Irgm1 also plays a role in the pathogenesis of experimental autoimmune encephalitis (EAE). We find that Irgm1/LRG-47 is a significant factor in the progression of EAE and multiple sclerosis (MS). Expression of Irgm1 was robustly elevated in MS-affected lesions and in the central nervous system (CNS) of myelin basic protein (MBP)-induced EAE mice, especially in cells of lymphoid and mononuclear phagocyte origin. Homozygous Irgm1 null mice were resistant to MBP-induced EAE, and CD4+ T cells in spleen and CNS of these mice displayed decreased proliferative capacity, increased apoptosis, and up-regulated interferon (IFN)-γ induction. Therefore, Irgm1-induced survival of autoreactive CD4+ T cells contributes significantly to the pathogenesis of EAE. Blockade of Irgm1 may be a potential therapeutic strategy for halting multiple sclerosis.—Xu, H., Wu, Z.-Y., Fang, F., Guo, L., Chen, D., Chen, J. X., Stern, D., Taylor, G. A., Jiang, H., Yan, S. S. Genetic deficiency of Irgm1 (LRG-47) suppresses induction of experimental autoimmune encephalomyelitis by promoting apoptosis of activated CD4+ T cells. PMID:20056715

  7. Regulatory T cells promote a protective Th17-associated immune response to intestinal bacterial infection with C. rodentium.

    PubMed

    Wang, Z; Friedrich, C; Hagemann, S C; Korte, W H; Goharani, N; Cording, S; Eberl, G; Sparwasser, T; Lochner, M

    2014-11-01

    Intestinal infection with the mouse pathogen Citrobacter rodentium induces a strong local Th17 response in the colon. Although this inflammatory immune response helps to clear the pathogen, it also induces inflammation-associated pathology in the gut and thus, has to be tightly controlled. In this project, we therefore studied the impact of Foxp3(+) regulatory T cells (Treg) on the infectious and inflammatory processes elicited by the bacterial pathogen C. rodentium. Surprisingly, we found that depletion of Treg by diphtheria toxin in the Foxp3(DTR) (DEREG) mouse model resulted in impaired bacterial clearance in the colon, exacerbated body weight loss, and increased systemic dissemination of bacteria. Consistent with the enhanced susceptibility to infection, we found that the colonic Th17-associated T-cell response was impaired in Treg-depleted mice, suggesting that the presence of Treg is crucial for the establishment of a functional Th17 response after the infection in the gut. As a consequence of the impaired Th17 response, we also observed less inflammation-associated pathology in the colons of Treg-depleted mice. Interestingly, anti-interleukin (IL)-2 treatment of infected Treg-depleted mice restored Th17 induction, indicating that Treg support the induction of a protective Th17 response during intestinal bacterial infection by consumption of local IL-2. PMID:24646939

  8. Interleukin-10 deficiency impairs regulatory T cell-derived neuropilin-1 functions and promotes Th1 and Th17 immunity

    PubMed Central

    Wang, Shimin; Gao, Xiang; Shen, Guobo; Wang, Wei; Li, Jingyu; Zhao, Jingyi; Wei, Yu-Quan; Edwards, Carl K.

    2016-01-01

    Regulatory T cells (Tregs) expand in peripheral lymphoid organs and can produce immunosuppressive cytokines to support tumor growth. IL-10 abrogation efficiently induces Treg formation but dampens tumoral neuropilin-1 (Nrp-1) Treg signaling, which simultaneously augments Th1 and Th17 immunity. These effects are associated with the plasticity and stability of Tregs and effector T cell functions that can limit tumorigenesis. Within the tumor microenvironment, there appears to be a “mutual antagonism” between immunoenhancement and immunosuppression mechanisms, eventually leading to decreased metastasis. In contrast, tumor progression is paralleled by a reduction in Nrp-1-producing Tregs controlled by the IL-10 and TGF-β1 levels. However, Th1, Th17 and Treg immunity is primarily regulated by IL-10 or Nrp-1 and not TGF-β1 except when combined with IL-10. These results emphasize the important implications for the therapeutic use of Tregs. The number of Treg cells must be maintained in a healthy and dynamic homeostatic range to prevent malignant diseases. Moreover, Treg-mediated immunosuppression can be limited by reducing tumor-derived Treg Nrp-1 levels. PMID:27075020

  9. Tadalafil Reduces Myeloid-Derived Suppressor Cells and Regulatory T Cells and Promotes Tumor Immunity in Patients with Head and Neck Squamous Cell Carcinoma

    PubMed Central

    Vella, Jennifer L.; Reis, Isildinha M.; De la fuente, Adriana C.; Gomez, Carmen; Sargi, Zoukaa; Nazarian, Ronen; Califano, Joseph; Borrello, Ivan

    2015-01-01

    Purpose Myeloid-derived suppressor cells (MDSC) and regulatory T cells (Treg) play a key role in the progression of head and neck squamous cell carcinoma (HNSCC). On the basis of our preclinical data demonstrating that phosphodiesterase-5 (PDE5) inhibition can modulate these cell populations, we evaluated whether the PDE5 inhibitor tadalafil can revert tumor-induced immunosuppression and promote tumor immunity in patients with HNSCC. Experimental Design First, we functionally and phenotypically characterized MDSCs in HNSCCs and determined, retrospectively, whether their presence at the tumor site correlates with recurrence. Then, we performed a prospective single-center, double-blinded, randomized, three-arm study in which patients with HNSCC undergoing definitive surgical resection of oral and oropharyngeal tumors were treated with tadalafil 10 μg/day, 20 μg/day, or placebo for at least 20 days preoperatively. Blood and tumor MDSC and Treg presence and CD8+ T-cell reactivity to tumor antigens were evaluated before and after treatment. Results MDSCs were characterized in HNSCC and their intratumoral presence significantly correlates with recurrence. Tadalafil treatment was well tolerated and significantly reduced both MDSCs and Treg concentrations in the blood and in the tumor (P < 0.05). In addition, the concentration of blood CD8+ T cells reactive to autologous tumor antigens significantly increased after treatment (P < 0.05). Tadalafil immunomodulatory activity was maximized at an intermediate dose but not at higher doses. Mechanistic analysis suggests a possible off-target effect on PDE11 at high dosages that, by increasing intracellular cAMP, may negatively affect antitumor immunity. Conclusions Tadalafil seems to beneficially modulate the tumor micro- and macro-environment in patients with HNSCC by lowering MDSCs and Tregs and increasing tumor-specific CD8+ T cells in a dose-dependent fashion. PMID:25320361

  10. CD40 promotes MHC class II expression on adipose tissue macrophages and regulates adipose tissue CD4+ T cells with obesity.

    PubMed

    Morris, David L; Oatmen, Kelsie E; Mergian, Taleen A; Cho, Kae Won; DelProposto, Jennifer L; Singer, Kanakadurga; Evans-Molina, Carmella; O'Rourke, Robert W; Lumeng, Carey N

    2016-06-01

    Obesity activates both innate and adaptive immune responses in adipose tissue, but the mechanisms critical for regulating these responses remain unknown. CD40/CD40L signaling provides bidirectional costimulatory signals between antigen-presenting cells and CD4(+) T cells, and CD40L expression is increased in obese humans. Therefore, we examined the contribution of CD40 to the progression of obesity-induced inflammation in mice. CD40 was highly expressed on adipose tissue macrophages in mice, and CD40/CD40L signaling promoted the expression of antigen-presenting cell markers in adipose tissue macrophages. When fed a high fat diet, Cd40-deficient mice had reduced accumulation of conventional CD4(+) T cells (Tconv: CD3(+)CD4(+)Foxp3(-)) in visceral fat compared with wild-type mice. By contrast, the number of regulatory CD4(+) T cells (Treg: CD3(+)CD4(+)Foxp3(+)) in lean and obese fat was similar between wild-type and knockout mice. Adipose tissue macrophage content and inflammatory gene expression in fat did not differ between obese wild-type and knockout mice; however, major histocompatibility complex class II and CD86 expression on adipose tissue macrophages was reduced in visceral fat from knockout mice. Similar results were observed in chimeric mice with hematopoietic Cd40-deficiency. Nonetheless, neither whole body nor hematopoietic disruption of CD40 ameliorated obesity-induced insulin resistance in mice. In human adipose tissue, CD40 expression was positively correlated with CD80 and CD86 expression in obese patients with type 2 diabetes. These findings indicate that CD40 signaling in adipose tissue macrophages regulates major histocompatibility complex class II and CD86 expression to control the expansion of CD4(+) T cells; however, this is largely dispensable for the development of obesity-induced inflammation and insulin resistance in mice. PMID:26658005

  11. Splenectomy promotes indirect elimination of intraocular tumors by CD8+ T cells that is associated with IFNγ- and Fas/FasL-dependent activation of intratumoral macrophages.

    PubMed

    Miller, Maxine R; Mandell, Jonathan B; Beatty, Kelly M; Harvey, Stephen A K; Rizzo, Michael J; Previte, Dana M; Thorne, Stephen H; McKenna, Kyle C

    2014-12-01

    Ocular immune privilege (IP) limits the immune surveillance of intraocular tumors as certain immunogenic tumor cell lines (P815, E.G7-OVA) that are rejected when transplanted in the skin grow progressively when placed in the anterior chamber of the eye. As splenectomy (SPLNX) is known to terminate ocular IP, we characterized the immune mechanisms responsible for rejection of intraocular tumors in SPLNX mice as a first step toward identifying how to restore tumoricidal activity within the eye. CD8(+) T cells, IFNγ, and FasL, but not perforin, or TNFα were required for the elimination of intraocular E.G7-OVA tumors that culminated in destruction of the eye (ocular phthisis). IFNγ and FasL did not target tumor cells directly as the majority of SPLNX IFNγR1(-/-) mice and Fas-defective lpr mice failed to eliminate intraocular E.G7-OVA tumors that expressed Fas and IFNγR1. Bone marrow chimeras revealed that IFNγR1 and Fas expression on immune cells was most critical for rejection, and SPLNX increased the frequency of activated macrophages (Mϕ) within intraocular tumors in an IFNγ- and Fas/FasL-dependent manner, suggesting an immune cell target of IFNγ and Fas. As depletion of Mϕs limited CD8 T cell-mediated rejection of intraocular tumors in SPLNX mice, our data support a model in which IFNγ- and Fas/FasL-dependent activation of intratumoral Mϕs by CD8(+) T cells promotes severe intraocular inflammation that indirectly eliminates intraocular tumors by inducing phthisis, and suggests that immunosuppressive mechanisms that maintain ocular IP interfere with the interaction between CD8(+) T cells and Mϕs to limit the immunosurveillance of intraocular tumors. PMID:25248763

  12. Bicistronic DNA Vaccines Simultaneously Encoding HIV, HSV and HPV Antigens Promote CD8+ T Cell Responses and Protective Immunity

    PubMed Central

    Santana, Vinicius C.; Diniz, Mariana O.; Cariri, Francisco A. M. O.; Ventura, Armando M.; Cunha-Neto, Edécio; Almeida, Rafael R.; Campos, Marco A.; Lima, Graciela K.; Ferreira, Luís C. S.

    2013-01-01

    Millions of people worldwide are currently infected with human papillomavirus (HPV), herpes simplex virus (HSV) or human immunodeficiency virus (HIV). For this enormous contingent of people, the search for preventive and therapeutic immunological approaches represents a hope for the eradication of latent infection and/or virus-associated cancer. To date, attempts to develop vaccines against these viruses have been mainly based on a monovalent concept, in which one or more antigens of a virus are incorporated into a vaccine formulation. In the present report, we designed and tested an immunization strategy based on DNA vaccines that simultaneously encode antigens for HIV, HSV and HPV. With this purpose in mind, we tested two bicistronic DNA vaccines (pIRES I and pIRES II) that encode the HPV-16 oncoprotein E7 and the HIV protein p24 both genetically fused to the HSV-1 gD envelope protein. Mice i.m. immunized with the DNA vaccines mounted antigen-specific CD8+ T cell responses, including in vivo cytotoxic responses, against the three antigens. Under experimental conditions, the vaccines conferred protective immunity against challenges with a vaccinia virus expressing the HIV-derived protein Gag, an HSV-1 virus strain and implantation of tumor cells expressing the HPV-16 oncoproteins. Altogether, our results show that the concept of a trivalent HIV, HSV, and HPV vaccine capable to induce CD8+ T cell-dependent responses is feasible and may aid in the development of preventive and/or therapeutic approaches for the control of diseases associated with these viruses. PMID:23951135

  13. Human Cytomegalovirus variant peptides adapt by decreasing their total coordination upon binding to a T cell receptor

    PubMed Central

    Antipas, Georgios S.E.; Germenis, Anastasios E.

    2015-01-01

    The tertiary structure of the native Cytomegalovirus peptide (NLV) presented by HLA-A2 and bound to the RA14 T cell receptor was used as a reference for the calculation of atomic coordination differences of both the NLV as well as of a number of singly substituted NLV variants in the absence of TCR. Among the pMHC complexes, the native peptide was found to exhibit the highest total coordination difference in respect to the reference structure, suggesting that it experienced the widest structural adaptation upon recognition by the TCR. In addition, the peptide on the isolated NLV-MHC complex was over-coordinated as compared to the rest of the variants. Moreover, the trend was found to account for a set of measured dissociation constants and critical concentrations for target-cell lysis for all variants in complexation with RA14: functionally, all variant peptides were established to be either weak agonists or null peptides, while, at the same time, our current study established that they were also under-coordinated in respect to NLV. It could, thus, be argued that the most ‘efficient’ structural adaptation upon pMHC recognition by the TCR requires of the peptide to undergo the widest under-coordination possible. The main structural characteristic which differentiated the NLV in respect to the variants was a the presence of 16 oxygen atoms (waters) in the former׳s second coordination shell which accounted for over-coordination of roughly 100% and 30% in the O–O and C–O partials respectively. In fact, in the absence of second shell oxygens, the NLV peptide was decidedly under-coordinated in respect to all of the variants, as also suggested by the C–C partial. PMID:26958591

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

    PubMed

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

    2016-03-01

    STK11/LKB1 is among the most commonly inactivated tumor suppressors in non-small cell lung cancer (NSCLC), especially in tumors harboring KRAS mutations. Many oncogenes promote immune escape, undermining the effectiveness of immunotherapies, but it is unclear whether the inactivation of tumor suppressor genes, such as STK11/LKB1, exerts similar effects. In this study, we investigated the consequences of STK11/LKB1 loss on the immune microenvironment in a mouse model of KRAS-driven NSCLC. Genetic ablation of STK11/LKB1 resulted in accumulation of neutrophils with T-cell-suppressive effects, along with a corresponding increase in the expression of T-cell exhaustion markers and tumor-promoting cytokines. The number of tumor-infiltrating lymphocytes was also reduced in LKB1-deficient mouse and human tumors. Furthermore, STK11/LKB1-inactivating mutations were associated with reduced expression of PD-1 ligand PD-L1 in mouse and patient tumors as well as in tumor-derived cell lines. Consistent with these results, PD-1-targeting antibodies were ineffective against Lkb1-deficient tumors. In contrast, treating Lkb1-deficient mice with an IL6-neutralizing antibody or a neutrophil-depleting antibody yielded therapeutic benefits associated with reduced neutrophil accumulation and proinflammatory cytokine expression. Our findings illustrate how tumor suppressor mutations can modulate the immune milieu of the tumor microenvironment, and they offer specific implications for addressing STK11/LKB1-mutated tumors with PD-1-targeting antibody therapies. PMID:26833127

  15. The Ubiquitin Ligase Stub1 Negatively Modulates Regulatory T cell Suppressive Activity by Promoting Degradation of the Transcription Factor Foxp3

    PubMed Central

    Yang, Huang-Yu; Li, Zhiyuan; Gao, Yayi; Jinasena, Dilini; Fu, Juan; Lin, Fang; Chen, Chen; Zhang, Jing; Yu, Ning; Li, Xiangpei; Shan, Zhao; Nie, Jia; Gao, Zhimei; Tian, Hong; Li, Yangyang; Yao, Zhengju; Zheng, Ying; Park, Benjamin V.; Pan, Ziyi; Zhang, Jing; Dang, Eric; Li, Zhiguang; Wang, Honglin; Luo, Weibo; Li, Liwu; Semenza, Gregg L.; Zheng, Song-Guo; Loser, Karin; Tsun, Andy; Greene, Mark I.; Pardoll, Drew M.; Pan, Fan; Li, Bin

    2013-01-01

    SUMMARY Regulatory T (Treg) cells suppress inflammatory immune responses and autoimmunity caused by self-reactive T cells. The key Treg cell transcription factor Foxp3 is downregulated during inflammation to allow for the acquisition of effector T cell-like functions. Here, we demonstrate that stress signals elicited by proinflammatory cytokines and lipopolysaccharide lead to the degradation of Foxp3 through the action of the E3 ubiquitin ligase Stub1. Stub1 interacted with Foxp3 to promote its K48-linked polyubiquitination in an Hsp70-dependent manner. Knockdown of endogenous Stub1 or Hsp70 prevented Foxp3 degradation. Furthermore, the overexpression of Stub1 in Treg cells abrogated their ability to suppress inflammatory immune responses in vitro and in vivo, and conferred a T helper 1 (Th1) cell-like phenotype. Our results demonstrate the critical role of the stress-activated Stub1-Hsp70 complex in promoting Treg cell inactivation, thus providing a potential therapeutic target for the intervention against autoimmune disease, infection and cancer. PMID:23973223

  16. Crystal structure of human T cell leukemia virus type 1 gp21 ectodomain crystallized as a maltose-binding protein chimera reveals structural evolution of retroviral transmembrane proteins

    PubMed Central

    Kobe, Bostjan; Center, Rob J.; Kemp, Bruce E.; Poumbourios, Pantelis

    1999-01-01

    Retroviral entry into cells depends on envelope glycoproteins, whereby receptor binding to the surface-exposed subunit triggers membrane fusion by the transmembrane protein (TM) subunit. We determined the crystal structure at 2.5-Å resolution of the ectodomain of gp21, the TM from human T cell leukemia virus type 1. The gp21 fragment was crystallized as a maltose-binding protein chimera, and the maltose-binding protein domain was used to solve the initial phases by the method of molecular replacement. The structure of gp21 comprises an N-terminal trimeric coiled coil, an adjacent disulfide-bonded loop that stabilizes a chain reversal, and a C-terminal sequence structurally distinct from HIV type 1/simian immunodeficiency virus gp41 that packs against the coil in an extended antiparallel fashion. Comparison of the gp21 structure with the structures of other retroviral TMs contrasts the conserved nature of the coiled coil-forming region and adjacent disulfide-bonded loop with the variable nature of the C-terminal ectodomain segment. The structure points to these features having evolved to enable the dual roles of retroviral TMs: conserved fusion function and an ability to anchor diverse surface-exposed subunit structures to the virion envelope and infected cell surface. The structure of gp21 implies that the N-terminal fusion peptide is in close proximity to the C-terminal transmembrane domain and likely represents a postfusion conformation. PMID:10200260

  17. Neutrophils confer T cell resistance to myeloid-derived suppressor cell-mediated suppression to promote chronic inflammation.

    PubMed

    Ryan, Sean O; Johnson, Jenny L; Cobb, Brian A

    2013-05-15

    Low-grade chronic inflammation can persist in aging humans unnoticed for years or even decades, inflicting continuous damage that can culminate later in life as organ dysfunction, physical frailty, and some of the most prominent debilitating and deadly age-associated diseases, including rheumatoid arthritis, diabetes, heart disease, and cancer. Despite the near universal acceptance of these associations, the mechanisms underlying unresolved inflammation remain poorly understood. In this study, we describe a novel inducible method to examine systemic chronic inflammation using susceptible animal models. Induced inflammation results in unresolved innate cellular responses and persistence of the same serum proinflammatory molecules used as diagnostic biomarkers and therapeutic targets for chronic inflammation in humans. Surprisingly, we found long-term persistence of an inflammation-associated neutrophil cell population constitutively producing the proinflammatory IFN-γ cytokine, which until now has only been detected transiently in acute inflammatory responses. Interestingly, these cells appear to confer T cell resistance to the otherwise potent anti-inflammatory function of myeloid-derived suppressor cells, revealing a novel mechanism for the maintenance of chronic inflammatory responses over time. This discovery represents an attractive target to resolve inflammation and prevent the inflammation-induced pathologies that are of critical concern for the well-being of the aging population. PMID:23576679

  18. MHC Class I-Restricted Myelin Epitopes Are Cross-Presented by Tip-DCs That Promote Determinant Spreading to CD8+ T Cells

    PubMed Central

    Ji, Qingyong; Castelli, Luca; Goverman, Joan M.

    2013-01-01

    Myelin presentation to T cells within the central nervous system (CNS) sustains inflammation in multiple sclerosis (MS). CD4+ and CD8+ T cells contribute to MS; however, only cells that present myelin to CD4+ T cells have been identified. We show that MHC class I-restricted myelin basic protein (MBP) was presented by oligodendrocytes and cross-presented by Tip-dendritic cells (DCs) during experimental autoimmune encephalomyelitis (EAE), an animal model of MS initiated by CD4+ T cells. Tip-DCs activated naïve and effector CD8+ T cells ex vivo, and naïve MBP-specific CD8+ T cells were activated within the CNS during CD4+ T cell-induced EAE. These results demonstrate that CD4+ T cell-mediated CNS autoimmunity leads to determinant spreading to myelin-specific CD8+ T cells that are capable of direct recognition of oligodendrocytes. PMID:23291597

  19. A novel regulatory pathway for autoimmune disease: Binding of partial MHC class II constructs to monocytes reduces CD74 expression and induces both specific and bystander T-cell tolerance

    PubMed Central

    Andrew, Shayne; Huan, Jianya; Chou, Yuan K.; Buenafe, Abigail C.; Dahan, Rony; Reiter, Yoram; Mooney, Jeffery L.; Offner, Halina; Burrows, Gregory G.

    2012-01-01

    Treatment with partial (p)MHC class II-β1α1 constructs (also referred to as recombinant T-cell receptor ligands – RTL) linked to antigenic peptides can induce T-cell tolerance, inhibit recruitment of inflammatory cells and reverse autoimmune diseases. Here we demonstrate a novel regulatory pathway that involves RTL binding to CD11b+ mononuclear cells through a receptor comprised of MHC class II invariant chain (CD74), cell-surface histones and MHC class II itself for treatment of experimental autoimmune encephalomyelitis (EAE). Binding of RTL constructs with CD74 involved a previously unrecognized MHC class II-α1/CD74 interaction that inhibited CD74 expression, blocked activity of its ligand, macrophage migration inhibitory factor, and reduced EAE severity. These findings implicate binding of RTL constructs to CD74 as a key step in both antigen-driven and bystander T-cell tolerance important in treatment of inflammatory diseases. PMID:23026773

  20. The Adenylate Cyclase Toxins of Bacillus anthracis and Bordetella pertussis Promote Th2 Cell Development by Shaping T Cell Antigen Receptor Signaling

    PubMed Central

    Rossi Paccani, Silvia; Benagiano, Marisa; Capitani, Nagaja; Zornetta, Irene; Ladant, Daniel; Montecucco, Cesare; D'Elios, Mario M.; Baldari, Cosima T.

    2009-01-01

    The adjuvanticity of bacterial adenylate cyclase toxins has been ascribed to their capacity, largely mediated by cAMP, to modulate APC activation, resulting in the expression of Th2–driving cytokines. On the other hand, cAMP has been demonstrated to induce a Th2 bias when present during T cell priming, suggesting that bacterial cAMP elevating toxins may directly affect the Th1/Th2 balance. Here we have investigated the effects on human CD4+ T cell differentiation of two adenylate cyclase toxins, Bacillus anthracis edema toxin (ET) and Bordetella pertussis CyaA, which differ in structure, mode of cell entry, and subcellular localization. We show that low concentrations of ET and CyaA, but not of their genetically detoxified adenylate cyclase defective counterparts, potently promote Th2 cell differentiation by inducing expression of the master Th2 transcription factors, c-maf and GATA-3. We also present evidence that the Th2–polarizing concentrations of ET and CyaA selectively inhibit TCR–dependent activation of Akt1, which is required for Th1 cell differentiation, while enhancing the activation of two TCR–signaling mediators, Vav1 and p38, implicated in Th2 cell differentiation. This is at variance from the immunosuppressive toxin concentrations, which interfere with the earliest step in TCR signaling, activation of the tyrosine kinase Lck, resulting in impaired CD3ζ phosphorylation and inhibition of TCR coupling to ZAP-70 and Erk activation. These results demonstrate that, notwithstanding their differences in their intracellular localization, which result in focalized cAMP production, both toxins directly affect the Th1/Th2 balance by interfering with the same steps in TCR signaling, and suggest that their adjuvanticity is likely to result from their combined effects on APC and CD4+ T cells. Furthermore, our results strongly support the key role of cAMP in the adjuvanticity of these toxins. PMID:19266022

  1. Regulatory T cells and antigen-specific tolerance.

    PubMed

    Kretschmer, Karsten; Apostolou, Irina; Verginis, Panos; von Boehmer, Harald

    2008-01-01

    Foxp3-expressing regulatory T cells (Tregs) have an essential function of preventing autoimmune disease in man and mouse. Foxp3 binds to forkhead motifs of about 1,100 genes and the strength of binding increases upon PMA/ionomycin stimulation. In Foxp3-expressing T-cell hybridomas, Foxp3 promoter binding does not lead to activation or suppression of genes which becomes only visible after T-cell activation. These findings are in line with observations by others that Foxp3 exerts important functions through association with Tcell receptor (TCR)-dependent transcription factors in a DNAbinding complex. Tregs can be generated when developing T cells encounter TCR agonist ligands in the thymus. This pro - cess requires costimulatory signals. In contrast, extra thymic conversion of naive T cells into Tregs is inhibited by costimulation. In fact, DC-derived retinoic acid (RA) helps the conversion process by counteracting the negative impact of costimulation. Since AP-1 is produced after costimulation and appears to interfere with Foxp3-NFAT transcription complexes, it is of interest to note that RA interferes with AP-1-dependent transcription. Thus, RA may interfere with the negative impact of costimulation on Treg conversion by interfering with the generation and/or function of AP-1. PMID:18802332

  2. Infection by HIV-1 blocked by binding of dextrin 2-sulphate to the cell surface of activated human peripheral blood mononuclear cells and cultured T-cells.

    PubMed Central

    Shaunak, S; Gooderham, N J; Edwards, R J; Payvandi, N; Javan, C M; Baggett, N; MacDermot, J; Weber, J N; Davies, D S

    1994-01-01

    1. Structural analogues of a sulphated polysaccharide, dextrin sulphate, were synthesized and tested for their ability to block infection by HIV-1. Using the T-cell lines, C8166 and HPB-ALL, and the laboratory adapted strains of HIV-1.MN, HIV-1.IIIb and HIV-1.RF, dextrin 2-sulphate (D2S) combined the best combination of high anti-HIV-1 activity (95% inhibitory concentration (IC95) = 230 nM) and low anticoagulant activity. It also blocked infection of activated peripheral blood mononuclear (PBMN) cells by five primary viral isolates at an IC95 of 230-3700 nM depending upon the primary viral isolate tested. 2. In saturation binding studies, [3H]-D2S bound to a cell surface protein on HPB-ALL cells in a specific and saturable manner with a Kd of 82 +/- 14 nM and a Bmax of 4.8 +/- 0.3 pmol/10(6) cells. It bound to other human T-cell lines in a similar manner. 3. There was very little binding of [3H]-D2S to freshly isolated PBMN cells (Bmax 0.18 +/- 0.03 pmol/10(6) cells) and these cells could not be infected by HIV-1. Culture of PBMN cells in lymphocyte growth medium (LGM) containing IL-2 did not significantly change the Bmax of [3H]-D2S. In contrast, PBMN cells which had been cultured with phytohaemagglutinin (PHA; 5 micrograms ml-1) for 72 h had a Bmax of [3H]-D2S binding of 7.2 +/- 0.1 pmol/10(6) cells and these cells could be infected by HIV-1. Removal of the PHA and further culture of the PBMN cells in LGM containing IL-2 resulted in a fall in the Bmax to 2.0 +/- 0.1 pmol/10(6) cells.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:7812605

  3. IL-2 / α-IL-2 Complex Treatment Cannot Be Substituted for the Adoptive Transfer of Regulatory T cells to Promote Bone Marrow Engraftment

    PubMed Central

    Mahr, Benedikt; Unger, Lukas; Hock, Karin; Pilat, Nina; Baranyi, Ulrike; Schwarz, Christoph; Maschke, Svenja; Farkas, Andreas Michael; Wekerle, Thomas

    2016-01-01

    Cell therapy with recipient Tregs achieves engraftment of allogeneic bone marrow (BM) without the need for cytoreductive conditioning (i.e., without irradiation or cytotoxic drugs). Thereby mixed chimerism and transplantation tolerance are established in recipients conditioned solely with costimulation blockade and rapamycin. However, clinical translation would be substantially facilitated if Treg-stimulating pharmaceutical agents could be used instead of individualized cell therapy. Recently, it was shown that interleukin-2 (IL-2) complexed with a monoclonal antibody (mAb) (clone JES6-1A12) against IL-2 (IL-2 complexes) potently expands and activates Tregs in vivo. Therefore, we investigated whether IL-2 complexes can replace Treg therapy in a costimulation blockade-based and irradiation-free BM transplantation (BMT) model. Unexpectedly, the administration of IL-2 complexes at the time of BMT (instead of Tregs) failed to induce BM engraftment in non-irradiated recipients (0/6 with IL-2 complexes vs. 3/4 with Tregs, p<0.05). Adding IL-2 complexes to an otherwise effective regimen involving recipient irradiation (1Gy) but no Treg transfer indeed actively triggered donor BM rejection at higher doses (0/8 with IL-2 complexes vs. 9/11 without, p<0.01) and had no detectable effect at two lower doses (3/5 vs. 9/11, p>0.05). CD8 T cells and NK cells of IL-2 complex-treated naïve mice showed an enhanced proliferative response towards donor antigens in vitro despite the marked expansion of Tregs. However, IL-2 complexes also expanded conventional CD4 T cells, CD8 T cells, NK cells, NKT cells and notably even B cells, albeit to a lesser extent. Notably, IL-2 complex expanded Tregs featured less potent suppressive functions than in vitro activated Tregs in terms of T cell suppression in vitro and BM engraftment in vivo. In conclusion, these data suggest that IL-2 complexes are less effective than recipient Tregs in promoting BM engraftment and in contrast actually trigger BM

  4. IL-2/α-IL-2 Complex Treatment Cannot Be Substituted for the Adoptive Transfer of Regulatory T cells to Promote Bone Marrow Engraftment.

    PubMed

    Mahr, Benedikt; Unger, Lukas; Hock, Karin; Pilat, Nina; Baranyi, Ulrike; Schwarz, Christoph; Maschke, Svenja; Farkas, Andreas Michael; Wekerle, Thomas

    2016-01-01

    Cell therapy with recipient Tregs achieves engraftment of allogeneic bone marrow (BM) without the need for cytoreductive conditioning (i.e., without irradiation or cytotoxic drugs). Thereby mixed chimerism and transplantation tolerance are established in recipients conditioned solely with costimulation blockade and rapamycin. However, clinical translation would be substantially facilitated if Treg-stimulating pharmaceutical agents could be used instead of individualized cell therapy. Recently, it was shown that interleukin-2 (IL-2) complexed with a monoclonal antibody (mAb) (clone JES6-1A12) against IL-2 (IL-2 complexes) potently expands and activates Tregs in vivo. Therefore, we investigated whether IL-2 complexes can replace Treg therapy in a costimulation blockade-based and irradiation-free BM transplantation (BMT) model. Unexpectedly, the administration of IL-2 complexes at the time of BMT (instead of Tregs) failed to induce BM engraftment in non-irradiated recipients (0/6 with IL-2 complexes vs. 3/4 with Tregs, p<0.05). Adding IL-2 complexes to an otherwise effective regimen involving recipient irradiation (1Gy) but no Treg transfer indeed actively triggered donor BM rejection at higher doses (0/8 with IL-2 complexes vs. 9/11 without, p<0.01) and had no detectable effect at two lower doses (3/5 vs. 9/11, p>0.05). CD8 T cells and NK cells of IL-2 complex-treated naïve mice showed an enhanced proliferative response towards donor antigens in vitro despite the marked expansion of Tregs. However, IL-2 complexes also expanded conventional CD4 T cells, CD8 T cells, NK cells, NKT cells and notably even B cells, albeit to a lesser extent. Notably, IL-2 complex expanded Tregs featured less potent suppressive functions than in vitro activated Tregs in terms of T cell suppression in vitro and BM engraftment in vivo. In conclusion, these data suggest that IL-2 complexes are less effective than recipient Tregs in promoting BM engraftment and in contrast actually trigger BM

  5. Tandem CAR T cells targeting HER2 and IL13Rα2 mitigate tumor antigen escape.

    PubMed

    Hegde, Meenakshi; Mukherjee, Malini; Grada, Zakaria; Pignata, Antonella; Landi, Daniel; Navai, Shoba A; Wakefield, Amanda; Fousek, Kristen; Bielamowicz, Kevin; Chow, Kevin K H; Brawley, Vita S; Byrd, Tiara T; Krebs, Simone; Gottschalk, Stephen; Wels, Winfried S; Baker, Matthew L; Dotti, Gianpietro; Mamonkin, Maksim; Brenner, Malcolm K; Orange, Jordan S; Ahmed, Nabil

    2016-08-01

    In preclinical models of glioblastoma, antigen escape variants can lead to tumor recurrence after treatment with CAR T cells that are redirected to single tumor antigens. Given the heterogeneous expression of antigens on glioblastomas, we hypothesized that a bispecific CAR molecule would mitigate antigen escape and improve the antitumor activity of T cells. Here, we created a CAR that joins a HER2-binding scFv and an IL13Rα2-binding IL-13 mutein to make a tandem CAR exodomain (TanCAR) and a CD28.ζ endodomain. We determined that patient TanCAR T cells showed distinct binding to HER2 or IL13Rα2 and had the capability to lyse autologous glioblastoma. TanCAR T cells exhibited activation dynamics that were comparable to those of single CAR T cells upon encounter of HER2 or IL13Rα2. We observed that TanCARs engaged HER2 and IL13Rα2 simultaneously by inducing HER2-IL13Rα2 heterodimers, which promoted superadditive T cell activation when both antigens were encountered concurrently. TanCAR T cell activity was more sustained but not more exhaustible than that of T cells that coexpressed a HER2 CAR and an IL13Rα2 CAR, T cells with a unispecific CAR, or a pooled product. In a murine glioblastoma model, TanCAR T cells mitigated antigen escape, displayed enhanced antitumor efficacy, and improved animal survival. Thus, TanCAR T cells show therapeutic potential to improve glioblastoma control by coengaging HER2 and IL13Rα2 in an augmented, bivalent immune synapse that enhances T cell functionality and reduces antigen escape. PMID:27427982

  6. Cutting edge: identification of a novel chemokine receptor that binds dendritic cell- and T cell-active chemokines including ELC, SLC, and TECK.

    PubMed

    Gosling, J; Dairaghi, D J; Wang, Y; Hanley, M; Talbot, D; Miao, Z; Schall, T J

    2000-03-15

    Searching for new receptors of dendritic cell- and T cell-active chemokines, we used a combination of techniques to interrogate orphan chemokine receptors. We report here on human CCX CKR, previously represented only by noncontiguous expressed sequence tags homologous to bovine PPR1, a putative gustatory receptor. We employed a two-tiered process of ligand assignment, where immobilized chemokines constructed on stalks (stalkokines) were used as bait for adhesion of cells expressing CCX CKR. These cells adhered to stalkokines representing ELC, a chemokine previously thought to bind only CCR7. Adhesion was abolished in the presence of soluble ELC, SLC (CCR7 ligands), and TECK (a CCR9 ligand). Complete ligand profiles were further determined by radiolabeled ligand binding and competition with >80 chemokines. ELC, SLC, and TECK comprised high affinity ligands (IC50 <15 nM); lower affinity ligands include BLC and vMIP-II (IC50 <150 nM). With its high affinity for CC chemokines and homology to CC receptors, we provisionally designate this new receptor CCR10. PMID:10706668

  7. Liposomal vaccines incorporating molecular adjuvants and intrastructural T-cell help promote the immunogenicity of HIV membrane-proximal external region peptides

    PubMed Central

    Hanson, Melissa C.; Abraham, Wuhbet; Crespo, Monica P.; Chen, Stephanie H.; Liu, Haipeng; Szeto, Greg Lee; Kim, Mikyung; Reinherz, Ellis L.; Irvine, Darrell J.

    2015-01-01

    An HIV vaccine capable of inducing high and durable levels of broadly neutralizing antibodies has thus far proven elusive. A promising antigen is the membrane-proximal external region (MPER) from gp41, a segment of the viral envelope recognized by a number of broadly neutralizing antibodies. Though an attractive vaccine target due to the linear nature of the epitope and its highly conserved sequence, MPER peptides are poorly immunogenic and may require display on membranes to achieve a physiological conformation matching the native virus. Here we systematically explored how the structure and composition of liposomes displaying MPER peptides impacts the strength and durability of humoral responses to this antigen as well as helper T-cell responses in mice. Administration of MPER peptides anchored to the surface of liposomes induced MPER-specific antibodies whereas MPER administered in oil-based emulsion adjuvants or alum did not, even when combined with Toll like receptor agonists. High-titer IgG responses to liposomal MPER required the inclusion of molecular adjuvants such as monophosphoryl lipid A. Anti-MPER humoral responses were further enhanced by incorporating high-Tm lipids in the vesicle bilayer and optimizing the MPER density to a mean distance of ~10–15 nm between peptides on the liposomes' surfaces. Encapsulation of helper epitopes within the vesicles allowed efficient “intrastructural” T-cell help, which promoted IgG responses to MPER while minimizing competing B-cell responses against the helper sequence. These results define several key properties of liposome formulations that promote durable, high-titer antibody responses against MPER peptides, which will be a prerequisite for a successful MPER-targeting vaccine. PMID:25559188

  8. Complement component C5a Promotes Expression of IL-22 and IL-17 from Human T cells and its Implication in Age-related Macular Degeneration

    PubMed Central

    2011-01-01

    Background Age related macular degeneration (AMD) is the leading cause of irreversible blindness in elderly populations worldwide. Inflammation, among many factors, has been suggested to play an important role in AMD pathogenesis. Recent studies have demonstrated a strong genetic association between AMD and complement factor H (CFH), the down-regulatory factor of complement activation. Elevated levels of complement activating molecules including complement component 5a (C5a) have been found in the serum of AMD patients. Our aim is to study whether C5a can impact human T cells and its implication in AMD. Methods Human peripheral blood mononuclear cells (PBMCs) were isolated from the blood of exudative form of AMD patients using a Ficoll gradient centrifugation protocol. Intracellular staining and enzyme-linked immunosorbent assays were used to measure protein expression. Apoptotic cells were detected by staining of cells with the annexin-V and TUNEL technology and analyzed by a FACS Caliber flow cytometer. SNP genotyping was analyzed by TaqMan genotyping assay using the Real-time PCR system 7500. Results We show that C5a promotes interleukin (IL)-22 and IL-17 expression by human CD4+ T cells. This effect is dependent on B7, IL-1β and IL-6 expression from monocytes. We have also found that C5a could protect human CD4+ cells from undergoing apoptosis. Importantly, consistent with a role of C5a in promoting IL-22 and IL-17 expression, significant elevation in IL-22 and IL-17 levels was found in AMD patients as compared to non-AMD controls. Conclusions Our results support the notion that C5a may be one of the factors contributing to the elevated serum IL-22 and IL-17 levels in AMD patients. The possible involvement of IL-22 and IL-17 in the inflammation that contributes to AMD may herald a new approach to treat AMD. PMID:21762495

  9. TpF1 from Treponema pallidum activates inflammasome and promotes the development of regulatory T cells.

    PubMed

    Babolin, Chiara; Amedei, Amedeo; Ozolins, Dzintars; Zilevica, Aija; D'Elios, Mario Milco; de Bernard, Marina

    2011-08-01

    Human syphilis is a multistage disease, with diverse and wide-ranging manifestations caused by Treponema pallidum. Despite the fact that a cell-mediated immune response takes part in the course of syphilis, T. pallidum often manages to evade host immunity and, in untreated individuals, may trigger chronic infection. With this study, we demonstrate for the first time, to our knowledge, that Treponema pallidum induces a regulatory T (Treg) response in patients with secondary syphilis and we found that the miniferritin TpF1, produced by the bacterium, is able to expand this response and promote the production of TGF-β. Accordingly, TpF1 stimulates monocytes to release IL-10 and TGF-β, the key cytokines in driving Treg cell differentiation. Interestingly, we also found that TpF1 stimulates monocytes to synthesize and release several proinflammatory cytokines, such as TNF-α, IL-6, and IL-1β, the latter following the activation of the multiprotein complex inflammasome. Collectively, these data strongly support a central role for TpF1 both in the inflammation process, which occurs in particular during the early stage of syphilis, and in the long-term persistence of the spirochete within the host by promoting Treg response and TGF-β production. PMID:21709157

  10. Transcription Factor Binding Site Positioning in Yeast: Proximal Promoter Motifs Characterize TATA-Less Promoters

    PubMed Central

    Erb, Ionas; van Nimwegen, Erik

    2011-01-01

    The availability of sequence specificities for a substantial fraction of yeast's transcription factors and comparative genomic algorithms for binding site prediction has made it possible to comprehensively annotate transcription factor binding sites genome-wide. Here we use such a genome-wide annotation for comprehensively studying promoter architecture in yeast, focusing on the distribution of transcription factor binding sites relative to transcription start sites, and the architecture of TATA and TATA-less promoters. For most transcription factors, binding sites are positioned further upstream and vary over a wider range in TATA promoters than in TATA-less promoters. In contrast, a group of ‘proximal promoter motifs’ (GAT1/GLN3/DAL80, FKH1/2, PBF1/2, RPN4, NDT80, and ROX1) occur preferentially in TATA-less promoters and show a strong preference for binding close to the transcription start site in these promoters. We provide evidence that suggests that pre-initiation complexes are recruited at TATA sites in TATA promoters and at the sites of the other proximal promoter motifs in TATA-less promoters. TATA-less promoters can generally be classified by the proximal promoter motif they contain, with different classes of TATA-less promoters showing different patterns of transcription factor binding site positioning and nucleosome coverage. These observations suggest that different modes of regulation of transcription initiation may be operating in the different promoter classes. In addition we show that, across all promoter classes, there is a close match between nucleosome free regions and regions of highest transcription factor binding site density. This close agreement between transcription factor binding site density and nucleosome depletion suggests a direct and general competition between transcription factors and nucleosomes for binding to promoters. PMID:21931670

  11. Tumor-induced senescent T cells promote the secretion of pro-inflammatory cytokines and angiogenic factors by human monocytes/macrophages through a mechanism that involves Tim-3 and CD40L

    PubMed Central

    Ramello, M C; Tosello Boari, J; Canale, F P; Mena, H A; Negrotto, S; Gastman, B; Gruppi, A; Acosta Rodríguez, E V; Montes, C L

    2014-01-01

    Solid tumors are infiltrated by immune cells where macrophages and senescent T cells are highly represented. Within the tumor microenvironment, a cross-talk between the infiltrating cells may occur conditioning the characteristic of the in situ immune response. Our previous work showed that tumors induce senescence of T cells, which are powerful suppressors of lympho-proliferation. In this study, we report that Tumor-Induced Senescent (TIS)-T cells may also modulate monocyte activation. To gain insight into this interaction, CD4+ or CD8+TIS-T or control-T cells were co-incubated with autologous monocytes under inflammatory conditions. After co-culture with CD4+ or CD8+TIS-T cells, CD14+ monocytes/macrophages (Mo/Ma) exhibit a higher expression of CD16+ cells and a reduced expression of CD206. These Mo/Ma produce nitric oxide and reactive oxygen species; however, TIS-T cells do not modify phagocyte capacity of Mo/Ma. TIS-T modulated-Mo/Ma show a higher production of pro-inflammatory cytokines (TNF, IL-1β and IL-6) and angiogenic factors (MMP-9, VEGF-A and IL-8) and a lower IL-10 and IP-10 secretion than monocytes co-cultured with controls. The mediator(s) present in the supernatant of TIS-T cell/monocyte-macrophage co-cultures promote(s) tubulogenesis and tumor-cell survival. Monocyte-modulation induced by TIS-T cells requires cell-to-cell contact. Although CD4+ shows different behavior from CD8+TIS-T cells, blocking mAbs against T-cell immunoglobulin and mucin protein 3 and CD40 ligand reduce pro-inflammatory cytokines and angiogenic factors production, indicating that these molecules are involved in monocyte/macrophage modulation by TIS-T cells. Our results revealed a novel role for TIS-T cells in human monocyte/macrophage modulation, which may have deleterious consequences for tumor progression. This modulation should be considered to best tailor the immunotherapy against cancer. PMID:25375372

  12. Follicular Helper T Cells.

    PubMed

    Vinuesa, Carola G; Linterman, Michelle A; Yu, Di; MacLennan, Ian C M

    2016-05-20

    Although T cell help for B cells was described several decades ago, it was the identification of CXCR5 expression by B follicular helper T (Tfh) cells and the subsequent discovery of their dependence on BCL6 that led to the recognition of Tfh cells as an independent helper subset and accelerated the pace of discovery. More than 20 transcription factors, together with RNA-binding proteins and microRNAs, control the expression of chemotactic receptors and molecules important for the function and homeostasis of Tfh cells. Tfh cells prime B cells to initiate extrafollicular and germinal center antibody responses and are crucial for affinity maturation and maintenance of humoral memory. In addition to the roles that Tfh cells have in antimicrobial defense, in cancer, and as HIV reservoirs, regulation of these cells is critical to prevent autoimmunity. The realization that follicular T cells are heterogeneous, comprising helper and regulatory subsets, has raised questions regarding a possible division of labor in germinal center B cell selection and elimination. PMID:26907215

  13. The immunodominant CD8+ T cell epitope region of Theiler’s virus in resistant C57BL/6 mice is critical for anti-viral immune responses, viral persistence, and binding to the host cells.

    PubMed Central

    Myoung, Jinjong; Hou, Wanqiu; Kang, Bongsu; Lyman, Michael A.; Kang, Jeong-Ah; Kim, Byung S.

    2007-01-01

    Theiler’s virus infection induces an immune-mediated demyelinating disease, providing a relevant animal model of human multiple sclerosis. VP2121–130-specific CD8+ T cells in resistant H-2b mice account for the majority of CNS-infiltrating CD8+ T cells. To further study the role of the CD8+ T cells, we generated a panel of mutant viruses substituted with L, G, or T at the anchor residue (M130) of the VP2121–130 epitope. M130L virus (M130L-V) with a substitution of M with L displayed similar properties as wild-type virus (WT-V). However, M130G-V and M130T-V could not establish a persistent infection in the CNS. The level of both virus-specific CD8+ and CD4+ T cell responses is significantly reduced in mice infected with these variant viruses. While all mutant and wild-type viruses replicate comparably in BHK cells, replication of M130G-V and M130T-V in macrophages was significantly lower compared to those infected with WT-V and M130L-V. Interestingly, these mutant viruses deficient in replication in primary mouse cells showed drastically reduced binding ability to the cells. These results suggest that the anchor residue of the predominant CD8+ T cell epitope of TMEV in resistant mice is critical for the virus to infect target cells and this deficiency may result in poor viral persistence leading to correspondingly low T cell responses in the periphery and CNS. Thus, selection of the cellular binding region of the virus as the predominant epitope for CD8+ T cells in resistant mice may provide a distinct advantage in controlling viral persistence by preventing escape mutations. PMID:17095033

  14. Crystallization of a trimeric human T cell leukemia virus type 1 gp21 ectodomain fragment as a chimera with maltose-binding protein.

    PubMed Central

    Center, R. J.; Kobe, B.; Wilson, K. A.; Teh, T.; Howlett, G. J.; Kemp, B. E.; Poumbourios, P.

    1998-01-01

    We present a novel protein crystallization strategy, applied to the crystallization of human T cell leukemia virus type 1 (HTLV-1) transmembrane protein gp21 lacking the fusion peptide and the transmembrane domain, as a chimera with the Escherichia coli maltose binding protein (MBP). Crystals could not be obtained with a MBP/gp21 fusion protein in which fusion partners were separated by a flexible linker, but were obtained after connecting the MBP C-terminal alpha-helix to the predicted N-terminal alpha-helical sequence of gp21 via three alanine residues. The gp21 sequences conferred a trimeric structure to the soluble fusion proteins as assessed by sedimentation equilibrium and X-ray diffraction, consistent with the trimeric structures of other retroviral transmembrane proteins. The envelope protein precursor, gp62, is likewise trimeric when expressed in mammalian cells. Our results suggest that MBP may have a general application for the crystallization of proteins containing N-terminal alpha-helical sequences. PMID:9684894

  15. Inhaled house dust programs pulmonary dendritic cells to promote type 2 T-cell responses by an indirect mechanism.

    PubMed

    Moran, Timothy P; Nakano, Keiko; Whitehead, Gregory S; Thomas, Seddon Y; Cook, Donald N; Nakano, Hideki

    2015-11-15

    The induction of allergen-specific T helper 2 (Th2) cells by lung dendritic cells (DCs) is a critical step in allergic asthma development. Airway delivery of purified allergens or microbial products can promote Th2 priming by lung DCs, but how environmentally relevant quantities and combinations of these factors affect lung DC function is unclear. Here, we investigated the ability of house dust extract (HDE), which contains a mixture of environmental adjuvants, to prime Th2 responses against an innocuous inhaled antigen. Inhalational exposure to HDE conditioned lung conventional DCs, but not monocyte-derived DCs, to induce antigen-specific Th2 differentiation. Conditioning of DCs by HDE was independent of Toll-like receptor 4 signaling, indicating that environmental endotoxin is dispensable for programming DCs to induce Th2 responses. DCs directly treated with HDE underwent maturation but were poor stimulators of Th2 differentiation. In contrast, DCs treated with bronchoalveolar lavage fluid (BALF) from HDE-exposed mice induced robust Th2 differentiation. DC conditioning by BALF was independent of the proallergic cytokines IL-25, IL-33, and thymic stromal lymphopoietin. BALF treatment of DCs resulted in upregulation of CD80 but low expression of CD40, CD86, and IL-12p40, which was associated with Th2 induction. These findings support a model whereby environmental adjuvants in house dust indirectly program DCs to prime Th2 responses by triggering the release of endogenous soluble factor(s) by airway cells. Identifying these factors could lead to novel therapeutic targets for allergic asthma. PMID:26386119

  16. miR-664 negatively regulates PLP2 and promotes cell proliferation and invasion in T-cell acute lymphoblastic leukaemia

    SciTech Connect

    Zhu, Hong; Miao, Mei-hua; Ji, Xue-qiang; Xue, Jun; Shao, Xue-jun

    2015-04-03

    MicroRNAs (miRNAs) play important roles in the pathogenesis of many types of cancers by negatively regulating gene expression at posttranscriptional level. However, the role of microRNAs in leukaemia, particularly T-cell acute lymphoblastic leukaemia (T-ALL), has remained elusive. Here, we identified miR-664 and its predicted target gene PLP2 were differentially expressed in T-ALL using bioinformatics methods. In T-ALL cell lines, CCK-8 proliferation assay indicated that the cell proliferation was promoted by miR-664, while miR-664 inhibitor could significantly inhibited the proliferation. Moreover, migration and invasion assay showed that overexpression of miR-664 could significantly promoted the migration and invasion of T-ALL cells, whereas miR-664 inhibitor could reduce cell migration and invasion. luciferase assays confirmed that miR-664 directly bound to the 3'untranslated region of PLP2, and western blotting showed that miR-664 suppressed the expression of PLP2 at the protein levels. This study indicated that miR-664 negatively regulates PLP2 and promotes proliferation and invasion of T-ALL cell lines. Thus, miR-664 may represent a potential therapeutic target for T-ALL intervention. - Highlights: • miR-664 mimics promote the proliferation and invasion of T-ALL cells. • miR-664 inhibitors inhibit the proliferation and invasion of T-ALL cells. • miR-664 targets 3′ UTR of PLP2 in T-ALL cells. • miR-664 negatively regulates PLP2 in T-ALL cells.

  17. Transfer of regulatory T cells into abortion-prone mice promotes the expansion of uterine mast cells and normalizes early pregnancy angiogenesis

    PubMed Central

    Woidacki, Katja; Meyer, Nicole; Schumacher, Anne; Goldschmidt, Alexandra; Maurer, Marcus; Zenclussen, Ana Claudia

    2015-01-01

    Implantation of the fertilized egg depends on the coordinated interplay of cells and molecules that prepare the uterus for this important event. In particular, regulatory T cells (Tregs) are key regulators as their ablation hinders implantation by rendering the uterus hostile for the embryo. In addition, the adoptive transfer of Tregs can avoid early abortion in mouse models. However, it is still not defined which mechanisms underlie Treg function during this early period. Cells of the innate immune system have been reported to support implantation, in part by promoting angiogenesis. In particular, uterine mast cells (uMCs) emerge as novel players at the fetal-maternal interface. Here, we studied whether the positive action of Tregs is based on the expansion of uMCs and the promotion of angiogenesis. We observed that abortion-prone mice have insufficient numbers of uMCs that could be corrected by the adoptive transfer of Tregs. This in turn positively influenced the remodeling of spiral arteries and placenta development as well as the levels of soluble fms-like tyrosine kinase 1 (sFlt-1). Our data suggest an interplay between Tregs and uMCs that is relevant for the changes required at the feto-maternal interface for the normal development of pregnancy. PMID:26355667

  18. Transfer of regulatory T cells into abortion-prone mice promotes the expansion of uterine mast cells and normalizes early pregnancy angiogenesis.

    PubMed

    Woidacki, Katja; Meyer, Nicole; Schumacher, Anne; Goldschmidt, Alexandra; Maurer, Marcus; Zenclussen, Ana Claudia

    2015-01-01

    Implantation of the fertilized egg depends on the coordinated interplay of cells and molecules that prepare the uterus for this important event. In particular, regulatory T cells (Tregs) are key regulators as their ablation hinders implantation by rendering the uterus hostile for the embryo. In addition, the adoptive transfer of Tregs can avoid early abortion in mouse models. However, it is still not defined which mechanisms underlie Treg function during this early period. Cells of the innate immune system have been reported to support implantation, in part by promoting angiogenesis. In particular, uterine mast cells (uMCs) emerge as novel players at the fetal-maternal interface. Here, we studied whether the positive action of Tregs is based on the expansion of uMCs and the promotion of angiogenesis. We observed that abortion-prone mice have insufficient numbers of uMCs that could be corrected by the adoptive transfer of Tregs. This in turn positively influenced the remodeling of spiral arteries and placenta development as well as the levels of soluble fms-like tyrosine kinase 1 (sFlt-1). Our data suggest an interplay between Tregs and uMCs that is relevant for the changes required at the feto-maternal interface for the normal development of pregnancy. PMID:26355667

  19. Shifting the Evolving CAR T Cell Platform into Higher Gear.

    PubMed

    Holohan, Daniel R; Lee, James C; Bluestone, Jeffrey A

    2015-10-12

    In this issue of Cancer Cell, Zhao and colleagues test various chimeric antigen receptor (CAR) T cells to show that CD28-CD3ζ CAR T cells that constitutively express 4-1BBL promote T cell expansion and tumor eradication while reducing exhaustion. The results have important implications for the development of effective CAR T cell therapies in cancer patients. PMID:26461084

  20. Nab2 regulates secondary CD8+ T-cell responses through control of TRAIL expression

    PubMed Central

    Gerlach, Carmen; Arens, Ramon; Janssen, Edith M.; Fitzgerald, Patrick; Schumacher, Ton N.; Medema, Jan Paul; Green, Douglas R.; Schoenberger, Stephen P.

    2012-01-01

    CD4+ Th cells are pivotal for the generation and maintenance of CD8+ T-cell responses. “Helped” CD8+ T cells receive signals during priming that prevent the induction of the proapoptotic molecule TNF-related apoptosis-inducing ligand (TRAIL) during reactivation, thereby enabling robust secondary expansion. Conversely, “helpless” CD8+ T cells primed in the absence of Th induce TRAIL expression after restimulation and undergo activation-induced cell death. In the present study, we investigated the molecular basis for the differential regulation of TRAIL in helped versus helpless CD8+ T cells by comparing their transcriptional profiles, and have identified a transcriptional corepressor, NGFI-A binding protein 2 (Nab2), that is selectively induced in helped CD8+ T cells. Enforced expression of Nab2 prevents TRAIL induction after restimulation of primary helpless CD8+ T cells, and expression of a dominant-negative form of Nab2 in helped CD8+ T cells impairs their secondary proliferative response that is reversible by TRAIL blockade. Finally, we observe that the CD8+ T-cell autocrine growth factor IL-2 coordinately increases Nab2 expression and decreases TRAIL expression. These findings identify Nab2 as a mediator of Th-dependent CD8+ T-cell memory responses through the regulation of TRAIL and the promotion of secondary expansion, and suggest a mechanism through which this operates. PMID:22128144

  1. Tregs utilize beta-galactoside-binding protein to transiently inhibit PI3K/p21ras activity of human CD8+ T cells to block their TCR-mediated ERK activity and proliferation.

    PubMed

    Baatar, Dolgor; Olkhanud, Purevdorj B; Wells, Valerie; Indig, Fred E; Mallucci, Livio; Biragyn, Arya

    2009-10-01

    Regulatory T cells (Tregs) and beta-galactoside-binding protein (betaGBP), a regulatory protein often found expressed at sites of immunological privilege, have similar functions. Their presence affects the outcome of harmful autoimmunity and cancers, including experimental autoimmune encephalomyelitis and malignant gliomas. Here we report a novel pathway by which Tregs express and utilize betaGBP to control CD8(+) T cell responses partially activating TCR signaling but blocking PI3K activity. As a result, this leads to a loss of p21(ras), ERK and Akt activities despite activation of TCR proximal signals, such as phosphorylation of CD3zeta, Zap70, Lat and PKCtheta. Although non-processive TCR signaling often leads to cell anergy, Tregs/betaGBP did not affect cell viability. Instead, betaGBP/Tregs transiently prevented activation of CD8(+) T cells with self-antigens, while keeping their responses to xenogeneic antigens unaffected. PMID:19520156

  2. Regulation of the T cell response.

    PubMed

    Romagnani, S

    2006-11-01

    The T cell branch of the immune system can respond to a virtually infinite variety of exogenous antigens, thus including the possibility of self-antigen recognition and dangerous autoimmune reactions. Therefore, regulatory mechanisms operate both during ontogeny within the thymus and after birth in the periphery. The control of self-reactive T cells occurs through a process of negative selection that results in apoptosis of T cells showing high affinity for self-peptides expressed at the thymic level by means of promiscuous gene expression. Self-reactive T cells escaped to negative selection are controlled in the periphery by other regulatory mechanisms, the most important being natural Foxp3+ T regulatory (Treg) cells. Regulation is also required to control excessive effector T cell responses against exogenous antigens, when they become dangerous for the body. Three types of effector T cells have been recognized: T helper 1 (Th1) cells, which are protective against intracellular bacteria; Th2 cells, which play some role in the protection against nematodes, but are responsible for allergic reactions; Th17 cells, which are probably effective in the protection against extracellular bacteria, but also play a role in the amplification of autoimmune disorders. Abnormal or excessive Th effector responses are regulated by different mechanisms. Redirection or immune deviation of Th1- or Th2-dominated responses is provided by cytokines [interferon-gamma (IFN-gamma) vs. interleukin-4 (IL-4)] produced by the same cell types and by the CXCR3-binding chemokines CXCL4 and CXCL10. Moreover, both Th1 and Th2 responses can be suppressed by adaptive Treg cells through contact-dependent mechanisms and/or the production of IL-10 and transforming growth factor-beta (TGF-beta). Finally, TGF-beta1 can promote the development of both Th17 effector and adaptive Treg cells, while the contemporaneous production of IL-6 contributes to the development of Th17 cells, but inhibits Treg cells

  3. T cell responses to cytomegalovirus.

    PubMed

    Klenerman, Paul; Oxenius, Annette

    2016-06-01

    Human cytomegalovirus (HCMV) establishes a latent infection that generally remains asymptomatic in immune-competent hosts for decades but can cause serious illness in immune-compromised individuals. The long-term control of CMV requires considerable effort from the host immune system and has a lasting impact on the profile of the immune system. One hallmark of CMV infection is the maintenance of large populations of CMV-specific memory CD8(+) T cells - a phenomenon termed memory inflation - and emerging data suggest that memory inflation is associated with impaired immunity in the elderly. In this Review, we discuss the molecular triggers that promote memory inflation, the idea that memory inflation could be considered a natural pathway of T cell maturation that could be harnessed in vaccination, and the broader implications of CMV infection and the T cell responses it elicits. PMID:27108521

  4. IL-21-Induced MHC Class II+ NK Cells Promote the Expansion of Human Uncommitted CD4+ Central Memory T Cells in a Macrophage Migration Inhibitory Factor-Dependent Manner.

    PubMed

    Loyon, Romain; Picard, Emilie; Mauvais, Olivier; Queiroz, Lise; Mougey, Virginie; Pallandre, Jean-René; Galaine, Jeanne; Mercier-Letondal, Patricia; Kellerman, Guillaume; Chaput, Nathalie; Wijdenes, John; Adotévi, Olivier; Ferrand, Christophe; Romero, Pedro; Godet, Yann; Borg, Christophe

    2016-07-01

    NK cells are critical for innate immunity-mediated protection. The main roles of NK cells rely on their cytotoxic functions or depend on the tuning of Th1 adaptive immunity by IFN-γ. However, the precise influence of inflammatory cytokines on NK cell and CD4 T lymphocyte interactions was never investigated. In this study, we provide evidence that IL-21, a cytokine produced during chronic inflammation or infectious diseases, promotes the differentiation of a specific subset of NK cells coexpressing CD86 and HLA-DR and lacking NKp44. More importantly, IL-21-propagated HLA-DR(+) NK cells produce macrophage migration inhibitory factor and provide costimulatory signaling during naive CD4(+) T cell priming inducing the differentiation of uncommitted central memory T cells. Central memory T cells expanded in the presence of HLA-DR(+) NK cells are CXCR3(+)CCR6(-)CCR4(-)CXCR5(-) and produce IL-2, as well as low levels of TNF-α. Costimulation of CD4(+) T cells by HLA-DR(+) NK cells prevents the acquisition of effector memory phenotype induced by IL-2. Moreover, we identified this population of NK HLA-DR(+) macrophage migration inhibitory factor(+) cells in inflammatory human appendix. Collectively, these results demonstrate a novel function for IL-21 in tuning NK and CD4(+) T cell interactions promoting a specific expansion of central memory lymphocytes. PMID:27233967

  5. Antigen-presenting cells containing multiple costimulatory molecules promote activation and expansion of antigen-specific memory CD8+ T cells

    PubMed Central

    Yang, Sixun; Schlom, Jeffrey

    2009-01-01

    We have previously demonstrated that multiple immunizations with vector-based vaccines containing transgenes for tumor Ags and a triad of costimulatory molecules (TRICOM) enhance the expansion and functional avidity of Ag-specific memory CD8+ T cells in a mouse model. However, the effect of enhanced costimulation on human memory CD8+ T cells is still unclear. The study reported here was an in vitro investigation of the proliferation and function of CEA-specific human memory CD8+ T cells following enhanced costimulation. Our results demonstrated that TRICOM costimulation enhanced production of multiple cytokines and expansion of CEA-specific memory CD8+ T cells. The lytic capacity of memory CTLs toward CEA+ tumors was also significantly enhanced. IL-2Rα (CD25) was upregulated dramatically following APC-TRICOM stimulation, suggesting that the enhanced expansion of memory CD8+ T cells may be mediated by increased expression of IL-2R on memory T cells. The enhanced cytokine production and proliferation following TRICOM signaling was completely blocked by the combination of neutralizing Abs against B7-1, ICAM-1, and LFA-3, the costimulatory molecules comprising TRICOM. No difference in T-cell apoptosis was observed between APC-TRICOM and APC-wild-type groups, as determined by annexin V, Bcl-2, and active caspase-3 staining. Results indicated that enhanced costimulation greatly expanded human CEA-specific CD8+ T cells and enhanced T-cell function, without inducing increased apoptosis of CEA-specific memory CD8+ T cells. PMID:18690438

  6. Tumor-evoked regulatory B cells promote breast cancer metastasis by converting resting CD4⁺ T cells to T-regulatory cells.

    PubMed

    Olkhanud, Purevdorj B; Damdinsuren, Bazarragchaa; Bodogai, Monica; Gress, Ronald E; Sen, Ranjan; Wejksza, Katarzyna; Malchinkhuu, Enkhzol; Wersto, Robert P; Biragyn, Arya

    2011-05-15

    Pulmonary metastasis of breast cancer requires recruitment and expansion of T-regulatory cells (Treg) that promote escape from host protective immune cells. However, it remains unclear precisely how tumors recruit Tregs to support metastatic growth. Here we report the mechanistic involvement of a unique and previously undescribed subset of regulatory B cells. These cells, designated tumor-evoked Bregs (tBreg), phenotypically resemble activated but poorly proliferative mature B2 cells (CD19(+) CD25(High) CD69(High)) that express constitutively active Stat3 and B7-H1(High) CD81(High) CD86(High) CD62L(Low) IgM(Int). Our studies with the mouse 4T1 model of breast cancer indicate that the primary role of tBregs in lung metastases is to induce TGF-β-dependent conversion of FoxP3(+) Tregs from resting CD4(+) T cells. In the absence of tBregs, 4T1 tumors cannot metastasize into the lungs efficiently due to poor Treg conversion. Our findings have important clinical implications, as they suggest that tBregs must be controlled to interrupt the initiation of a key cancer-induced immunosuppressive event that is critical to support cancer metastasis. PMID:21444674

  7. Monocytic CCR2+ Myeloid Derived Suppressor Cells Promote Immune Escape By Limiting Activated CD8 T Cell Infiltration Into The Tumor Microenvironment

    PubMed Central

    Lesokhin, Alexander M.; Hohl, Tobias M.; Kitano, Shigehisa; Cortez, Czrina; Hirschhorn-Cymerman, Daniel; Avogadri, Francesca; Rizzuto, Gabrielle A.; Lazarus, John J.; Pamer, Eric G.; Houghton, Alan N.; Merghoub, Taha; Wolchok, Jedd D.

    2011-01-01

    Myeloid derived suppressor cells (MDSC) are a heterogeneous population of cells that accumulate during tumor formation, facilitate immune escape and enable tumor progression. MDSC are important contributors to the development of an immunosuppressive tumor microenvironment that blocks the action of cytotoxic anti-tumor T effector cells. Heterogeneity in these cells poses a significant barrier to studying the in vivo contributions of individual MDSC subtypes. Herein, we demonstrate that granulocyte-macrophage colony stimulating factor (GM-CSF), a cytokine critical for the numeric and functional development of MDSC populations, promotes expansion of a monocyte-derived MDSC population characterized by expression of CD11b and the chemokine receptor CCR2. Utilizing a toxin mediated ablation strategy to target CCR2-expressing cells, we show that these monocytic MDSCs regulate entry of activated CD8 T cells into the tumor site, thereby limiting the efficacy of immunotherapy. Our results argue that therapeutic targeting of monocytic MDSCs would enhance outcomes in immunotherapy. PMID:22174368

  8. Barley-derived β-glucans increases gut permeability, ex vivo epithelial cell binding to E. coli, and naive T-cell proportions in weanling pigs.

    PubMed

    Ewaschuk, J B; Johnson, I R; Madsen, K L; Vasanthan, T; Ball, R; Field, C J

    2012-08-01

    Weaning in young animals is associated with an increased incidence of gastrointestinal infections. β-glucans exert numerous physiological effects, including altering immune function. The objective of this study was to determine the effects of feeding barley (Hordeum vulgare L.)-derived β-glucans on immune and intestinal function in weanling pigs (Sus scrofa). Thirty-one individually-housed Dutch Landrace pigs (21 d; initial BW, 6,298 ± 755 g) were weaned and fed a wheat-based diet (control) or a low (Lo-BG), medium (Med-BG), or high β-glucan-containing barley-based diet (Hi-BG) for 2 wk with 7 or 8 pigs/treatment. Intestinal segments were analyzed for permeability using Ussing chambers and K88 Escherichia coli adhesion to enterocytes was assessed ex vivo. Immune cells from mesenteric lymph nodes, peripheral blood, and Peyer's patches were analyzed for lymphocyte subsets by indirect immunofluorescence and the ability to respond ex vivo to mitogens by (3)H-thymidine incorporation. Hematology and neutrophil function were determined by flow cytometry. Neutrophil burst, size, and granularity, lymphocyte proliferation, and B-cell distribution in peripheral blood lymphocytes, Peyer's patches, and mesenteric lymph nodes were not affected by β-glucans content of the diet. The β-glucans content of the diet altered blood concentrations of erythrocytes and leukocytes, CD4, CD45RA, and CD8 blood cells (P < 0.05). In addition, feeding β-glucan resulted in increased (P < 0.05) percentage CD45RA positive cells in peripheral blood lymphocytes, Peyer's patches, and mesenteric lymph nodes. Mannitol permeability and tissue conductance were increased (P < 0.05) in Hi-BG fed pigs compared with control pigs. Percentage maximum K88-E.coli binding was increased in proportion to the β-glucan content of the diet (P < 0.05). Although β-glucan feeding during the weaning period increased blood lymphocytes and the proportion of naïve T-cells, it also increased E. coli

  9. Human endogenous retrovirus envelope proteins target dendritic cells to suppress T-cell activation.

    PubMed

    Hummel, Jonas; Kämmerer, Ulrike; Müller, Nora; Avota, Elita; Schneider-Schaulies, Sibylle

    2015-06-01

    Though mostly defective, human endogenous retroviruses (HERV) can retain open reading frames, which are especially expressed in the placenta. There, the envelope (env) proteins of HERV-W (Syncytin-1), HERV-FRD (Syncytin-2), and HERV-K (HML-2) were implicated in tolerance against the semi-allogenic fetus. Here, we show that the known HERV env-binding receptors ASCT-1 and -2 and MFSD2 are expressed by DCs and T-cells. When used as effectors in coculture systems, CHO cells transfected to express Syncytin-1, -2, or HML-2 did not affect T-cell expansion or overall LPS-driven phenotypic DC maturation, however, promoted release of IL-12 and TNF-α rather than IL-10. In contrast, HERV env expressing choriocarcinoma cell lines suppressed T-cell proliferation and LPS-induced TNF-α and IL-12 release, however, promoted IL-10 accumulation, indicating that these effects might not rely on HERV env interactions. However, DCs conditioned by choriocarcinoma, but also transgenic CHO cells failed to promote allogenic T-cell expansion. This was associated with a loss of DC/T-cell conjugate frequencies, impaired Ca(2+) mobilization, and aberrant patterning of f-actin and tyrosine phosphorylated proteins in T-cells. Altogether, these findings suggest that HERV env proteins target T-cell activation indirectly by modulating the stimulatory activity of DCs. PMID:25752285

  10. Exposure to Bisphenol A Prenatally or in Adulthood Promotes TH2 Cytokine Production Associated with Reduction of CD4+CD25+ Regulatory T Cells

    PubMed Central

    Yan, Huimin; Takamoto, Masaya; Sugane, Kazuo

    2008-01-01

    Background Bisphenol A (BPA) is a widespread endocrine-disrupting chemical that can affect humans and animals. Objectives We investigated the effects of adult or prenatal exposure to BPA on T-helper (TH)1/TH2 immune responses and the mechanisms underlying these effects. Methods To evaluate the effects of exposure to BPA in adulthood, male Leishmania major–susceptible BALB/c and –resistant C57BL/6 mice were subcutaneously injected with 0.625, 1.25, 2.5, and 5 μmol BPA 1 week before being infected with L. major. To evaluate prenatal exposure, female mice were given BPA-containing drinking water at concentrations of 1, 10, and 100 nM for 2 weeks, then mated, and given BPA for another week. Male 10-week-old offspring were infected with L. major. Footpad swelling was assessed as a measure of the course of infection. Results Mice exposed to BPA prenatally or in adulthood showed a dose-dependent increase in footpad swelling after being infected with L. major. Exposure to BPA in adulthood significantly promoted antigen-stimulated production of interleukin (IL)-4, IL-10, and IL-13 but not interferon-γ (IFN-γ). However, mice prenatally exposed to BPA showed increased production of not only IL-4 but also IFN-γ. The percentages of CD4+CD25+ cells were decreased in mice exposed to BPA either prenatally or in adulthood. Effects of prenatal BPA exposure were far more pronounced than effects of exposure in adulthood. Conclusion BPA promotes the development of TH2 cells in adulthood and both TH1 and TH2 cells in prenatal stages by reducing the number of regulatory T cells. PMID:18414636

  11. Multiple myeloma causes clonal T-cell immunosenescence: identification of potential novel targets for promoting tumour immunity and implications for checkpoint blockade.

    PubMed

    Suen, H; Brown, R; Yang, S; Weatherburn, C; Ho, P J; Woodland, N; Nassif, N; Barbaro, P; Bryant, C; Hart, D; Gibson, J; Joshua, D

    2016-08-01

    Tumour-induced dysfunction of cytotoxic T cells in patients with multiple myeloma (MM) may contribute to immune escape and be responsible for the lack of therapeutic efficacy of immune checkpoint blockade. We therefore investigated dysfunctional clonal T cells in MM and demonstrated immunosenescence but not exhaustion as a predominant feature. T-cell clones were detected in 75% of MM patients and their prognostic significance was revalidated in a new post-immunomodulatory drug cohort. The cells exhibited a senescent secretory effector phenotype: KLRG-1+/CD57+/CD160+/CD28-. Normal-for-age telomere lengths indicate that senescence is telomere independent and potentially reversible. p38-mitogen-activated protein kinase, p16 and p21 signalling pathways known to induce senescence were not elevated. Telomerase activity was found to be elevated and this may explain how normal telomere lengths are maintained in senescent cells. T-cell receptor signalling checkpoints were normal but elevated SMAD levels associated with T-cell inactivation were detected and may provide a potential target for the reversal of clonal T-cell dysfunction in MM. Low programmed death 1 and cytotoxic T-lymphocyte-associated antigen 4 expression detected on T-cell clones infers that these cells are not exhausted but suggests that there would be a suboptimal response to immune checkpoint blockade in MM. Our data suggest that other immunostimulatory strategies are required in MM. PMID:27102208

  12. T cell factor-4 functions as a co-activator to promote NF-κB-dependent MMP-15 expression in lung carcinoma cells.

    PubMed

    Liu, Yuliang; Xu, Yu; Guo, Shuliang; Chen, Hong

    2016-01-01

    Both TCF-4 and MMP-15 are closely linked to the development of lung cancer, while the regulatory role of TCF-4 in MMP-15 expression is still obscure. Here we found that expression of TCF-4 and MMP-15 was increased in lung cancer cells or tissues versus the normal ones. With gain-or loss-of -function studies, we demonstrated that TCF-4 positively regulated MMP-15 expression in mRNA and protein levels. With reporter gene assay, we found that TCF-4 regulated MMP-15 expression via a potential NF-κB binding element locating at -2833/-2824 in the mouse MMP-15 promoter. With ChIP and immunoblotting assays, we identified that TCF-4 functioned as a co-activator to potentiate the binding between p65 and MMP-15 promoter. Functionally, TCF-4 silence attenuated the migration activity of LLC cells, while additional overexpression of MMP-15 rescued this effect in cell scratch test and transwell migration assay. In xenograft model, TCF-4 silence-improved tumor lesions in lungs and survival time of LLC-tumor bearing mice were abolished by MMP-15 overexpression. In conclusion, we are the first to identify TCF-4 as a co-activator of NF-κB p65 to promote MMP-15 transcription and potentiate the migration activity of the lung cancer cells. Our findings shed light on the therapeutic strategies of this malignancy. PMID:27046058

  13. T cell factor-4 functions as a co-activator to promote NF-κB-dependent MMP-15 expression in lung carcinoma cells

    PubMed Central

    Liu, Yuliang; Xu, Yu; Guo, Shuliang; Chen, Hong

    2016-01-01

    Both TCF-4 and MMP-15 are closely linked to the development of lung cancer, while the regulatory role of TCF-4 in MMP-15 expression is still obscure. Here we found that expression of TCF-4 and MMP-15 was increased in lung cancer cells or tissues versus the normal ones. With gain-or loss-of -function studies, we demonstrated that TCF-4 positively regulated MMP-15 expression in mRNA and protein levels. With reporter gene assay, we found that TCF-4 regulated MMP-15 expression via a potential NF-κB binding element locating at -2833/-2824 in the mouse MMP-15 promoter. With ChIP and immunoblotting assays, we identified that TCF-4 functioned as a co-activator to potentiate the binding between p65 and MMP-15 promoter. Functionally, TCF-4 silence attenuated the migration activity of LLC cells, while additional overexpression of MMP-15 rescued this effect in cell scratch test and transwell migration assay. In xenograft model, TCF-4 silence-improved tumor lesions in lungs and survival time of LLC-tumor bearing mice were abolished by MMP-15 overexpression. In conclusion, we are the first to identify TCF-4 as a co-activator of NF-κB p65 to promote MMP-15 transcription and potentiate the migration activity of the lung cancer cells. Our findings shed light on the therapeutic strategies of this malignancy. PMID:27046058

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

    SciTech Connect

    Warford, Jordan; Doucette, Carolyn D.; Hoskin, David W.; Easton, Alexander S.

    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

  15. The Immunomodulator VacA Promotes Immune Tolerance and Persistent Helicobacter pylori Infection through Its Activities on T-Cells and Antigen-Presenting Cells

    PubMed Central

    Djekic, Aleksandra; Müller, Anne

    2016-01-01

    VacA is a pore-forming toxin that has long been known to induce vacuolization in gastric epithelial cells and to be linked to gastric disorders caused by H. pylori infection. Its role as a major colonization and persistence determinant of H. pylori is less well-understood. The purpose of this review is to discuss the various target cell types of VacA and its mechanism of action; specifically, we focus on the evidence showing that VacA targets myeloid cells and T-cells to directly and indirectly prevent H. pylori-specific T-cell responses and immune control of the infection. In particular, the ability of VacA-proficient H. pylori to skew T-cell responses towards regulatory T-cells and the effects of Tregs on H. pylori chronicity are highlighted. The by-stander effects of VacA-driven immunomodulation on extragastric diseases are discussed as well. PMID:27322319

  16. The Immunomodulator VacA Promotes Immune Tolerance and Persistent Helicobacter pylori Infection through Its Activities on T-Cells and Antigen-Presenting Cells.

    PubMed

    Djekic, Aleksandra; Müller, Anne

    2016-01-01

    VacA is a pore-forming toxin that has long been known to induce vacuolization in gastric epithelial cells and to be linked to gastric disorders caused by H. pylori infection. Its role as a major colonization and persistence determinant of H. pylori is less well-understood. The purpose of this review is to discuss the various target cell types of VacA and its mechanism of action; specifically, we focus on the evidence showing that VacA targets myeloid cells and T-cells to directly and indirectly prevent H. pylori-specific T-cell responses and immune control of the infection. In particular, the ability of VacA-proficient H. pylori to skew T-cell responses towards regulatory T-cells and the effects of Tregs on H. pylori chronicity are highlighted. The by-stander effects of VacA-driven immunomodulation on extragastric diseases are discussed as well. PMID:27322319

  17. Cell adhesion molecules involved in intrathymic T cell development.

    PubMed

    Patel, D D; Haynes, B F

    1993-08-01

    During stem cell migration to the thymus, intrathymic maturation of T cells, and emigration of mature T cells out of the thymus, intercellular interactions of developing T cells with a myriad of cell types are required for normal T cell development. Intercellular interactions of T cell precursors with endothelial cells, thymic epithelial cells, fibroblasts, thymic macrophages and dendritic cells are all mediated by adhesion molecules on immature T cells binding to ligands on thymic microenvironment cells. While many receptor-ligand interactions that are important in intrathymic T cell development are known, the adhesion molecules that are important for migration of T cell precursors to the thymus and for emigration of mature thymocytes from the thymus are poorly understood. An emerging concept is that select adhesion molecules at discrete stages of T cell maturation participate in and regulate the complex processes of T cell development. PMID:7693023

  18. Interferon-γ Promotes Inflammation and Development of T-Cell Lymphoma in HTLV-1 bZIP Factor Transgenic Mice

    PubMed Central

    Mitagami, Yu; Yasunaga, Jun-ichirou; Kinosada, Haruka; Ohshima, Koichi; Matsuoka, Masao

    2015-01-01

    Human T-cell leukemia virus type 1 (HTLV-1) is an etiological agent of several inflammatory diseases and a T-cell malignancy, adult T-cell leukemia (ATL). HTLV-1 bZIP factor (HBZ) is the only viral gene that is constitutively expressed in HTLV-1-infected cells, and it has multiple functions on T-cell signaling pathways. HBZ has important roles in HTLV-1-mediated pathogenesis, since HBZ transgenic (HBZ-Tg) mice develop systemic inflammation and T-cell lymphomas, which are similar phenotypes to HTLV-1-associated diseases. We showed previously that in HBZ-Tg mice, HBZ causes unstable Foxp3 expression, leading to an increase in regulatory T cells (Tregs) and the consequent induction of IFN-γ-producing cells, which in turn leads to the development of inflammation in the mice. In this study, we show that the severity of inflammation is correlated with the development of lymphomas in HBZ-Tg mice, suggesting that HBZ-mediated inflammation is closely linked to oncogenesis in CD4+ T cells. In addition, we found that IFN-γ-producing cells enhance HBZ-mediated inflammation, since knocking out IFN-γ significantly reduced the incidence of dermatitis as well as lymphoma. Recent studies show the critical roles of the intestinal microbiota in the development of Tregs in vivo. We found that even germ-free HBZ-Tg mice still had an increased number of Tregs and IFN-γ-producing cells, and developed dermatitis, indicating that an intrinsic activity of HBZ evokes aberrant T-cell differentiation and consequently causes inflammation. These results show that immunomodulation by HBZ is implicated in both inflammation and oncogenesis, and suggest a causal connection between HTLV-1-associated inflammation and ATL. PMID:26296091

  19. Prostaglandin E2 promotes survival of naive UCB T cells via the Wnt/β-catenin pathway and alters immune reconstitution after UCBT

    PubMed Central

    Li, L; Kim, H T; Nellore, A; Patsoukis, N; Petkova, V; McDonough, S; Politikos, I; Nikiforow, S; Soiffer, R; Antin, J H; Ballen, K; Cutler, C; Ritz, J; Boussiotis, V A

    2014-01-01

    The outcome of umbilical cord blood transplantation (UCBT) is compromised by low hematopoietic stem cell (HSC) doses leading to prolonged time to engraftment, delayed immunological reconstitution and late memory T-cell skewing. Exposure of UCB to dimethyl-prostaglandin E2 (dmPGE2) increases HSC in vivo. We determined that exposure of UCB T lymphocytes to dmPGE2 modified Wnt signaling resulting in T cell factor (TCF)-mediated transcription. Wnt signaling upregulated interleukin (IL)-7R and IL-2Rβ, resulting in enhanced survival mediated by the homeostatic cytokines IL-7 and IL-15. dmPGE2 also induced components of the Wnt pathway and Wnt receptors, thereby priming UCB T cells to receive signals via Wnt ligands in vivo. We observed that the Wnt transcription factor TCF7 and its target EOMES were elevated in the T cells of patients who received PGE2-treated UCBs. Consistent with the role of Wnt/β-catenin signaling to induce and maintain naive, memory precursors and long-lived central memory CD8+ cells, these patients also had increased fractions of CD8+CD45RO-CD62L+ plus CD8+CD45RO+CD62L+ subsets encompassing these T-cell populations. These effects of the PGE2/Wnt/β-catenin axis may have significant implications for harnessing immunity in the context of UCBT, where impaired immune reconstitution is associated with late memory T-cell skewing. PMID:24442207

  20. Antigenic epitopes fused to cationic peptide bound to oligonucleotides facilitate Toll-like receptor 9-dependent, but CD4+ T cell help-independent, priming of CD8+ T cells.

    PubMed

    Schirmbeck, Reinhold; Riedl, Petra; Zurbriggen, Rinaldo; Akira, Shizuo; Reimann, Jörg

    2003-11-15

    A priority in current vaccine research is the development of adjuvants that support the efficient priming of long-lasting, CD4(+) T cell help-independent CD8(+) T cell immunity. Oligodeoxynucleotides (ODN) with immune-stimulating sequences (ISS) containing CpG motifs facilitate the priming of MHC class I-restricted CD8(+) T cell responses to proteins or peptides. We show that the adjuvant effect of ISS(+) ODN on CD8(+) T cell priming to large, recombinant Ag is enhanced by binding them to short, cationic (arginine-rich) peptides that themselves have no adjuvant activity in CD8(+) T cell priming. Fusing antigenic epitopes to cationic (8- to 10-mer) peptides bound to immune-stimulating ISS(+) ODN or nonstimulating NSS(+) ODN (without CpG-containing sequences) generated immunogens that efficiently primed long-lasting, specific CD8(+) T cell immunity of high magnitude. Different MHC class I-binding epitopes fused to short cationic peptides of different origins showed this adjuvant activity. Quantitative ODN binding to cationic peptides strikingly reduced the toxicity of the latter, suggesting that it improves the safety profile of the adjuvant. CD8(+) T cell priming supported by this adjuvant was Toll-like receptor 9 dependent, but required no CD4(+) T cell help. ODN (with or without CpG-containing sequences) are thus potent Th1-promoting adjuvants when bound to cationic peptides covalently linked to antigenic epitopes, a mode of Ag delivery prevailing in many viral nucleocapsids. PMID:14607920

  1. miRNA-149* promotes cell proliferation and suppresses apoptosis by mediating JunB in T-cell acute lymphoblastic leukemia.

    PubMed

    Fan, Sheng-Jin; Li, Hui-Bo; Cui, Gang; Kong, Xiao-Lin; Sun, Li-Li; Zhao, Yan-Qiu; Li, Ying-Hua; Zhou, Jin

    2016-02-01

    MicroRNA-149* (miRNA-149*) functions as an oncogenic regulator in human melanoma. However, the effect of miRNA-149* on T-cell acute lymphoblastic leukemia (T-ALL) is unclear. Here we aimed to analyze the effects of miRNA-149* on in vitro T-ALL cells and to uncover the target for miRNA-149* in these cells. The miRNA-149* level was determined in multiple cell lines and bone marrow cells derived from patients with T-ALL, B acute lymphoblastic leukemia (B-ALL), acute myelocytic leukemia (AML), and healthy donors. We found that miRNA-149* was highly expressed in T-ALL cell lines and T-ALL patients' bone marrow samples. JunB was identified as a direct target of miR-149*. miRNA-149* mimics downregulated JunB levels in Molt-4 and Jurkat cells, while miRNA-149* inhibitors dramatically upregulated JunB expression in these cells. miRNA-149* mimics promoted proliferation, decreased the proportion of cells in G1 phase, and reduced cell apoptosis in T-ALL cells, while miRNA-149* inhibitors prevented these effects. miRNA-149* mimics downregulated p21 and upregulated cyclinD1, 4EBP1, and p70s6k in Molt-4 and Jurkat cells. Again, inhibitors prevented these effects. Our findings demonstrate that miRNA-149* may serve as an oncogenic regulator in T-ALL by negatively regulating JunB. PMID:26725775

  2. T Cells in Fish

    PubMed Central

    Nakanishi, Teruyuki; Shibasaki, Yasuhiro; Matsuura, Yuta

    2015-01-01

    Cartilaginous and bony fish are the most primitive vertebrates with a thymus, and possess T cells equivalent to those in mammals. There are a number of studies in fish demonstrating that the thymus is the essential organ for development of T lymphocytes from early thymocyte progenitors to functionally competent T cells. A high number of T cells in the intestine and gills has been reported in several fish species. Involvement of CD4+ and CD8α+ T cells in allograft rejection and graft-versus-host reaction (GVHR) has been demonstrated using monoclonal antibodies. Conservation of CD4+ helper T cell functions among teleost fishes has been suggested in a number studies employing mixed leukocyte culture (MLC) and hapten/carrier effect. Alloantigen- and virus-specific cytotoxicity has also been demonstrated in ginbuna and rainbow trout. Furthermore, the important role of cell-mediated immunity rather than humoral immunity has been reported in the protection against intracellular bacterial infection. Recently, the direct antibacterial activity of CD8α+, CD4+ T-cells and sIgM+ cells in fish has been reported. In this review, we summarize the recent progress in T cell research focusing on the tissue distribution and function of fish T cells. PMID:26426066

  3. Adenovirus Improves the Efficacy of Adoptive T-cell Therapy by Recruiting Immune Cells to and Promoting Their Activity at the Tumor.

    PubMed

    Tähtinen, Siri; Grönberg-Vähä-Koskela, Susanna; Lumen, Dave; Merisalo-Soikkeli, Maiju; Siurala, Mikko; Airaksinen, Anu J; Vähä-Koskela, Markus; Hemminki, Akseli

    2015-08-01

    Despite the rapid progress in the development of novel adoptive T-cell therapies, the clinical benefits in treatment of established tumors have remained modest. Several immune evasion mechanisms hinder T-cell entry into tumors and their activity within the tumor. Of note, oncolytic adenoviruses are intrinsically immunogenic due to inherent pathogen-associated molecular patterns. Here, we studied the capacity of adenovirus to overcome resistance of chicken ovalbumin-expressing B16.OVA murine melanoma tumors to adoptive ovalbumin-specific CD8(+) T-cell (OT-I) therapy. Following intraperitoneal transfer of polyclonally activated OT-I lymphocytes, control of tumor growth was superior in mice given intratumoral adenovirus compared with control mice, even in the absence of oncolytic virus replication. Preexisting antiviral immunity against serotype 5 did not hinder the therapeutic efficacy of the combination treatment. Intratumoral adenovirus injection was associated with an increase in proinflammatory cytokines, CD45(+) leukocytes, CD8(+) lymphocytes, and F4/80(+) macrophages, suggesting enhanced tumor immunogenicity. The proinflammatory effects of adenovirus on the tumor microenvironment led to expression of costimulatory signals on CD11c(+) antigen-presenting cells and subsequent activation of T cells, thus breaking the tumor-induced peripheral tolerance. An increased number of CD8(+) T cells specific for endogenous tumor antigens TRP-2 and gp100 was detected in combination-treated mice, indicating epitope spreading. Moreover, the majority of virus/T-cell-treated mice rejected the challenge of parental B16.F10 tumors, suggesting that systemic antitumor immunity was induced. In summary, we provide proof-of-mechanism data on combining adoptive T-cell therapy and adenovirotherapy for the treatment of cancer. PMID:25977260

  4. The phosphodiesterase 4 inhibitor roflumilast augments the Th17-promoting capability of dendritic cells by enhancing IL-23 production, and impairs their T cell stimulatory activity due to elevated IL-10.

    PubMed

    Bros, Matthias; Montermann, Evelyn; Cholaszczyńska, Anna; Reske-Kunz, Angelika B

    2016-06-01

    Phosphodiesterase 4 (PDE4) inhibitors serve to prevent degradation of the intracellular second messenger cAMP, resulting in broad anti-inflammatory effects on different cell types including immune cells. Agents that elevate cAMP levels via activation of adenylate cyclase have been shown to imprint a Th17-promoting capacity in dendritic cells (DCs). Therefore, we studied the potential of therapeutically relevant PDE inhibitors to induce a pronounced Th17-skewing capacity in DCs. Here we show that mouse bone marrow-derived (BM-) DCs when treated with the PDE4 inhibitor roflumilast (ROF, trade name: Daxas) in the course of stimulation with LPS (ROF-DCs) evoked elevated IL-17 levels in cocultured allogeneic T cells. In addition, as compared with control settings, levels of IFN-γ remained unaltered, while contents of Th2 cytokines (IL-5, IL-10) were diminished. ROF enhanced expression of the Th17-promoting factor IL-23 in BM-DCs. In line, neutralizing antibodies specific for IL-23 or IL-6 when applied to DC/T cell cocultures partially inhibited the IL17-promoting effect of ROF-DCs. Furthermore, ROF-DCs displayed a markedly diminished allogeneic T cell stimulatory capacity due to enhanced production of IL-10, which was restored upon application of IL-10 specific neutralizing antibody to DC/T cell cocultures. Both the IL-17-inducing and impaired T cell stimulatory capacity of BM-DCs were mimicked by a specific activator of protein kinase A, while stimulation of EPACs (exchange proteins of activated cAMP) did not yield such effects. Taken together, our findings suggest that PDE4 inhibitors aside from their broad overall anti-inflammatory effects may enhance the Th17-polarizing capacity in DCs as an unwanted side effect. PMID:27070502

  5. T cell subpopulations.

    PubMed

    Romagnani, Sergio

    2014-01-01

    The role of allergen-specific CD4+ effector type 2 helper (Th2) cells in the pathogenesis of allergic disorders is an established fact. Th2 cells produce interleukin (IL)-4 and IL-13, which induce immunoglobulin E production by B cells, and IL-5 that allows recruitment of eosinophils. Two main mechanisms control the Th2-mediated allergic inflammation: immune deviation (or Th1 redirection) and immune regulation. Regulatory T (Treg) cells exhibit a CD4+ phenotype and include Foxp3-positive thymic and induced Tregs, as well as Foxp3-negative IL-10-producing cells. Both immune deviation and immune regulation evoked by the maternal and newborn microbial environment probably operate in preventing allergen-specific Th2 responses. However, microbe-related protection from allergy seems to mainly depend on epigenetically controlled acetylation of the IFNG promoter of CD4+ T cells. Even Th17 and Th9 cells, as well as invariant NKT cells, have been implicated in the pathogenesis of allergic disorders, but their role is certainly more limited. Recently, innate lymphoid type 2 cells (ILC2) have been found to be able to produce high amounts of IL-5 and IL-13 in response to stimulation with IL-25 and IL-33 produced by non-immune cells. Together with Th2 cells, ILC2 may contribute to the induction and maintenance of allergic inflammation. PMID:24925396

  6. Identification and functional analysis of a second RBF-2 binding site within the HIV-1 promoter

    SciTech Connect

    Dahabieh, Matthew S.; Ooms, Marcel; Malcolm, Tom; Simon, Viviana; Sadowski, Ivan

    2011-09-15

    Transcription from the HIV-1 long terminal repeat (LTR) is mediated by numerous host transcription factors. In this study we characterized an E-box motif (RBE1) within the core promoter that was previously implicated in both transcriptional activation and repression. We show that RBE1 is a binding site for the RBF-2 transcription factor complex (USF1, USF2, and TFII-I), previously shown to bind an upstream viral element, RBE3. The RBE1 and RBE3 elements formed complexes of identical mobility and protein constituents in gel shift assays, both with Jurkat T-cell nuclear extracts and recombinant USF/TFII-I. Furthermore, both elements are regulators of HIV-1 expression; mutations in LTR-luciferase reporters and in HIV-1 molecular clones resulted in decreased transcription, virion production, and proviral expression in infected cells. Collectively, our data indicate that RBE1 is a bona fide RBF-2 binding site and that the RBE1 and RBE3 elements are necessary for mediating proper transcription from the HIV-1 LTR.

  7. A BTLA-mediated bait and switch strategy permits Listeria expansion in CD8α(+) DCs to promote long-term T cell responses.

    PubMed

    Yang, Xuanming; Zhang, Xunmin; Sun, Yonglian; Tu, Tony; Fu, May Lynne; Miller, Mendy; Fu, Yang-Xin

    2014-07-01

    Listeria monocytogenes infected CD8α(+) DCs in the spleen are essential for CD8(+) T cell generation. CD8α(+) DCs are also necessary for Listeria expansion and dissemination within the host. The mechanisms that regulate CD8α(+) DCs to allow Listeria expansion are unclear. We find that activating the B and T lymphocyte attenuator (BTLA), a coinhibitory receptor for T cells, suppresses, while blocking BTLA enhances, both the primary and memory CD8 T cell responses against Listeria. Btla(-/-) mice have lower effector and memory CD8(+) T cells while paradoxically also being more resistant to Listeria. Although bacterial entry into Btla(-/-) CD8α(+) DCs is unaffected, Listeria fails to expand within these cells. BTLA signaling limits Fas/FasL-mediated suppression of Listeria expansion within CD8α(+) DCs to more effectively alert adaptive immune cells. This study uncovers a BTLA-mediated strategy used by the host that permits Listeria proliferation to enable increasing T cell responses for long-term protection. PMID:25011109

  8. Dietary fish oil and DHA down-regulate antigen-activated CD4+ T-cells while promoting the formation of liquid-ordered mesodomains.

    PubMed

    Kim, Wooki; Barhoumi, Rola; McMurray, David N; Chapkin, Robert S

    2014-01-28

    We have demonstrated previously that n-3 PUFA endogenously produced by fat-1 transgenic mice regulate CD4+ T-cell function by affecting the formation of lipid rafts, liquid-ordered mesodomains in the plasma membrane. In the present study, we tested the effects of dietary sources of n-3 PUFA, i.e. fish oil (FO) or purified DHA, when compared with an n-6 PUFA-enriched maize oil control diet in DO11.10 T-cell receptor transgenic mice. Dietary n-3 PUFA were enriched in CD4+ T-cells, resulting in the increase of the n-3:n-6 ratio. Following antigen-specific CD4+ T-cell activation by B-lymphoma cells pulsed with the ovalbumin 323-339 peptide, the formation of liquid-ordered mesodomains at the immunological synapse relative to the whole CD4+ T-cell, as assessed by Laurdan labelling, was increased (P< 0·05) in the FO-fed group. The FO diet also suppressed (P< 0·05) the co-localisation of PKCθ with ganglioside GM1 (monosialotetrahexosylganglioside), a marker for lipid rafts, which is consistent with previous observations. In contrast, the DHA diet down-regulated (P< 0·05) PKCθ signalling by moderately affecting the membrane liquid order at the immunological synapse, suggesting the potential contribution of the other major n-3 PUFA components of FO, including EPA. PMID:23962659

  9. Inflammatory regulatory T cells in the microenvironments of ulcerative colitis and colon carcinoma.

    PubMed

    Kryczek, Ilona; Wang, Lin; Wu, Ke; Li, Wei; Zhao, Ende; Cui, Tracy; Wei, Shuang; Liu, Yan; Wang, Yin; Vatan, Linda; Szeliga, Wojciech; Greenson, Joel K; Roliński, Jacek; Zgodzinski, Witold; Huang, Emina; Tao, Kaixiong; Wang, Guobin; Zou, Weiping

    2016-08-01

    Foxp3(+)CD4(+) regulatory T (Treg) cells are thought to express negligible levels of effector cytokines, and inhibit immune responses and inflammation. Here, we have identified a population of IL-8(+)Foxp3(+)CD4(+) T cells in human peripheral blood, which is selectively increased in the microenvironments of ulcerative colitis and colon carcinoma. Phenotypically, this population is minimally overlapping with IL-17(+)Foxp3(+)CD4(+) T cells, and is different from IL-8(-)Foxp3(+)CD4(+) T cells in the same microenvironment. 40-60% of IL-8(+)Foxp3(+)CD4(+) T cells exhibit naive phenotype and express CD127, whereas IL-8(-)Foxp3(+)CD4(+) cells are basically memory T cells and express minimal CD127. The levels of CXCR5 expression are higher in IL-8(+)Foxp3(+) cells than in IL-8(-)Foxp3(+) cells. IL-2 and TGFβ induce IL-8(+)Foxp3(+) T cells. Exogenous Foxp3 expression promotes IL-8(+)Foxp3(+) T cells and inhibits effector cytokine IFNγ and IL-2 expression. Furthermore, Foxp3 binds to IL-8 proximal promoter and increases its activity. Functionally, IL-8(+)Foxp3(+) T cells inhibit T cell proliferation and effector cytokine production, but stimulate inflammatory cytokine production in the colon tissues, and promote neutrophil trafficking through IL-8. Thus, IL-8(+)Foxp3(+) cells may be an "inflammatory" Treg subset, and possess inflammatory and immunosuppressive dual biological activities. Given their dual roles and localization, these cells may be in a unique position to support tumor initiation and development in human chronic inflammatory environment. PMID:27622054

  10. Measurement of phenotype and absolute number of circulating heparin-binding hemagglutinin, ESAT-6 and CFP-10, and purified protein derivative antigen-specific CD4 T cells can discriminate active from latent tuberculosis infection.

    PubMed

    Hutchinson, Paul; Barkham, Timothy M S; Tang, Wenying; Kemeny, David M; Chee, Cynthia Bin-Eng; Wang, Yee T

    2015-02-01

    The tuberculin skin test (TST) and interferon gamma (IFN-γ) release assays (IGRAs) are used as adjunctive tests for the evaluation of suspected cases of active tuberculosis (TB). However, a positive test does not differentiate latent from active TB. We investigated whether flow cytometric measurement of novel combinations of intracellular cytokines and surface makers on CD4 T cells could differentiate between active and latent TB after stimulation with Mycobacterium tuberculosis-specific proteins. Blood samples from 60 patients referred to the Singapore Tuberculosis Control Unit for evaluation for active TB or as TB contacts were stimulated with purified protein derivative (PPD), ESAT-6 and CFP-10, or heparin-binding hemagglutinin (HBHA). The CD4 T cell cytokine response (IFN-γ, interleukin-2 [IL-2], interleukin-17A [IL-17A], interleukin-22 [IL-22], granulocyte-macrophage colony-stimulating factor [GM-CSF], and tumor necrosis factor alpha [TNF-α]) and surface marker expression (CD27, CXCR3, and CD154) were then measured. We found that the proportion of PPD-specific CD4 T cells, defined as CD154(+) TNF-α(+) cells that were negative for CD27 and positive for GM-CSF, gave the strongest discrimination between subjects with latent and those with active TB (area under the receiver operator characteristic [ROC] curve of 0.9277; P < 0.0001). Also, the proportions and absolute numbers of HBHA-specific CD4 T cells were significantly higher in those with latent TB infection, particularly CD154(+) TNF-α(+) IFN-γ(+) IL-2(+) and CD154(+) TNF-α(+) CXCR3(+). Finally, we found that the ratio of ESAT-6- and CFP-10-responding to HBHA-responding CD4 T cells was significantly different between the two study populations. In conclusion, we found novel markers of M. tuberculosis-specific CD4 cells which differentiate between active and latent TB. PMID:25520147

  11. T cell recognition of beryllium.

    PubMed

    Dai, Shaodong; Falta, Michael T; Bowerman, Natalie A; McKee, Amy S; Fontenot, Andrew P

    2013-12-01

    Chronic beryllium disease (CBD) is a granulomatous lung disorder caused by a hypersensitivity to beryllium and characterized by the accumulation of beryllium-specific CD4(+) T cells in the lung. Genetic susceptibility to beryllium-induced disease is strongly associated with HLA-DP alleles possessing a glutamic acid at the 69th position of the β-chain (βGlu69). The structure of HLA-DP2, the most prevalent βGlu69-containing molecule, revealed a unique solvent-exposed acidic pocket that includes βGlu69 and represents the putative beryllium-binding site. The delineation of mimotopes and endogenous self-peptides that complete the αβTCR ligand for beryllium-specific CD4(+) T cells suggests a unique role of these peptides in metal ion coordination and the generation of altered self-peptides, blurring the distinction between hypersensitivity and autoimmunity. PMID:23978481

  12. Kinetic discrimination in T-cell activation.

    PubMed Central

    Rabinowitz, J D; Beeson, C; Lyons, D S; Davis, M M; McConnell, H M

    1996-01-01

    We propose a quantitative model for T-cell activation in which the rate of dissociation of ligand from T-cell receptors determines the agonist and antagonist properties of the ligand. The ligands are molecular complexes between antigenic peptides and proteins of the major histocompatibility complex on the surfaces of antigen-presenting cells. Binding of ligand to receptor triggers a series of biochemical reactions in the T cell. If the ligand dissociates after these reactions are complete, the T cell receives a positive activation signal. However, dissociation of ligand after completion of the first reaction but prior to generation of the final products results in partial T-cell activation, which acts to suppress a positive response. Such a negative signal is brought about by T-cell ligands containing the variants of antigenic peptides referred to as T-cell receptor antagonists. Results of recent experiments with altered peptide ligands compare favorably with T-cell responses predicted by this model. PMID:8643643

  13. T-Cell Lymphoma

    MedlinePlus

    ... are extremely rare. T-cell lymphomas can be aggressive (fast-growing) or indolent (slow-growing). Lymphomas are ... also be involved. This group of PTCLs is aggressive and requires combination chemotherapy upon diagnosis. For more ...

  14. VLA-4 blockade promotes differential routes into human CNS involving PSGL-1 rolling of T cells and MCAM-adhesion of TH17 cells

    PubMed Central

    Schneider-Hohendorf, Tilman; Rossaint, Jan; Mohan, Hema; Böning, Daniel; Breuer, Johanna; Kuhlmann, Tanja; Gross, Catharina C.; Flanagan, Ken; Sorokin, Lydia; Vestweber, Dietmar; Zarbock, Alexander

    2014-01-01

    The focus of this study is the characterization of human T cell blood–brain barrier migration and corresponding molecular trafficking signatures. We examined peripheral blood and cerebrospinal fluid immune cells from patients under long-term anti–very late antigen-4 (VLA-4)/natalizumab therapy (LTNT) and from CNS specimens. LTNT patients’ cerebrospinal fluid T cells exhibited healthy central-/effector-memory ratios, but lacked CD49d and showed enhanced myeloma cell adhesion molecule (MCAM) expression. LTNT led to an increase of PSGL-1 expression on peripheral T cells. Although vascular cell adhesion molecule-1 (VLA-4 receptor) was expressed at all CNS barriers, P-selectin (PSGL-1-receptor) was mainly detected at the choroid plexus. Accordingly, in vitro experiments under physiological flow conditions using primary human endothelial cells and LTNT patients’ T cells showed increased PSGL-1–mediated rolling and residual adhesion, even under VLA-4 blockade. Adhesion of MCAM+/TH17 cells was not affected by VLA-4 blocking alone, but was abrogated when both VLA-4 and MCAM were inhibited. Consistent with these data, MCAM+ cells were detected in white matter lesions, and in gray matter of multiple sclerosis patients. Our data indicate that lymphocyte trafficking into the CNS under VLA-4 blockade can occur by using the alternative adhesion molecules, PSGL-1 and MCAM, the latter representing an exclusive pathway for TH17 cells to migrate over the blood–brain barrier. PMID:25135296

  15. TALEN-Mediated Inactivation of PD-1 in Tumor-Reactive Lymphocytes Promotes Intratumoral T-cell Persistence and Rejection of Established Tumors.

    PubMed

    Menger, Laurie; Sledzinska, Anna; Bergerhoff, Katharina; Vargas, Frederick Arce; Smith, Julianne; Poirot, Laurent; Pule, Martin; Hererro, Javier; Peggs, Karl S; Quezada, Sergio A

    2016-04-15

    Despite the promising efficacy of adoptive cell therapies (ACT) in melanoma, complete response rates remain relatively low and outcomes in other cancers are less impressive. The immunosuppressive nature of the tumor microenvironment and the expression of immune-inhibitory ligands, such as PD-L1/CD274 by the tumor and stroma are considered key factors limiting efficacy. The addition of checkpoint inhibitors (CPI) to ACT protocols bypasses some mechanisms of immunosuppression, but associated toxicities remain a significant concern. To overcome PD-L1-mediated immunosuppression and reduce CPI-associated toxicities, we used TALEN technology to render tumor-reactive T cells resistant to PD-1 signaling. Here, we demonstrate that inactivation of the PD-1 gene in melanoma-reactive CD8(+) T cells and in fibrosarcoma-reactive polyclonal T cells enhanced the persistence of PD-1 gene-modified T cells at the tumor site and increased tumor control. These results illustrate the feasibility and potency of approaches incorporating advanced gene-editing technologies into ACT protocols to silence immune checkpoints as a strategy to overcome locally active immune escape pathways. Cancer Res; 76(8); 2087-93. ©2016 AACR. PMID:27197251

  16. Activation of Flucloxacillin-Specific CD8+ T-Cells With the Potential to Promote Hepatocyte Cytotoxicity in a Mouse Model.

    PubMed

    Nattrass, Ryan; Faulkner, Lee; Vocanson, Marc; Antoine, Daniel J; Kipar, Anja; Kenna, Gerry; Nicolas, Jean-Francois; Park, B Kevin; Naisbitt, Dean J

    2015-07-01

    There are currently no animal models of drug-induced liver injury (DILI) where the adaptive immune system has been shown to damage the liver. Thus, it is difficult to explore the mechanistic basis of the tissue injury. The aim of this study was to use C57BL/6 CD4+-deficient mice with a mutation in the αβ gene encoding for Major histocompatibilty complex (MHC) class II molecules to (1) develop a mouse model of flucloxacillin sensitization, (2) explore whether drug-specific CD8+ kill primary hepatocytes, and (3) analyze perturbations in liver integrity following oral exposure to flucloxacillin. CD8+ T-cells from lymph nodes of flucloxacillin-sensitized mice were stimulated to proliferate, secrete interferon (IFN-γ) and granzyme B, and induce hepatocyte apoptosis in a concentration-dependent manner following ex vivo stimulation. The T-cell response was antigen-specific; T-cells were not activated with other β-lactam antibiotics. Furthermore, T-cell responses only occurred in the presence of flucloxacillin-pulsed antigen presenting cells. In separate experiments, flucloxacillin-specific T-cells were induced to migrate to the mesenteric lymph nodes using retinoic acid, prior to administration of oral flucloxacillin, and analysis of plasma biomarkers of liver injury. Oral exposure to flucloxacillin resulted in mild elevations in alanine aminotransferase, liver, and gall bladder leukocyte infiltration and a marked swelling of the gall bladder. Thus, CD4+-deficient mice represent a promising model to study the role of the adaptive immune system in DILI. PMID:25877613

  17. Vaccines targeting tumor blood vessel antigens promote CD8(+) T cell-dependent tumor eradication or dormancy in HLA-A2 transgenic mice.

    PubMed

    Zhao, Xi; Bose, Anamika; Komita, Hideo; Taylor, Jennifer L; Chi, Nina; Lowe, Devin B; Okada, Hideho; Cao, Ying; Mukhopadhyay, Debabrata; Cohen, Peter A; Storkus, Walter J

    2012-02-15

    We have recently shown that effective cytokine gene therapy of solid tumors in HLA-A2 transgenic (HHD) mice lacking murine MHC class I molecule expression results in the generation of HLA-A2-restricted CD8(+) T effector cells selectively recognizing tumor blood vessel-associated pericytes and/or vascular endothelial cells. Using an HHD model in which HLA-A2(neg) tumor (MC38 colon carcinoma or B16 melanoma) cells are not recognized by the CD8(+) T cell repertoire, we now show that vaccines on the basis of tumor-associated blood vessel Ags (TBVA) elicit protective Tc1-dependent immunity capable of mediating tumor regression or extending overall survival. Vaccine efficacy was not observed if (HLA-A2(neg)) wild-type C57BL/6 mice were instead used as recipient animals. In the HHD model, effective vaccination resulted in profound infiltration of tumor lesions by CD8(+) (but not CD4(+)) T cells, in a coordinate reduction of CD31(+) blood vessels in the tumor microenvironment, and in the "spreading" of CD8(+) T cell responses to alternate TBVA that were not intrinsic to the vaccine. Protective Tc1-mediated immunity was durable and directly recognized pericytes and/or vascular endothelial cells flow-sorted from tumor tissue but not from tumor-uninvolved normal kidneys harvested from these same animals. Strikingly, the depletion of CD8(+), but not CD4(+), T cells at late time points after effective therapy frequently resulted in the recurrence of disease at the site of the regressed primary lesion. This suggests that the vaccine-induced anti-TBVA T cell repertoire can mediate the clinically preferred outcomes of either effectively eradicating tumors or policing a state of (occult) tumor dormancy. PMID:22246626

  18. Identification of novel Notch target genes in T cell leukaemia

    PubMed Central

    Chadwick, Nicholas; Zeef, Leo; Portillo, Virginia; Fennessy, Carl; Warrander, Fiona; Hoyle, Sarah; Buckle, Anne-Marie

    2009-01-01

    Background Dysregulated Notch signalling is believed to play an important role in the development and maintenance of T cell leukaemia. At a cellular level, Notch signalling promotes proliferation and inhibits apoptosis of T cell acute lymphoblastic leukaemia (T-ALL) cells. In this study we aimed to identify novel transcriptional targets of Notch signalling in the T-ALL cell line, Jurkat. Results RNA was prepared from Jurkat cells retrovirally transduced with an empty vector (GFP-alone) or vectors containing constitutively active forms of Notch (N1ΔE or N3ΔE), and used for Affymetrix microarray analysis. A subset of genes found to be regulated by Notch was chosen for real-time PCR validation and in some cases, validation at the protein level, using several Notch-transduced T-ALL and non-T-ALL leukaemic cell lines. As expected, several known transcriptional target of Notch, such as HES1 and Deltex, were found to be overexpressed in Notch-transduced cells, however, many novel transcriptional targets of Notch signalling were identified using this approach. These included the T cell costimulatory molecule CD28, the anti-apoptotic protein GIMAP5, and inhibitor of DNA binding 1 (1D1). Conclusion The identification of such downstream Notch target genes provides insights into the mechanisms of Notch function in T cell leukaemia, and may help identify novel therapeutic targets in this disease. PMID:19508709

  19. P2X7 on Mouse T Cells: One Channel, Many Functions

    PubMed Central

    Rissiek, Björn; Haag, Friedrich; Boyer, Olivier; Koch-Nolte, Friedrich; Adriouch, Sahil

    2015-01-01

    The P2X7 receptor is an adenosine triphosphate (ATP)-gated cation channel that is expressed by several cells of the immune system. P2X7 is best known for its proinflammatory role in promoting inflammasome formation and release of mature interleukin (IL)-1β by innate immune cells. Mounting evidence indicates that P2X7 is also an important regulatory receptor of murine and human T cell functions. Murine T cells express a sensitive splice variant of P2X7 that can be activated either by non-covalent binding of ATP or, in the presence of nicotinamide adenine dinucleotide, by its covalent ADP-ribosylation catalyzed by the ecto-ADP-ribosyltransferase ARTC2.2. Prolonged activation of P2X7 by either one of these pathways triggers the induction of T cell death. Conversely, lower concentrations of ATP can activate P2X7 to enhance T cell proliferation and production of IL-2. In this review, we will highlight the molecular and cellular consequences of P2X7 activation on mouse T cells and its versatile role in T cell homeostasis and activation. Further, we will discuss important differences in the function of P2X7 on human and murine T cells. PMID:26042119

  20. Phosphorylation of a Tyrosine Residue on Zap70 by Lck and Its Subsequent Binding via an SH2 Domain May Be a Key Gatekeeper of T Cell Receptor Signaling In Vivo.

    PubMed

    Thill, Peter A; Weiss, Arthur; Chakraborty, Arup K

    2016-09-15

    The initiation of signaling in T lymphocytes in response to the binding of the T cell receptor (TCR) to cognate ligands is a key step in the emergence of adaptive immune responses. Conventional models posit that TCR signaling is initiated by the phosphorylation of receptor-associated immune receptor activation motifs (ITAMs). The cytoplasmic tyrosine kinase Zap70 binds to phosphorylated ITAMs, is subsequently activated, and then propagates downstream signaling. While evidence for such models is provided by experiments with cell lines, in vivo, Zap70 is bound to phosphorylated ITAMs in resting T cells. However, Zap70 is activated only upon TCR binding to cognate ligand. We report the results of computational studies of a new model for the initiation of TCR signaling that incorporates these in vivo observations. Importantly, the new model is shown to allow better and faster TCR discrimination between self-ligands and foreign ligands. The new model is consistent with many past experimental observations, and experiments that could further test the model are proposed. PMID:27354065

  1. Cannabinoid receptor-2 (CB2) agonist ameliorates colitis in IL-10{sup −/−} mice by attenuating the activation of T cells and promoting their apoptosis

    SciTech Connect

    Singh, Udai P.; Singh, Narendra P.; Singh, Balwan; Price, Robert L.; Nagarkatti, Mitzi; Nagarkatti, Prakash S.

    2012-01-15

    Inflammatory bowel disease (IBD) is a chronic intestinal inflammation caused by hyperactivated effector immune cells that produce pro-inflammatory cytokines. Recent studies have shown that the cannabinoid system may play a critical role in mediating protection against intestinal inflammation. However, the effect of cannabinoid receptor induction after chronic colitis progression has not been investigated. Here, we investigate the effect of cannabinoid receptor-2 (CB2) agonist, JWH-133, after chronic colitis in IL-10{sup −/−} mice. JWH-133 effectively attenuated the overall clinical score, and reversed colitis-associated pathogenesis and decrease in body weight in IL-10{sup −/−} mice. After JWH-133 treatment, the percentage of CD4{sup +} T cells, neutrophils, mast cells, natural killer (NK1.1) cells, and activated T cells declined in the intestinal lamina propria (LP) and mesenteric lymph nodes (MLN) of mice with chronic colitis. JWH-133 was also effective in ameliorating dextran sodium sulfate (DSS)-induced colitis. In this model, JWH-133 reduced the number and percentage of macrophages and IFN-γ expressing cells that were induced during colitis progression. Treatment with aminoalkylindole 6-iodo-pravadoline (AM630), a CB2 receptor antagonist, reversed the colitis protection provided by JWH-133 treatment. Also, activated T cells were found to undergo apoptosis following JWH-133 treatment both in-vivo and in-vitro. These findings suggest that JWH-133 mediates its effect through CB2 receptors, and ameliorates chronic colitis by inducing apoptosis in activated T cells, reducing the numbers of activated T cells, and suppressing induction of mast cells, NK cells, and neutrophils at sites of inflammation in the LP. These results support the idea that the CB2 receptor agonists may serve as a therapeutic modality against IBD. -- Highlights: ► JWH-133, a cannnabinoid receptor-2 agonist ameliorates experimental colitis. ► JWH-133 suppressed inflammation and

  2. IL-10 promotes homeostatic proliferation of human CD8(+) memory T cells and, when produced by CD1c(+) DCs, shapes naive CD8(+) T-cell priming.

    PubMed

    Nizzoli, Giulia; Larghi, Paola; Paroni, Moira; Crosti, Maria Cristina; Moro, Monica; Neddermann, Petra; Caprioli, Flavio; Pagani, Massimiliano; De Francesco, Raffaele; Abrignani, Sergio; Geginat, Jens

    2016-07-01

    IL-10 is an anti-inflammatory cytokine that inhibits maturation and cytokine production of dendritic cells (DCs). Although mature DCs have the unique capacity to prime CD8(+) CTL, IL-10 can promote CTL responses. To understand these paradoxic findings, we analyzed the role of IL-10 produced by human APC subsets in T-cell responses. IL-10 production was restricted to CD1c(+) DCs and CD14(+) monocytes. Interestingly, it was differentially regulated, since R848 induced IL-10 in DCs, but inhibited IL-10 in monocytes. Autocrine IL-10 had only a weak inhibitory effect on DC maturation, cytokine production, and CTL priming with high-affinity peptides. Nevertheless, it completely blocked cross-priming and priming with low-affinity peptides of a self/tumor-antigen. IL-10 also inhibited CD1c(+) DC-induced CD4(+) T-cell priming and enhanced Foxp3 induction, but was insufficient to induce T-cell IL-10 production. CD1c(+) DC-derived IL-10 had also no effect on DC-induced secondary expansions of memory CTL. However, IL-15-driven, TCR-independent proliferation of memory CTL was enhanced by IL-10. We conclude that DC-derived IL-10 selects high-affinity CTL upon priming. Moreover, IL-10 preserves established CTL memory by enhancing IL-15-dependent homeostatic proliferation. These combined effects on CTL priming and memory maintenance provide a plausible mechanism how IL-10 promotes CTL responses in humans. PMID:27129615

  3. Memory T Cell Migration

    PubMed Central

    Zhang, Qianqian; Lakkis, Fadi G.

    2015-01-01

    Immunological memory is a key feature of adaptive immunity. It provides the organism with long-lived and robust protection against infection. In organ transplantation, memory T cells pose a significant threat by causing allograft rejection that is generally resistant to immunosuppressive therapy. Therefore, a more thorough understanding of memory T cell biology is needed to improve the survival of transplanted organs without compromising the host’s ability to fight infections. This review will focus on the mechanisms by which memory T cells migrate to the site where their target antigen is present, with particular emphasis on their migration to transplanted organs. First, we will define the known subsets of memory T cells (central, effector, and tissue resident) and their circulation patterns. Second, we will review the cellular and molecular mechanisms by which memory T cells migrate to inflamed and non-inflamed tissues and highlight the emerging paradigm of antigen-driven, trans-endothelial migration. Third, we will discuss the relevance of this knowledge to organ transplantation and the prevention or treatment of allograft rejection. PMID:26483794

  4. Activation of CD1d-restricted natural killer T cells can inhibit cancer cell proliferation during chemotherapy by promoting the immune responses in murine mesothelioma.

    PubMed

    Wu, Licun; Yun, Zhihong; Tagawa, Tetsuzo; De la Maza, Luis; Wu, Matthew Onn; Yu, Julie; Zhao, Yidan; de Perrot, Marc

    2014-12-01

    We studied the impact of natural killer T (NKT) cell activation by alpha-galactocysylceramide (α-GalCer, α-GC) on cancer cell repopulation during chemotherapy in murine mesothelioma. The number of NKT cells was found to be increased during the development of murine mesothelioma. NKT cells specifically recognize α-GC through CD1d resulting in their activation and expansion. Tumor-bearing mice were treated with chemotherapy once weekly, and α-GC was followed after each cycle of chemotherapy. Anti-tumor effect was evaluated on wild-type (WT) and CD1d knockout (CD1dKO) mice. Cancer cell proliferation and apoptosis were evaluated by Ki67 and TUNEL immunohistochemistry. CD4(+) and CD8(+) T cell proportion and activation in tumor, spleen, draining lymph node and peripheral blood were determined by flow cytometry, and gene expression of activated T cell-related cytokines was quantified by reverse transcription PCR. NKT cells were identified by CD1d-α-GC-tetramer staining. In WT mice, tumor growth delay was achieved by cisplatin (Cis), and this effect was improved in combination with α-GC, but α-GC alone had little effect. Cancer cell proliferation during chemotherapy was significantly inhibited by α-GC, while cancer cell death was significantly upregulated. α-GC following chemotherapy resulted in NKT cell expansion and an increase of interferon-γ production in the draining lymph node, blood and spleen. Gene expression of immune-associated cytokines was upregulated. Strikingly, the percentage of inducible T cell co-stimulator(+)CD4 T cells, Th17/Tc17 cells increased in splenocytes. In CD1d KO mice, however, Cis alone was less effective and Cis + α-GC provided no additional benefit over Cis alone. α-GC alone had minimal effect in both mice. NKT activation between cycles of chemotherapy could improve the outcome of mesothelioma treatment. PMID:25183171

  5. IL-15 Trans-Signaling with the Superagonist RLI Promotes Effector/Memory CD8+ T Cell Responses and Enhances Antitumor Activity of PD-1 Antagonists.

    PubMed

    Desbois, Mélanie; Le Vu, Pauline; Coutzac, Clélia; Marcheteau, Elie; Béal, Coralie; Terme, Magali; Gey, Alain; Morisseau, Sébastien; Teppaz, Géraldine; Boselli, Lisa; Jacques, Yannick; Béchard, David; Tartour, Eric; Cassard, Lydie; Chaput, Nathalie

    2016-07-01

    Tumors with the help of the surrounding environment facilitate the immune suppression in patients, and immunotherapy can counteract this inhibition. Among immunotherapeutic strategies, the immunostimulatory cytokine IL-15 could represent a serious candidate for the reactivation of antitumor immunity. However, exogenous IL-15 may have a limited impact on patients with cancer due to its dependency on IL-15Rα frequently downregulated in cancer patients. In this work, we studied the antitumor activity of the IL-15 superagonist receptor-linker-IL-15 (RLI), designed to bypass the need of endogenous IL-15Rα. RLI consists of human IL-15 covalently linked to the human IL-15Rα sushi(+) domain. In a mouse model of colorectal carcinoma, RLI as a stand-alone treatment could limit tumor outgrowth only when initiated at an early time of tumor development. At a later time, RLI was not effective, coinciding with the strong accumulation of terminally exhausted programmed cell death-1 (PD-1)(high) T cell Ig mucin-3(+) CD8(+) T cells, suggesting that RLI was not able to reactivate terminally exhausted CD8(+) T cells. Combination with PD-1 blocking Ab showed synergistic activity with RLI, but not with IL-15. RLI could induce a greater accumulation of memory CD8(+) T cells and a stronger effector function in comparison with IL-15. Ex vivo stimulation of tumor-infiltrated lymphocytes from 16 patients with renal cell carcinoma demonstrated 56% of a strong tumor-infiltrated lymphocyte reactivation with the combination anti-PD-1/RLI compared with 43 and 6% with RLI or anti-PD-1, respectively. Altogether, this work provides evidence that the sushi-IL-15Rα/IL-15 fusion protein RLI enhances antitumor activity of anti-PD-1 treatment and is a promising approach to stimulate host immunity. PMID:27217584

  6. Virus-specific antibodies allow viral replication in the marginal zone, thereby promoting CD8(+) T-cell priming and viral control.

    PubMed

    Duhan, Vikas; Khairnar, Vishal; Friedrich, Sarah-Kim; Zhou, Fan; Gassa, Asmae; Honke, Nadine; Shaabani, Namir; Gailus, Nicole; Botezatu, Lacramioara; Khandanpour, Cyrus; Dittmer, Ulf; Häussinger, Dieter; Recher, Mike; Hardt, Cornelia; Lang, Philipp A; Lang, Karl S

    2016-01-01

    Clinically used human vaccination aims to induce specific antibodies that can guarantee long-term protection against a pathogen. The reasons that other immune components often fail to induce protective immunity are still debated. Recently we found that enforced viral replication in secondary lymphoid organs is essential for immune activation. In this study we used the lymphocytic choriomeningitis virus (LCMV) to determine whether enforced virus replication occurs in the presence of virus-specific antibodies or virus-specific CD8(+) T cells. We found that after systemic recall infection with LCMV-WE the presence of virus-specific antibodies allowed intracellular replication of virus in the marginal zone of spleen. In contrast, specific antibodies limited viral replication in liver, lung, and kidney. Upon recall infection with the persistent virus strain LCMV-Docile, viral replication in spleen was essential for the priming of CD8(+) T cells and for viral control. In contrast to specific antibodies, memory CD8(+) T cells inhibited viral replication in marginal zone but failed to protect mice from persistent viral infection. We conclude that virus-specific antibodies limit viral infection in peripheral organs but still allow replication of LCMV in the marginal zone, a mechanism that allows immune boosting during recall infection and thereby guarantees control of persistent virus. PMID:26805453

  7. Virus-specific antibodies allow viral replication in the marginal zone, thereby promoting CD8+ T-cell priming and viral control

    PubMed Central

    Duhan, Vikas; Khairnar, Vishal; Friedrich, Sarah-Kim; Zhou, Fan; Gassa, Asmae; Honke, Nadine; Shaabani, Namir; Gailus, Nicole; Botezatu, Lacramioara; Khandanpour, Cyrus; Dittmer, Ulf; Häussinger, Dieter; Recher, Mike; Hardt, Cornelia; Lang, Philipp A.; Lang, Karl S.

    2016-01-01

    Clinically used human vaccination aims to induce specific antibodies that can guarantee long-term protection against a pathogen. The reasons that other immune components often fail to induce protective immunity are still debated. Recently we found that enforced viral replication in secondary lymphoid organs is essential for immune activation. In this study we used the lymphocytic choriomeningitis virus (LCMV) to determine whether enforced virus replication occurs in the presence of virus-specific antibodies or virus-specific CD8+ T cells. We found that after systemic recall infection with LCMV-WE the presence of virus-specific antibodies allowed intracellular replication of virus in the marginal zone of spleen. In contrast, specific antibodies limited viral replication in liver, lung, and kidney. Upon recall infection with the persistent virus strain LCMV-Docile, viral replication in spleen was essential for the priming of CD8+ T cells and for viral control. In contrast to specific antibodies, memory CD8+ T cells inhibited viral replication in marginal zone but failed to protect mice from persistent viral infection. We conclude that virus-specific antibodies limit viral infection in peripheral organs but still allow replication of LCMV in the marginal zone, a mechanism that allows immune boosting during recall infection and thereby guarantees control of persistent virus. PMID:26805453

  8. Pseudomonas aeruginosa mannose-sensitive hemagglutinin promotes T-cell response via toll-like receptor 4-mediated dendritic cells to slow tumor progression in mice.

    PubMed

    Zhang, Min; Luo, Feifei; Zhang, Yufei; Wang, Luman; Lin, Wei; Yang, Mengxuan; Hu, Dali; Wu, Xiaofeng; Chu, Yiwei

    2014-05-01

    Pseudomonas aeruginosa-mannose-sensitive hemagglutinin (PA-MSHA) as a drug may kill tumor cells and has been used clinically. However, the antitumor immune response of PA-MSHA is not completely understood. In this study, we found that treating Lewis lung carcinoma (3LL)-bearing mice with PA-MSHA plus 3LL antigen led to slower tumor progression and longer survival. After PA-MSHA treatment, T-cell number and dendritic cell maturation were both increased significantly at the tumor site. In addition, PA-MSHA in vitro stimulation resulted in the maturation of bone marrow-derived dendritic cells (BMDCs) from naive mice, showing higher costimulatory molecule expression, more cytokine secretion, lower endocytic activity, and stronger capacity to enhance T-cell activation. Toll-like receptor (TLR)4 but not TLR2 was required in the maturation process. More importantly, PA-MSHA-induced DCs were essential for PA-MSHA to enhance activation, expansion, and interferon (IFN)-γ secretion of TLR4-mediated T cells, which play a role in the antitumor effect of PA-MSHA. Thus, this study reveals PA-MSHA as a novel TLR4 agonist that elicits antitumor immune response to slow tumor progression. PMID:24623801

  9. IL-27 promotes IL-10 production by effector Th1 CD4+ T cells; a critical mechanism for protection from severe immunopathology during malaria infection1

    PubMed Central

    Freitas do Rosário, Ana Paula; Lamb, Tracey; Spence, Philip; Stephens, Robin; Lang, Agathe; Roers, Axel; Muller, Werner; O’Garra, Anne; Langhorne, Jean

    2012-01-01

    Infection with the malaria parasite, Plasmodium, is characterized by excessive inflammation. The establishment of a precise balance between the pro- and anti-inflammatory responses is critical to guarantee control of the parasite and survival of the host. Interleukin (IL)-10, a key regulatory cytokine produced by many cells of the immune system, has been shown to protect mice against pathology during acute Plasmodium chabaudi chabaudi AS model of malaria. However, the critical cellular source of IL-10 is still unknown. Here, we demonstrate that T cell-derived IL-10 is necessary for the control of pathology during acute malaria, as mice bearing specific deletion of Il10 in T cells fully reproduce the phenotype observed in Il10−/− mice, with significant weight loss, drop in temperature and increased mortality. Furthermore, we show that IFN-γ+ Th1 cells are the main producers of IL-10 throughout acute infection, expressing high levels of CD44 and ICOS and low levels of CD127. Although Foxp3+ regulatory CD4+ T cells produce IL-10 during infection, highly activated IFN-γ+ Th1 cells were shown to be the essential and sufficient source of IL-10 to guarantee protection against severe immune-mediated pathology. Finally, in this model of malaria we demonstrate that the generation of protective IL10+IFN-γ+ Th1 cells is dependent on IL-27 signaling, and independent of IL-21. PMID:22205023

  10. Langerin negative dendritic cells promote potent CD8+ T-cell priming by skin delivery of live adenovirus vaccine microneedle arrays

    PubMed Central

    Bachy, Veronique; Hervouet, Catherine; Becker, Pablo D.; Chorro, Laurent; Carlin, Leo M.; Herath, Shanthi; Papagatsias, Timos; Barbaroux, Jean-Baptiste; Oh, Sea-Jin; Benlahrech, Adel; Athanasopoulos, Takis; Dickson, George; Patterson, Steven; Kwon, Sung-Yun; Geissmann, Frederic; Klavinskis, Linda S.

    2013-01-01

    Stabilization of virus protein structure and nucleic acid integrity is challenging yet essential to preserve the transcriptional competence of live recombinant viral vaccine vectors in the absence of a cold chain. When coupled with needle-free skin delivery, such a platform would address an unmet need in global vaccine coverage against HIV and other global pathogens. Herein, we show that a simple dissolvable microneedle array (MA) delivery system preserves the immunogenicity of vaccines encoded by live recombinant human adenovirus type 5 (rAdHu5). Specifically, dried rAdHu5 MA immunization induced CD8+ T-cell expansion and multifunctional cytokine responses equipotent with conventional injectable routes of immunization. Intravital imaging demonstrated MA cargo distributed both in the epidermis and dermis, with acquisition by CD11c+ dendritic cells (DCs) in the dermis. The MA immunizing properties were attributable to CD11c+ MHCIIhi CD8αneg epithelial cell adhesion molecule (EpCAMneg) CD11b+ langerin (Lang; CD207)neg DCs, but neither Langerhans cells nor Lang+ DCs were required for CD8+ T-cell priming. This study demonstrates an important technical advance for viral vaccine vectors progressing to the clinic and provides insights into the mechanism of CD8+ T-cell priming by live rAdHu5 MAs. PMID:23386724

  11. Homodimerization and Heterodimerization of Minimal Zinc(II)-Binding Domain Peptides of T-cell Proteins CD4, CD8α, and Lck

    PubMed Central

    Davis, Alisa M.; Berg, Jeremy M.

    2009-01-01

    Metal-mediated protein oligomerization is an emerging mode of protein-protein interaction. The C-terminal cytosolic domains of T-cell coreceptors CD4 and CD8α form zinc-bridged heterodimers with the N-terminal region of the kinase Lck, with each protein contributing two cysteinate ligands to the complex. Using size exclusion chromatography, 1H NMR, and UV/visible absorption spectroscopy with cobalt(II) as a spectroscopic probe, we demonstrate that small peptides derived from these regions form metal-bridged heterodimers but also homodimers, in contrast to previous reports. The Lck-CD4 and Lck-CD8α cobalt(II)-bridged heterodimer complexes are more stable than the corresponding (Lck)2cobalt(II) complex by factors of 11 ± 4 and 22 ± 9, respectively. These studies were aided by the discovery that cobalt(II) complexes with a cobalt(II)(-Cys-X-X-Cys-)(-Cys-X-Cys-) chromophore show unusual optical spectra with one component of the visible d to d (4A2 to 4T1(P)) transition red-shifted and well separated from the other components. These results provide insights into the basis of specificity of metal-bridged complex formation and on the potential biological significance of metal-bridged homodimers in T-cells. PMID:19624124

  12. T Cell factor 1 represses CD8+ effector T cell formation and function.

    PubMed

    Tiemessen, Machteld M; Baert, Miranda R M; Kok, Lianne; van Eggermond, Marja C J A; van den Elsen, Peter J; Arens, Ramon; Staal, Frank J T

    2014-12-01

    The Wnt-responsive transcription factor T cell factor 1 (Tcf1) is well known for its role in thymic T cell development and the formation of memory CD8(+) T cells. However, its role in the initial phases of CD8(+) T effector cell formation has remained unexplored. We report that high levels of Wnt signaling and Tcf1 are operational in naive and memory CD8(+) T cells, whereas Wnt signaling and Tcf1 were low in effector CD8(+) T cells. CD8(+) T cells deficient in Tcf1 produce IFN-γ more rapidly, coinciding with increased demethylation of the IFN-γ enhancer and higher expression of the transcription factors Tbet and Blimp1. Moreover, virus-specific Tcf1(-/-) CD8(+) T cells show accelerated expansion in acute infection, which is associated with increased IFN-γ and TNF production and lower viral load. Genetic complementation experiments with various Tcf1 isoforms indicate that Tcf1 dosage and protein stability are critical in suppressing IFN-γ production. Isoforms lacking the β-catenin binding domain are equally effective in inhibiting CD8(+) effector T cell formation. Thus, Tcf1 functions as a repressor of CD8(+) effector T cell formation in a β-catenin/Wnt-independent manner. PMID:25355919

  13. c-Jun and Ets2 proteins regulate expression of spleen tyrosine kinase in T cells.

    PubMed

    Ghosh, Debjani; Tsokos, George C; Kyttaris, Vasileios C

    2012-04-01

    Effector T cells and T cells from patients with systemic lupus erythematosus (SLE) express increased levels of the spleen tyrosine kinase (Syk). Syk binds to the T cell receptor (TCR)-CD3 complex and transduces the TCR-mediated signal in the cell more efficiently than the canonical CD3ζ chain. The reasons for the increased expression of Syk are unclear. In the present study, we found that Syk is regulated by the transcription factor c-Jun in cooperation with Ets2. c-Jun and Ets2 bound to the SYK promoter in close proximity and increased the promoter activity in a specific manner. Disruption of c-Jun and Ets2 expression by siRNA resulted in decreased expression of Syk. Overexpression of c-Jun but not Ets2 resulted in increase in Syk protein. c-Jun and Ets2 co-immunoprecipitated and had an additive effect on Syk expression. c-Jun-driven SYK promoter activation showed a similar pattern in B cells; however, as expected, basal promoter activity was much higher in B cells as compared with T cells. Overexpression of c-Jun led to increase in intracytoplasmic calcium flux following TCR stimulation. Moreover, we found that SLE T cells had increased levels of c-Jun at baseline and phosphorylated c-Jun upon activation. Finally, disruption of c-Jun and Ets2 in SLE T cells resulted in a decrease in calcium flux upon TCR stimulation. In conclusion, c-Jun in cooperation with Ets2 increases the expression of Syk and contributes to Syk-mediated heightened calcium responses in SLE T cells. PMID:22354960

  14. T cell traffic signals

    PubMed Central

    Van Epps, Heather L.

    2005-01-01

    In 1990, Charles Mackay and colleagues combined classical physiology with modern molecular biology to provide the first concrete evidence that naive and memory T cells follow distinct migratory routes out of the bloodstream— a discovery that helped invigorate the field of lymphocyte homing. PMID:16184630

  15. Neutrophils Contribute to Excess Serum BAFF Levels and Promote CD4+ T Cell and B Cell Responses in Lupus-Prone Mice

    PubMed Central

    Coquery, Christine M.; Wade, Nekeithia S.; Loo, William M.; Kinchen, Jason M.; Cox, Kelly M.; Jiang, Chao; Tung, Kenneth S.; Erickson, Loren D.

    2014-01-01

    Despite increased frequencies of neutrophils found in autoimmune diseases such as systemic lupus erythematosus (SLE), how they contribute to disease pathogenesis and the mechanisms that affect the accumulation of neutrophils are poorly understood. The aim of this study was to identify factors in autoantibody-mediated autoimmunity that controls the accumulation of spleen resident neutrophils and to determine whether neutrophils contribute to abnormal B cell responses. Increased levels of the cytokine BAFF have been linked to loss of B cell tolerance in autoimmunity, but the cellular source responsible for excess BAFF is unknown. B cell maturation antigen (BCMA) is a receptor for BAFF and is critical for the survival of bone marrow plasma cells. Paradoxically, BCMA deficiency exacerbates the formation of autoantibody-secreting plasma cells in spleens of lupus-prone mice and the reasons for this effect are not understood. Here we analyzed the phenotype, localization and function of neutrophils in spleens of healthy mice and congenic lupus-prone mice, and compared mice sufficient or deficient in BCMA expression. Neutrophils were found to be significantly increased in frequency and activation status in spleens of lupus-prone mice when BCMA was absent. Furthermore, neutrophils localized within T cell zones and enhanced CD4+ T cell proliferation and IFNγ production through the production of BAFF. Reduced BAFF and IFNγ serum levels, decreased frequencies of IFNγ-producing T cells, germinal center B cells, and autoantibody production after neutrophil depletion indicated the involvement of neutrophils in these autoimmune traits. Thus, we have identified a novel role for BCMA to control excess BAFF production in murine lupus through restraining the accumulation of BAFF-producing neutrophils. Our data suggests that devising therapeutic strategies to reduce neutrophils in autoimmunity may decrease BAFF levels and ameliorate disease. PMID:25010693

  16. Paxillin binding to the alpha 4 integrin subunit stimulates LFA-1 (integrin alpha L beta 2)-dependent T cell migration by augmenting the activation of focal adhesion kinase/proline-rich tyrosine kinase-2.

    PubMed

    Rose, David M; Liu, Shouchun; Woodside, Darren G; Han, Jaewon; Schlaepfer, David D; Ginsberg, Mark H

    2003-06-15

    Engagement of very late Ag-4 (integrin alpha(4)beta(1)) by ligands such as VCAM-1 markedly stimulates leukocyte migration mediated by LFA-1 (integrin alpha(L)beta(2)). This form of integrin trans-regulation in T cells requires the binding of paxillin to the alpha(4) integrin cytoplasmic domain. This conclusion is based on the abolition of trans-regulation in Jurkat T cells by an alpha(4) mutation (alpha(4)(Y991A)) that disrupts paxillin binding. Furthermore, cellular expression of an alpha(4)-binding fragment of paxillin that blocks the alpha(4)-paxillin interaction, selectively blocked VCAM-1 stimulation of alpha(L)beta(2)-dependent cell migration. The alpha(4)-paxillin association mediates trans-regulation by enhancing the activation of tyrosine kinases, focal adhesion kinase (FAK) and/or proline-rich tyrosine kinase-2 (Pyk2), based on two lines of evidence. First, disruption of the paxillin-binding site in the alpha(4) tail resulted in much less alpha(4)beta(1)-mediated phosphorylation of Pyk2 and FAK. Second, transfection with cDNAs encoding C-terminal fragments of Pyk2 and FAK, which block the function of the intact kinases, blocked alpha(4)beta(1) stimulation of alpha(L)beta(2)-dependent migration. These results define a proximal protein-protein interaction of an integrin cytoplasmic domain required for trans-regulation between integrins, and establish that augmented activation of Pyk2 and/or FAK is an immediate signaling event required for the trans-regulation of integrin alpha(L)beta(2) by alpha(4)beta(1). PMID:12794117

  17. T Cells and Gene Regulation: The Switching On and Turning Up of Genes after T Cell Receptor Stimulation in CD8 T Cells

    PubMed Central

    Conley, James M.; Gallagher, Michael P.; Berg, Leslie J.

    2016-01-01

    Signaling downstream of the T cell receptor (TCR) is directly regulated by the dose and affinity of peptide antigen. The strength of TCR signaling drives a multitude of T cell functions from development to differentiation. CD8 T cells differentiate into a diverse pool of effector and memory cells after activation, a process that is critical for pathogen clearance and is highly regulated by TCR signal strength. T cells rapidly alter their gene expression upon activation. Multiple signaling pathways downstream of the TCR activate transcription factors, which are critical for this process. The dynamics between proximal TCR signaling, transcription factor activation and CD8 T cell function are discussed here. We propose that inducible T cell kinase (ITK) acts as a rheostat for gene expression. This unique regulation of TCR signaling by ITK provides a possible signaling mechanism for the promotion of a diverse T cell repertoire in response to pathogen. PMID:26973653

  18. Promoter-distal RNA polymerase II binding discriminates active from inactive CCAAT/ enhancer-binding protein beta binding sites

    PubMed Central

    Savic, Daniel; Roberts, Brian S.; Carleton, Julia B.; Partridge, E. Christopher; White, Michael A.; Cohen, Barak A.; Cooper, Gregory M.; Gertz, Jason; Myers, Richard M.

    2015-01-01

    Transcription factors (TFs) bind to thousands of DNA sequences in mammalian genomes, but most of these binding events appear to have no direct effect on gene expression. It is unclear why only a subset of TF bound sites are actively involved in transcriptional regulation. Moreover, the key genomic features that accurately discriminate between active and inactive TF binding events remain ambiguous. Recent studies have identified promoter-distal RNA polymerase II (RNAP2) binding at enhancer elements, suggesting that these interactions may serve as a marker for active regulatory sequences. Despite these correlative analyses, a thorough functional validation of these genomic co-occupancies is still lacking. To characterize the gene regulatory activity of DNA sequences underlying promoter-distal TF binding events that co-occur with RNAP2 and TF sites devoid of RNAP2 occupancy using a functional reporter assay, we performed cis-regulatory element sequencing (CRE-seq). We tested more than 1000 promoter-distal CCAAT/enhancer-binding protein beta (CEBPB)-bound sites in HepG2 and K562 cells, and found that CEBPB-bound sites co-occurring with RNAP2 were more likely to exhibit enhancer activity. CEBPB-bound sites further maintained substantial cell-type specificity, indicating that local DNA sequence can accurately convey cell-type–specific regulatory information. By comparing our CRE-seq results to a comprehensive set of genome annotations, we identified a variety of genomic features that are strong predictors of regulatory element activity and cell-type–specific activity. Collectively, our functional assay results indicate that RNAP2 occupancy can be used as a key genomic marker that can distinguish active from inactive TF bound sites. PMID:26486725

  19. Commensal bacteria trigger a full dendritic cell maturation program that promotes the expansion of non-Tr1 suppressor T cells.

    PubMed

    Baba, Nobuyasu; Samson, Sandrine; Bourdet-Sicard, Raphaëlle; Rubio, Manuel; Sarfati, Marika

    2008-08-01

    Dendritic cells (DCs) orchestrate the immune response establishing immunity versus tolerance. These two opposite functions may be dictated by DC maturation status with maturity linked to immunogenicity. DCs directly interact with trillions of noninvasive intestinal bacteria in vivo, a process that contributes to gut homeostasis. We here evaluated the maturation program elicited in human DCs by direct exposure to commensal-related bacteria (CB) in the absence of inflammatory signals. We showed that eight gram(+) and gram(-) CB strains up-regulated costimulatory molecule expression in DCs and provoked a chemokine receptor switch similar to that activated by gram(+) pathogens. CB strains may be classified into three groups according to DC cytokine release: high IL-12 and low IL-10; low IL-12 and high IL-10; and low IL-12 and IL-10. All CB-treated DCs produced IL-1beta and IL-6 and almost no TGF-beta. Yet, CB instructed DCs to convert naive CD4+ T cells into hyporesponsive T cells that secreted low or no IFN-gamma, IL-10, and IL-17 and instead, displayed suppressor function. These data demonstrate that phenotypic DC maturation combined to an appropriate cytokine profile is insufficient to warrant Th1, IL-10-secreting T regulatory Type 1 (Tr1), or Th17 polarization. We propose that commensal flora and as such, probiotics manipulate DCs by a yet-unidentified pathway to enforce gut tolerance. PMID:18511576

  20. Genetic deficiency of Irgm1 (LRG-47) suppresses induction of experimental autoimmune encephalomyelitis by promoting apoptosis of activated CD4+ T cells.

    PubMed

    Xu, Hongwei; Wu, Zhi-Ying; Fang, Fang; Guo, Lan; Chen, Doris; Chen, John Xi; Stern, David; Taylor, Gregory A; Jiang, Hong; Yan, Shirley ShiDu

    2010-05-01

    The immunity-related GTPase Irgm1, also called LRG-47, is known to regulate host resistance to intracellular pathogens through multiple mechanisms that include controlling the survival of T lymphocytes. Here, we address whether Irgm1 also plays a role in the pathogenesis of experimental autoimmune encephalitis (EAE). We find that Irgm1/LRG-47 is a significant factor in the progression of EAE and multiple sclerosis (MS). Expression of Irgm1 was robustly elevated in MS-affected lesions and in the central nervous system (CNS) of myelin basic protein (MBP)-induced EAE mice, especially in cells of lymphoid and mononuclear phagocyte origin. Homozygous Irgm1 null mice were resistant to MBP-induced EAE, and CD4(+) T cells in spleen and CNS of these mice displayed decreased proliferative capacity, increased apoptosis, and up-regulated interferon (IFN)-gamma induction. Therefore, Irgm1-induced survival of autoreactive CD4(+) T cells contributes significantly to the pathogenesis of EAE. Blockade of Irgm1 may be a potential therapeutic strategy for halting multiple sclerosis. PMID:20056715

  1. Defective CD8 T Cell Memory Following Acute Infection Without CD4 T Cell Help

    NASA Astrophysics Data System (ADS)

    Sun, Joseph C.; Bevan, Michael J.

    2003-04-01

    The CD8+ cytotoxic T cell response to pathogens is thought to be CD4+ helper T cell independent because infectious agents provide their own inflammatory signals. Mice that lack CD4+ T cells mount a primary CD8 response to Listeria monocytogenes equal to that of wild-type mice and rapidly clear the infection. However, protective memory to a challenge is gradually lost in the former animals. Memory CD8+ T cells from normal mice can respond rapidly, but memory CD8+ T cells that are generated without CD4 help are defective in their ability to respond to secondary encounters with antigen. The results highlight a previously undescribed role for CD4 help in promoting protective CD8 memory development.

  2. A polymorphism in a phosphotyrosine signalling motif of CD229 (Ly9, SLAMF3) alters SH2 domain binding and T-cell activation.

    PubMed

    Margraf, Stefanie; Garner, Lee I; Wilson, Timothy J; Brown, Marion H

    2015-11-01

    Signalling lymphocyte activation molecule (SLAM) family members regulate activation and inhibition in the innate and adaptive immune systems. Genome-wide association studies identified their genetic locus (1q23) as highly polymorphic and associated with susceptibility to systemic lupus erythematosus (SLE). Here we show that the Val602 variant of the non-synonymous single nucleotide polymorphism (SNP) rs509749 in the SLAM family member CD229 (Ly9, SLAMF3) has a two-fold lower affinity compared with the SLE-associated Met602 variant for the small adaptor protein SAP. Comparison of the two variants in T-cell lines revealed the Val602 variant to be significantly more highly expressed than CD229 Met602 . Activation was diminished in cells expressing CD229 Val602 compared with CD229 Met602 as measured by up-regulation of CD69. There was no correlation between homozygosity at rs509749 and activation in peripheral blood mononuclear cells from healthy donors. These findings identify potential mechanisms by which a single SNP can perturb fine-tuning in the immune system with significant functional consequences. PMID:26221972

  3. Radiation-induced CXCL16 release by breast cancer cells attracts effector T cells1

    PubMed Central

    Matsumura, Satoko; Wang, Baomei; Kawashima, Noriko; Braunstein, Steve; Badura, Michelle; Cameron, Thomas O.; Babb, James S.; Schneider, Robert J.; Formenti, Silvia C.; Dustin, Michael L.; Demaria, Sandra

    2008-01-01

    Recruitment of effector T cells to inflamed peripheral tissues is regulated by chemokines and their receptors, but the factors regulating recruitment to tumors remain largely undefined. Ionizing radiation (IR) therapy is a common treatment modality for breast and other cancers. Used as a cytocidal agent for proliferating cancer cells, IR in combination with immunotherapy has been shown to promote immune-mediated tumor destruction in pre-clinical studies. Here we demonstrate that IR markedly enhanced the secretion by mouse and human breast cancer cells of CXCL16, a chemokine that binds to CXCR6 on Th1 and activated CD8 effector T cells, and plays an important role in their recruitment to sites of inflammation. Employing a poorly immunogenic mouse model of breast cancer, we found that irradiation increased the migration of CD8+CXCR6+ activated T cells to tumors in vitro and in vivo. CXCR6-deficient mice showed reduced infiltration of tumors by activated CD8 T cells and impaired tumor regression following treatment with local IR to the tumor and antibodies blocking the negative regulator of T cell activation CTLA-4. These results provide the first evidence that IR can induce the secretion by cancer cells of pro-inflammatory chemotactic factors that recruit anti-tumor effector T cells. The ability of IR to convert tumors into “inflamed” peripheral tissues could be exploited to overcome obstacles at the effector phase of the anti-tumor immune response and improve the therapeutic efficacy of immunotherapy. PMID:18713980

  4. Neem leaf glycoprotein promotes dual generation of central and effector memory CD8(+) T cells against sarcoma antigen vaccine to induce protective anti-tumor immunity.

    PubMed

    Ghosh, Sarbari; Sarkar, Madhurima; Ghosh, Tithi; Guha, Ipsita; Bhuniya, Avishek; Saha, Akata; Dasgupta, Shayani; Barik, Subhasis; Bose, Anamika; Baral, Rathindranath

    2016-03-01

    We have previously shown that Neem Leaf Glycoprotein (NLGP) mediates sustained tumor protection by activating host immune response. Now we report that adjuvant help from NLGP predominantly generates CD44(+)CD62L(high)CCR7(high) central memory (TCM; in lymph node) and CD44(+)CD62L(low)CCR7(low) effector memory (TEM; in spleen) CD8(+) T cells of Swiss mice after vaccination with sarcoma antigen (SarAg). Generated TCM and TEM participated either to replenish memory cell pool for sustained disease free states or in rapid tumor eradication respectively. TCM generated after SarAg+NLGP vaccination underwent significant proliferation and IL-2 secretion following SarAg re-stimulation. Furthermore, SarAg+NLGP vaccination helps in greater survival of the memory precursor effector cells at the peak of the effector response and their maintenance as mature memory cells, in comparison to single modality treatment. Such response is corroborated with the reduced phosphorylation of FOXO in the cytosol and increased KLF2 in the nucleus associated with enhanced CD62L, CCR7 expression of lymph node-resident CD8(+) T cells. However, spleen-resident CD8(+) T memory cells show superior efficacy for immediate memory-to-effector cell conversion. The data support in all aspects that SarAg+NLGP demonstrate superiority than SarAg vaccination alone that benefits the host by rapid effector functions whenever required, whereas, central-memory cells are thought to replenish the memory cell pool for ultimate sustained disease free survival till 60 days following post-vaccination tumor inoculation. PMID:26851529

  5. Non-invasive Imaging of Sendai Virus Infection in Pharmacologically Immunocompromised Mice: NK and T Cells, but not Neutrophils, Promote Viral Clearance after Therapy with Cyclophosphamide and Dexamethasone.

    PubMed

    Mostafa, Heba H; Vogel, Peter; Srinivasan, Ashok; Russell, Charles J

    2016-09-01

    In immunocompromised patients, parainfluenza virus (PIV) infections have an increased potential to spread to the lower respiratory tract (LRT), resulting in increased morbidity and mortality. Understanding the immunologic defects that facilitate viral spread to the LRT will help in developing better management protocols. In this study, we immunosuppressed mice with dexamethasone and/or cyclophosphamide then monitored the spread of viral infection into the LRT by using a noninvasive bioluminescence imaging system and a reporter Sendai virus (murine PIV type 1). Our results show that immunosuppression led to delayed viral clearance and increased viral loads in the lungs. After cessation of cyclophosphamide treatment, viral clearance occurred before the generation of Sendai-specific antibody responses and coincided with rebounds in neutrophils, T lymphocytes, and natural killer (NK) cells. Neutrophil suppression using anti-Ly6G antibody had no effect on infection clearance, NK-cell suppression using anti-NK antibody delayed clearance, and T-cell suppression using anti-CD3 antibody resulted in no clearance (chronic infection). Therapeutic use of hematopoietic growth factors G-CSF and GM-CSF had no effect on clearance of infection. In contrast, treatment with Sendai virus-specific polysera or a monoclonal antibody limited viral spread into the lungs and accelerated clearance. Overall, noninvasive bioluminescence was shown to be a useful tool to study respiratory viral progression, revealing roles for NK and T cells, but not neutrophils, in Sendai virus clearance after treatment with dexamethasone and cyclophosphamide. Virus-specific antibodies appear to have therapeutic potential. PMID:27589232

  6. The glycocalyx promotes cooperative binding and clustering of adhesion receptors.

    PubMed

    Xu, Guang-Kui; Qian, Jin; Hu, Jinglei

    2016-05-18

    Cell adhesion plays a pivotal role in various biological processes, e.g., immune responses, cancer metastasis, and stem cell differentiation. The adhesion behaviors depend subtly on the binding kinetics of receptors and ligands restricted at the cell-substrate interfaces. Although much effort has been directed toward investigating the kinetics of adhesion molecules, the role of the glycocalyx, anchored on cell surfaces as an exterior layer, is still unclear. In this paper, we propose a theoretical approach to study the collective binding kinetics of a few and a large number of binders in the presence of the glycocalyx, representing the cases of initial and mature adhesions of cells, respectively. The analytical results are validated by finding good agreement with our Monte Carlo simulations. In the force loading case, the on-rate and affinity increase as more bonds form, whereas this cooperative effect is not observed in the displacement loading case. The increased thickness and stiffness of the glycocalyx tend to decrease the affinity for a few bonds, while they have less influence on the affinity for a large number of bonds. Moreover, for a flexible membrane with thermally-excited shape fluctuations, the glycocalyx is exhibited to promote the formation of bond clusters, mainly due to the cooperative binding of binders. This study helps to understand the cooperative kinetics of adhesion receptors under physiologically relevant loading conditions and sheds light on the novel role of the glycocalyx in cell adhesion. PMID:27102288

  7. Fe(2+) binding on amyloid β-peptide promotes aggregation.

    PubMed

    Boopathi, Subramaniam; Kolandaivel, Ponmalai

    2016-09-01

    The metal ions Zn(2+) , Cu(2+) , and Fe(2+) play a significant role in the aggregation mechanism of Aβ peptides. However, the nature of binding between metal and peptide has remained elusive; the detailed information on this from the experimental study is very difficult. Density functional theory (dft) (M06-2X/6-311++G (2df,2pd) +LANL2DZ) has employed to determine the force field resulting due to metal and histidine interaction. We performed 200 ns molecular dynamics (MD) simulation on Aβ1-42 -Zn(2+) , Aβ1-42 -Cu(2+) , and Aβ1-42 -Fe(2+) systems in explicit water with different combination of coordinating residues including the three Histidine residues in the N-terminal. The present investigation, the Aβ1-42 -Zn(2+) system possess three turn conformations separated by coil structure. Zn(2+) binding caused the loss of the helical structure of N-terminal residues which transformed into the S-shaped conformation. Zn(2+) has reduced the coil and increases the turn content of the peptide compared with experimental study. On the other hand, the Cu(2+) binds with peptide, β sheet formation is observed at the N-terminal residues of the peptide. Fe(2+) binding is to promote the formation of Glu22-Lys28 salt-bridge which stabilized the turn conformation in the Phe19-Gly25 residues, subsequently β sheets were observed at His13-Lys18 and Gly29-Gly37 residues. The turn conformation facilitates the β sheets are arranged in parallel by enhancing the hydrophobic contact between Gly25 and Met35, Lys16 and Met35, Leu17 and Leu34, Val18 and Leu34 residues. The Fe(2+) binding reduced the helix structure and increases the β sheet content in the peptide, which suggested, Fe(2+) promotes the oligomerization by enhancing the peptide-peptide interaction. Proteins 2016; 84:1257-1274. © 2016 Wiley Periodicals, Inc. PMID:27214008

  8. Pharmacologic suppression of target cell recognition by engineered T cells expressing chimeric T-cell receptors.

    PubMed

    Alvarez-Vallina, L; Yañez, R; Blanco, B; Gil, M; Russell, S J

    2000-04-01

    Adoptive therapy with autologous T cells expressing chimeric T-cell receptors (chTCRs) is of potential interest for the treatment of malignancy. To limit possible T-cell-mediated damage to normal tissues that weakly express the targeted tumor antigen (Ag), we have tested a strategy for the suppression of target cell recognition by engineered T cells. Jurkat T cells were transduced with an anti-hapten chTCR tinder the control of a tetracycline-suppressible promoter and were shown to respond to Ag-positive (hapten-coated) but not to Ag-negative target cells. The engineered T cells were then reacted with hapten-coated target cells at different effector to target cell ratios before and after exposure to tetracycline. When the engineered T cells were treated with tetracycline, expression of the chTCR was greatly decreased and recognition of the hapten-coated target cells was completely suppressed. Tetracycline-mediated suppression of target cell recognition by engineered T cells may be a useful strategy to limit the toxicity of the approach to cancer gene therapy. PMID:10811469

  9. Designer T cells by T cell receptor replacement.

    PubMed

    Sommermeyer, Daniel; Neudorfer, Julia; Weinhold, Monika; Leisegang, Matthias; Engels, Boris; Noessner, Elfriede; Heemskerk, Mirjam H M; Charo, Jehad; Schendel, Dolores J; Blankenstein, Thomas; Bernhard, Helga; Uckert, Wolfgang

    2006-11-01

    T cell receptor (TCR) gene transfer is a convenient method to produce antigen-specific T cells for adoptive therapy. However, the expression of two TCR in T cells could impair their function or cause unwanted effects by mixed TCR heterodimers. With five different TCR and four different T cells, either mouse or human, we show that some TCR are strong--in terms of cell surface expression--and replace weak TCR on the cell surface, resulting in exchange of antigen specificity. Two strong TCR are co-expressed. A mouse TCR replaces human TCR on human T cells. Even though it is still poorly understood why some TCRalpha/beta combinations are preferentially expressed on T cells, our data suggest that, in the future, designer T cells with exclusive tumor reactivity can be generated by T cell engineering. PMID:17051621

  10. Current progress in γδ T-cell biology

    PubMed Central

    Hao, Jianlei; Wu, Xiaoli; Xia, Siyuan; Li, Zheng; Wen, Ti; Zhao, Na; Wu, Zhenzhou; Wang, Puyue; Zhao, Liqing; Yin, Zhinan

    2010-01-01

    T lymphocytes bearing γ- and δ-chain T-cell receptor heterodimers are named γδ T cells. Interestingly, γδ and αβ T cells share the same progenitors, and they undergo a fate decision in the thymus. Functional differentiation of γδ T cells occurs both inside and outside the thymus. Antigen recognition of γδ T-cell receptors is very unique, and the responses frequently exhibit innate characteristics. Nevertheless, peripheral γδ T cells exert a number of effector and regulatory functions. γδ T cells rapidly produce cytokines like interferon (IFN)-γ and IL-17 and promote inflammation, partly due to the inherent epigenetic and transcriptional programs, which facilitates a quick and extensive response. Moreover, γδ T cells lyse target cells directly, and this is necessary for pathogen or tumor clearance. γδ T cells can even serve as regulatory cells, and may contribute to immune suppression. Orchestration of γδ T-cell and other immune cell interactions may be critical for host defense and immune regulation. Recently, γδ T cells have been used for immunotherapy for infectious diseases and malignancy. In this review, we summarize the abstracts presented at the recent γδ T cell Conference held from 19 to 21 May 2010, in Kiel, Germany (please see the website for details: http://www.gammadelta-conference.uni-kiel.de/index.html). PMID:21042298

  11. Memory CD4 T cells in influenza.

    PubMed

    Zens, Kyra D; Farber, Donna L

    2015-01-01

    Influenza A virus is a significant cause of morbidity and mortality worldwide, particularly among young children and the elderly. Current vaccines induce neutralizing antibody responses directed toward highly variable viral surface proteins, resulting in limited heterosubtypic protection to new viral serotypes. By contrast, memory CD4 T cells recognize conserved viral proteins and are cross-reactive to multiple influenza strains. In humans, virus-specific memory CD4 T cells were found to be the protective correlate in human influenza challenge studies, suggesting their key role in protective immunity. In mouse models, memory CD4 T cells can mediate protective responses to secondary influenza infection independent of B cells or CD8 T cells, and can influence innate immune responses. Importantly, a newly defined, tissue-resident CD4 memory population has been demonstrated to be retained in lung tissue and promote optimal protective responses to an influenza infection. Here, we review the current state of results regarding the generation of memory CD4 T cells following primary influenza infection, mechanisms for their enhanced efficacy in protection from secondary challenge including their phenotype, localization, and function in the context of both mouse models and human infection. We also discuss the generation of memory CD4 T cells in response to influenza vaccines and its future implications for vaccinology. PMID:25005927

  12. MHC class II transactivator represses human IL-4 gene transcription by interruption of promoter binding with CBP/p300, STAT6 and NFAT1 via histone hypoacetylation

    PubMed Central

    Zhou, Xiaorong; Jiang, Yang; Lu, Liming; Ding, Qing; Jiao, Zhijun; Zhou, Yun; Xin, Lijun; Chou, Kuang-Yen

    2007-01-01

    In addition to its property of enhancing major histocompatibility complex (MHC) class II expression, the class II transactivator (CIITA) was recently demonstrated to be involved in T helper type 1/type 2 (Th1/Th2) differentiation by regulating interleukin-4 (IL-4) gene transcription. There was however, controversy regarding whether CIITA promotes or suppresses IL-4 expression in the experiments with transgenic mice. To clarify the discrepancy by using simpler experimental systems, human Jurkat T cells that express IL-4 but not interferon-γ, even if stimulated with phorbol 12-myristate 13-acetate plus ionomycin, were used for CIITA transfection. Significant suppression of IL-4 gene expression was demonstrated. Simultaneously, histones H3 and H4 in the IL-4 promoter were hypoacetylated. The suppression could be totally reversed by the histone deacetylatase inhibitor trichostatin A. Furthermore, the IL-4 expression was determined in primarily established human Th1/Th2 cells to which CIITA small interference RNA (siRNA) had been introduced. A substantially increased level of IL-4 was recorded in the CIITA siRNA-transfected Th1 cells, which was in parallel with significantly enhanced acetylation in histone H3 of the IL-4 promoter. Chromatin immunoprecipitation analysis indicated that CIITA abrogated the binding of coactivator CBP/p300 and transcription factors STAT6/NFAT1 to IL-4 promoter in the CIITA-transfected cells. In conclusion, CIITA was active in the repression of transcription activation of human IL-4 gene in both the T-cell line and the primary human CD4 T cells by preventing transcription factors from binding to IL-4 promoter through histone hypoacetylation. Our data confirm a potential significant role of CIITA in controlling Th1/Th2 differentiation via modulation of IL-4 gene activation. PMID:17645498

  13. I spy alloreactive T cells.

    PubMed

    Alegre, Maria-Luisa

    2015-01-28

    High-throughput sequencing of the T cell receptor Vβ CDR3 region allowed longitudinal tracking of alloreactive T cells in kidney transplant patients, revealing clonal deletion as a mechanism of transplantation tolerance (Morris et al., this issue). PMID:25632032

  14. Auto-reactive T cells revised. Overestimation based on methodology?

    PubMed

    Thorlacius-Ussing, Gorm; Sørensen, Jesper F; Wandall, Hans H; Pedersen, Anders Elm

    2015-05-01

    Autoreactive T cells have been identified in most autoimmune diseases and recently even in healthy individuals. Similar, T cells that recognize either wild-type or tumorspecific tumor antigens have been increasingly reported to develop spontaneously in cancer patients. This insight has become possible mainly due to novel immunoassays which have revolutionized the discovery of rare antigen specific T cells. At present, the major dogma that explains this increasing number of reports of autoreactive T cells is that autoreactive T cells are counteracted by CD4+CD25+ regulatory T (Treg) cells in vivo, in particular in healthy individuals, whereas dysfunction in Tregs or Treg responsiveness may unmask the autoreactive T cell responses in patients with autoimmune diseases. However, studies that identify autoreactive T cells are usually performed by culturing T cells with antigen presenting cells loaded with E. coli produced recombinant protein or unmodified synthetic HLA binding peptides. Our concern is that this approach may ignore the presence of natural genetic variation and post-translational modifications such as e.g. the complex nature of N- and O-linked glycosylation of mammalian proteins. Thus, T cell antigen reactivities identified with unmodified antigens in vitro may in part represent in vitro T cell activation against neo-epitopes and not true in vivo autoreactivity as postulated. This methodological problem may have implications for the interpretation of the frequent reporting of autoreactive T cells in autoimmunity, T cell responses to wild-type tumor antigens in cancer patients and most important for the increasing reports on naïve T cells with specificity against self-antigens in healthy individuals. Here, we discuss and provide examples for the possibility that the experimental methodology applied to document T cell reactivity against unmodified protein or peptide may lead to overinterpretation of the reported frequencies of autoreactive CD4+ and CD8+ T

  15. The transcription factor BATF operates as an essential differentiation checkpoint in early effector CD8+ T cells

    PubMed Central

    Kurachi, Makoto; Barnitz, R. Anthony; Yosef, Nir; Odorizzi, Pamela M.; Dilorio, Michael A.; Lemieux, Madeleine E.; Yates, Kathleen; Godec, Jernej; Klatt, Martin G.; Regev, Aviv; Wherry, E. John; Haining, W. Nicholas

    2014-01-01

    The transcription factor BATF is required for interleukin 17 (IL-17)-producing helper T cell (TH17) and follicular helper T cell (TFH) differentiation. Here, we show that BATF also has a fundamental role in regulating effector CD8+ T cell differentiation. BATF-deficient CD8+ T cells show profound defects in effector expansion and undergo proliferative and metabolic catastrophe early after antigen encounter. BATF, together with IRF4 and Jun proteins, binds to and promotes early expression of genes encoding lineage-specific transcription-factors (T-bet and Blimp-1) and cytokine receptors, while paradoxically repressing genes encoding effector molecules (IFN-γ and granzyme B). Thus, BATF amplifies TCR-dependent transcription factor expression and augments inflammatory signal propagation but restrains effector gene expression. This checkpoint prevents irreversible commitment to an effector fate until a critical threshold of downstream transcriptional activity has been achieved. PMID:24584090

  16. Ultra-low Dose Interleukin-2 Promotes Immune-modulating Function of Regulatory T Cells and Natural Killer Cells in Healthy Volunteers

    PubMed Central

    Ito, Sawa; Bollard, Catherine M; Carlsten, Mattias; Melenhorst, Jan Joseph; Biancotto, Angélique; Wang, Ena; Chen, Jinguo; Kotliarov, Yuri; Cheung, Foo; Xie, Zhi; Marincola, Francesco; Tanimoto, Kazushi; Battiwalla, Minoo; Olnes, Matthew J; Perl, Shira; Schum, Paula; Hughes, Thomas E; Keyvanfar, Keyvan; Hensel, Nancy; Muranski, Pawel; Young, Neal S; Barrett, A John

    2014-01-01

    Low-dose interleukin-2 (IL-2) expands regulatory T cells (Tregs) and natural killer (NK) cells after stem cell transplantation (SCT) and may reduce graft-versus-host disease (GVHD). We hypothesized that ultra-low dose (ULD) IL-2 could serve as an immune-modulating agent for stem cell donors to prevent GVHD following SCT. However, the safety, dose level, and immune signatures of ULD IL-2 in immune-competent healthy subjects remain unknown. Here, we have characterized the phenotype and function of Tregs and NK cells as well as the gene expression and cytokine profiles of 21 healthy volunteers receiving 50,000 to 200,000 units/m2/day IL-2 for 5 days. ULD IL-2 was well tolerated and induced a significant increase in the frequency of Tregs with increased suppressive function. There was a marked expansion of CD56bright NK cells with enhanced interferon-γ (IFN-γ) production. Serum cytokine profiling demonstrated increase of IFN-γ induced protein 10 (IP-10). Gene expression analysis revealed significant changes in a highly restricted set of genes, including FOXP3, IL-2RA, and CISH. This is the first study to evaluate global immune-modulating function of ULD IL-2 in healthy subjects and to support the future studies administrating ULD IL-2 to stem cell donors. PMID:24686272

  17. γδ T Cell-Dependent Regulatory T Cells Prevent the Development of Autoimmune Keratitis.

    PubMed

    Huang, Yafei; Yang, Zhifang; Huang, Chunjian; McGowan, Jessica; Casper, Tamara; Sun, Deming; Born, Willi K; O'Brien, Rebecca L

    2015-12-15

    To prevent potentially damaging inflammatory responses, the eye actively promotes local immune tolerance via a variety of mechanisms. Owing to trauma, infection, or other ongoing autoimmunity, these mechanisms sometimes fail, and an autoimmune disorder may develop in the eye. In mice of the C57BL/10 (B10) background, autoimmune keratitis often develops spontaneously, particularly in the females. Its incidence is greatly elevated in the absence of γδ T cells, such that ∼80% of female B10.TCRδ(-/-) mice develop keratitis by 18 wk of age. In this article, we show that CD8(+) αβ T cells are the drivers of this disease, because adoptive transfer of CD8(+), but not CD4(+), T cells to keratitis-resistant B10.TCRβ/δ(-/-) hosts induced a high incidence of keratitis. This finding was unexpected because in other autoimmune diseases, more often CD4(+) αβ T cells, or both CD4(+) and CD8(+) αβ T cells, mediate the disease. Compared with wild-type B10 mice, B10.TCRδ(-/-) mice also show increased percentages of peripheral memory phenotype CD8(+) αβ T cells, along with an elevated frequency of CD8(+) αβ T cells biased to produce inflammatory cytokines. In addition, B10.TCRδ-/- mice have fewer peripheral CD4(+) CD25(+) Foxp3(+) αβ regulatory T cells (Tregs), which express lower levels of receptors needed for Treg development and function. Together, these observations suggest that in B10 background mice, γδ T cells are required to generate adequate numbers of CD4(+) CD25(+) Foxp3(+) Tregs, and that in B10.TCRδ(-/-) mice a Treg deficiency allows dysregulated effector or memory CD8(+) αβ T cells to infiltrate the cornea and provoke an autoimmune attack. PMID:26566677

  18. Interleukin-6 expands homeostatic space for peripheral T cells.

    PubMed

    Hong, Changwan; Nam, Anna S; Keller, Hilary R; Ligons, Davinna L; Park, Joo-Young; Yoon, Hee-Won; Park, Joseph J; Luckey, Megan A; Park, Jung-Hyun

    2013-11-01

    T cell homeostasis and survival is dependent on interleukin-7 (IL-7). Immune activation, however, downregulates IL-7 receptor expression on T cells so that T cell survival during activation must be maintained independently of IL-7. The pro-inflammatory cytokine IL-6 shares common signaling pathways with IL-7 and can promote T cell survival in vitro. But whether IL-6 promotes T cell survival and homeostasis in vivo is not clear. Notably, IL-6 overexpression results in massive plasmacytosis and autoimmunity so that an IL-6 effect on in vivo T cell survival has remained untested. To overcome this limitation, here we generated IL-6 transgenic mice on an immunoglobulin heavy chain (IgH) deficient background which rendered them B cell deficient. Notably, such IgH(KO)IL6(Tg) mice were free of any signs of inflammation or autoimmunity and remained healthy throughout the course of analysis. In these mice, we found that IL-6 overexpression significantly increased peripheral T cell numbers, but importantly without increasing thymopoiesis. Moreover, IL-6 signaled T cells maintained their naïve phenotype and did not express activation/memory markers, suggesting that increased T cell numbers were due to increased T cell survival and not because of expansion of activated T cells. Mechanistically, we found that IL-6 signaling induced expression of pro-survival factors Mcl-1 and Pim-1/-2 but not Bcl-2. Thus, IL-6 is a T cell homeostatic cytokine that expands T cell space and can maintain the naïve T cell pool. PMID:23988623

  19. T Cell Intrinsic Heterodimeric Complexes between HVEM and BTLA Determine Receptivity to the Surrounding Microenvironment1

    PubMed Central

    Cheung, Timothy C.; Oborne, Lisa M.; Steinberg, Marcos W.; Macauley, Matthew G.; Fukuyama, Satoshi; Sanjo, Hideki; D’Souza, Claire; Norris, Paula S.; Pfeffer, Klaus; Murphy, Kenneth M.; Kronenberg, Mitchell; Spear, Patricia G.; Ware, Carl F.

    2010-01-01

    The inhibitory cosignaling pathway formed between the TNF receptor herpesvirus entry mediator (HVEM, TNFRSF14) and the Ig superfamily members, B and T lymphocyte attenuator (BTLA) and CD160, limits the activation of T cells. However, BTLA and CD160 can also serve as activating ligands for HVEM when presented in trans by adjacent cells, thus forming a bidirectional signaling pathway. BTLA and CD160 can directly activate the HVEM-dependent NF-κB RelA transcriptional complex raising the question of how NF-κB activation is repressed in naive T cells. In this study, we show BTLA interacts with HVEM in cis, forming a heterodimeric complex in naive T cells that inhibits HVEM-dependent NF-κB activation. The cis-interaction between HVEM and BTLA is the predominant form expressed on the surface of naive human and mouse T cells. The BTLA ectodomain acts as a competitive inhibitor blocking BTLA and CD160 from binding in trans to HVEM and initiating NF-κB activation. The TNF-related ligand, LIGHT (homologous to lymphotoxins, exhibits inducible expression, and competes with HSV glycoprotein D for HVEM, a receptor expressed by T lymphocytes, or TNFSF14) binds HVEM in the cis-complex, but NF-κB activation was attenuated, suggesting BTLA prevents oligomerization of HVEM in the cis-complex. Genetic deletion of BTLA or pharmacologic disruption of the HVEM-BTLA cis-complex in T cells promoted HVEM activation in trans. Interestingly, herpes simplex virus envelope glycoprotein D formed a cis-complex with HVEM, yet surprisingly, promoted the activation NF-κB RelA. We suggest that the HVEM-BTLA cis-complex competitively inhibits HVEM activation by ligands expressed in the surrounding microenvironment, thus helping maintain T cells in the naive state. PMID:19915044

  20. In silico prediction of the T-cell and IgE-binding epitopes of Per a 6 and Bla g 6 allergens in cockroaches.

    PubMed

    Chen, Hao; Yang, Hai-Wei; Wei, Ji-Fu; Tao, Ai-Lin

    2014-10-01

    Per a 6 and Bla g 6 are cockroach allergens found in Periplaneta americana and Blattella germanica, respectively. The objective of the present study was to predict the B‑ and T‑cell epitopes of the Per a 6 and Bla g 6 allergens. Three immunoinformatics tools, the DNAStar Protean system, the Bioinformatics Predicted Antigenic Peptides system and the BepiPred 1.0 server, were used to predict the potential B‑cell epitopes, while Net‑MHCIIpan‑2.0 and NetMHCII‑2.2 were used to predict the T‑cell epitopes of the two allergens. As a result, seven peptides were predicted in the Per a 6 allergen and seven peptides were predicted in the Bla g 6 allergen in the B‑cell epitope predictions. In the T‑cell prediction, the Per a 6 allergen was predicted to have nine strongly binding nonamer core epitope sequences (IC50<50 nm) and 28 weakly binding sequences (50 nm

  1. The Genotype of Early-Transmitting HIV gp120s Promotes α4β7 –Reactivity, Revealing α4β7+/CD4+ T cells As Key Targets in Mucosal Transmission

    PubMed Central

    Van Ryk, Donald; Block, Katharine E.; Jelicic, Katija; McNally, Jonathan P.; Ogundare, Olajumoke; Pascuccio, Massimiliano; Patel, Nikita; Wei, Danlan; Fauci, Anthony S.; Arthos, James

    2011-01-01

    Mucosal transmission of HIV is inefficient. The virus must breach physical barriers before it infects mucosal CD4+ T cells. Low-level viral replication occurs initially in mucosal CD4+ T cells, but within days high-level replication occurs in Peyer's patches, the gut lamina propria and mesenteric lymph nodes. Understanding the early events in HIV transmission may provide valuable information relevant to the development of an HIV vaccine. The viral quasispecies in a donor contracts through a genetic bottleneck in the recipient, such that, in low-risk settings, infection is frequently established by a single founder virus. Early-transmitting viruses in subtypes A and C mucosal transmission tend to encode gp120s with reduced numbers of N-linked glycosylation sites at specific positions throughout the V1-V4 domains, relative to typical chronically replicating isolates in the donor quasispecies. The transmission advantage gained by the absence of these N-linked glycosylation sites is unknown. Using primary α4β7+/CD4+ T cells and a flow-cytometry based steady-state binding assay we show that the removal of transmission-associated N-linked glycosylation sites results in large increases in the specific reactivity of gp120 for integrin- α4β7. High-affinity for integrin α4β7, although not found in many gp120s, was observed in early-transmitting gp120s that we analyzed. Increased α4β7 affinity is mediated by sequences encoded in gp120 V1/V2. α4β7-reactivity was also influenced by N-linked glycosylation sites located in C3/V4. These results suggest that the genetic bottleneck that occurs after transmission may frequently involve a relative requirement for the productive infection of α4β7+/CD4+ T cells. Early-transmitting gp120s were further distinguished by their dependence on avidity-effects to interact with CD4, suggesting that these gp120s bear unusual structural features not present in many well-characterized gp120s derived from chronically replicating viruses

  2. A stochastic T cell response criterion

    PubMed Central

    Currie, James; Castro, Mario; Lythe, Grant; Palmer, Ed; Molina-París, Carmen

    2012-01-01

    The adaptive immune system relies on different cell types to provide fast and coordinated responses, characterized by recognition of pathogenic challenge, extensive cellular proliferation and differentiation, as well as death. T cells are a subset of the adaptive immune cellular pool that recognize immunogenic peptides expressed on the surface of antigen-presenting cells by means of specialized receptors on their membrane. T cell receptor binding to ligand determines T cell responses at different times and locations during the life of a T cell. Current experimental evidence provides support to the following: (i) sufficiently long receptor–ligand engagements are required to initiate the T cell signalling cascade that results in productive signal transduction and (ii) counting devices are at work in T cells to allow signal accumulation, decoding and translation into biological responses. In the light of these results, we explore, with mathematical models, the timescales associated with T cell responses. We consider two different criteria: a stochastic one (the mean time it takes to have had N receptor–ligand complexes bound for at least a dwell time, τ, each) and one based on equilibrium (the time to reach a threshold number N of receptor–ligand complexes). We have applied mathematical models to previous experiments in the context of thymic negative selection and to recent two-dimensional experiments. Our results indicate that the stochastic criterion provides support to the thymic affinity threshold hypothesis, whereas the equilibrium one does not, and agrees with the ligand hierarchy experimentally established for thymic negative selection. PMID:22745227

  3. Estrogen Therapy Delays Autoimmune Diabetes and Promotes the Protective Efficiency of Natural Killer T-Cell Activation in Female Nonobese Diabetic Mice.

    PubMed

    Gourdy, Pierre; Bourgeois, Elvire A; Levescot, Anaïs; Pham, Linh; Riant, Elodie; Ahui, Marie-Louise; Damotte, Diane; Gombert, Jean-Marc; Bayard, Francis; Ohlsson, Claes; Arnal, Jean-François; Herbelin, André

    2016-01-01

    Therapeutic strategies focused on restoring immune tolerance remain the main avenue to prevent type 1 diabetes (T1D). Because estrogens potentiate FoxP3+ regulatory T cells (Treg) and invariant natural killer T (iNKT) cells, two regulatory lymphocyte populations that are functionally deficient in nonobese diabetic (NOD) mice, we investigated whether estradiol (E2) therapy influences the course of T1D in this model. To this end, female NOD mice were sc implanted with E2- or placebo-delivering pellets to explore the course of spontaneous and cyclophosphamide-induced diabetes. Treg-depleted and iNKT-cell-deficient (Jα18(-/-)) NOD mice were used to assess the respective involvement of these lymphocyte populations in E2 effects. Early E2 administration (from 4 wk of age) was found to preserve NOD mice from both spontaneous and cyclophosphamide-induced diabetes, and a complete protection was also observed throughout treatment when E2 treatment was initiated after the onset of insulitis (from 12 wk of age). This delayed E2 treatment remained fully effective in Treg-depleted mice but failed to entirely protect Jα18(-/-) mice. Accordingly, E2 administration was shown to restore the cytokine production of iNKT cells in response to in vivo challenge with the cognate ligand α-galactosylceramide. Finally, transient E2 administration potentiated the previously described protective action of α-galactosylceramide treatment in NOD females. This study provides original evidence that E2 therapy strongly protects NOD mice from T1D and reveals the estrogen/iNKT cell axis as a new effective target to counteract diabetes onset at the stage of insulitis. Estrogen-based therapy should thus be considered for T1D prevention. PMID:26485613

  4. An Oral Salmonella-Based Vaccine Inhibits Liver Metastases by Promoting Tumor-Specific T-Cell-Mediated Immunity in Celiac and Portal Lymph Nodes: A Preclinical Study

    PubMed Central

    Vendrell, Alejandrina; Mongini, Claudia; Gravisaco, María José; Canellada, Andrea; Tesone, Agustina Inés; Goin, Juan Carlos; Waldner, Claudia Inés

    2016-01-01

    Primary tumor excision is one of the most widely used therapies of cancer. However, the risk of metastases development still exists following tumor resection. The liver is a common site of metastatic disease for numerous cancers. Breast cancer is one of the most frequent sources of metastases to the liver. The aim of this work was to evaluate the efficacy of the orally administered Salmonella Typhi vaccine strain CVD 915 on the development of liver metastases in a mouse model of breast cancer. To this end, one group of BALB/c mice was orogastrically immunized with CVD 915, while another received PBS as a control. After 24 h, mice were injected with LM3 mammary adenocarcinoma cells into the spleen and subjected to splenectomy. This oral Salmonella-based vaccine produced an antitumor effect, leading to a decrease in the number and volume of liver metastases. Immunization with Salmonella induced an early cellular immune response in mice. This innate stimulation rendered a large production of IFN-γ by intrahepatic immune cells (IHIC) detected within 24 h. An antitumor adaptive immunity was found in the liver and celiac and portal lymph nodes (LDLN) 21 days after oral bacterial inoculation. The antitumor immune response inside the liver was associated with increased CD4+ and dendritic cell populations as well as with an inflammatory infiltrate located around liver metastatic nodules. Enlarged levels of inflammatory cytokines (IFN-γ and TNF) were also detected in IHIC. Furthermore, a tumor-specific production of IFN-γ and TNF as well as tumor-specific IFN-γ-producing CD8 T cells (CD8+IFN-γ+) were found in the celiac and portal lymph nodes of Salmonella-treated mice. This study provides first evidence for the involvement of LDLN in the development of an efficient cellular immune response against hepatic tumors, which resulted in the elimination of liver metastases after oral Salmonella-based vaccination. PMID:26973649

  5. T cell factor-1 negatively regulates expression of IL-17 family of cytokines and protects mice from experimental autoimmune encephalomyelitis.

    PubMed

    Yu, Qing; Sharma, Archna; Ghosh, Amalendu; Sen, Jyoti Misra

    2011-04-01

    Activated CD4 T cells are associated with protective immunity and autoimmunity. The manner in which the inflammatory potential of T cells and resultant autoimmunity is restrained is poorly understood. In this article, we demonstrate that T cell factor-1 (TCF1) negatively regulates the expression of IL-17 and related cytokines in activated CD4 T cells. We show that TCF1 does not affect cytokine signals and expression of transcription factors that have been shown to regulate Th17 differentiation. Instead, TCF1 regulates IL-17 expression, in part, by binding to the regulatory regions of the Il17 gene. Moreover, TCF1-deficient Th17 CD4 T cells express higher levels of IL-7Rα, which potentially promotes their survival and expansion in vivo. Accordingly, TCF1-deficient mice are hyperresponsive to experimental autoimmune encephalomyelitis. Thus, TCF1, a constitutively expressed T cell-specific transcription factor, is a critical negative regulator of the inflammatory potential of TCR-activated T cells and autoimmunity. PMID:21339363

  6. Memory T Cells in Transplantation

    PubMed Central

    Su, Charles A.; Fairchild, Robert L.

    2014-01-01

    Following infections and environmental exposures, memory T cells are generated that provide long-term protective immunity. Compared to their naïve T cell counterparts, memory T cells possess unique characteristics that endow them with the ability to quickly and robustly respond to foreign antigens. While such memory T cells are beneficial in protecting their hosts from recurrent infection, memory cells reactive to donor antigens pose a major barrier to successful transplantation and tolerance induction. Significant progress has been made over the past several decades contributing to our understanding of memory T cell generation, their distinct biology, and their detrimental impact in clinical and animal models of transplantation. This review focuses on the unique features which make memory T cells relevant to the transplant community and discusses potential therapies targeting memory T cells which may ameliorate allograft rejection. PMID:25435071

  7. Immunomodulatory Effects of Polysaccharide from Marine Fungus Phoma herbarum YS4108 on T Cells and Dendritic Cells

    PubMed Central

    Chen, Song; Zhou, Yan; Zhang, Xian; Zhu, Rui; Gao, Xiang-Dong

    2014-01-01

    YCP, as a kind of natural polysaccharides from the mycelium of marine filamentous fungus Phoma herbarum YS4108, has great antitumor potential via enhancement of host immune response, but little is known about the molecular mechanisms. In the present study, we mainly focused on the effects and mechanisms of YCP on the specific immunity mediated by dendritic cells (DCs) and T cells. T cell /DC activation-related factors including interferon- (IFN-) γ, interleukin-12 (IL-12), and IL-4 were examined with ELISA. Receptor knock-out mice and fluorescence-activated cell sorting are used to analyze the YCP-binding receptor of T cells and DCs. RT-PCR is utilized to measure MAGE-A3 for analyzing the tumor-specific killing effect. In our study, we demonstrated YCP can provide the second signal for T cell activation, proliferation, and IFN-γ production through binding to toll-like receptor- (TLR-) 2 and TLR-4. YCP could effectively promote IL-12 secretion and expression of markers (CD80, CD86, and MHC II) via TLR-4 on DCs. Antigen-specific immunity against mouse melanoma cells was strengthened through the activation of T cells and the enhancement of capacity of DCs by YCP. The data supported that YCP can exhibit specific immunomodulatory capacity mediated by T cells and DCs. PMID:25525304

  8. Immediate Dysfunction of Vaccine-Elicited CD8+ T Cells Primed in the Absence of CD4+ T Cells.

    PubMed

    Provine, Nicholas M; Larocca, Rafael A; Aid, Malika; Penaloza-MacMaster, Pablo; Badamchi-Zadeh, Alexander; Borducchi, Erica N; Yates, Kathleen B; Abbink, Peter; Kirilova, Marinela; Ng'ang'a, David; Bramson, Jonathan; Haining, W Nicholas; Barouch, Dan H

    2016-09-01

    CD4(+) T cell help is critical for optimal CD8(+) T cell memory differentiation and maintenance in many experimental systems. In addition, many reports have identified reduced primary CD8(+) T cell responses in the absence of CD4(+) T cell help, which often coincides with reduced Ag or pathogen clearance. In this study, we demonstrate that absence of CD4(+) T cells at the time of adenovirus vector immunization of mice led to immediate impairments in early CD8(+) T cell functionality and differentiation. Unhelped CD8(+) T cells exhibited a reduced effector phenotype, decreased ex vivo cytotoxicity, and decreased capacity to produce cytokines. This dysfunctional state was imprinted within 3 d of immunization. Unhelped CD8(+) T cells expressed elevated levels of inhibitory receptors and exhibited transcriptomic exhaustion and anergy profiles by gene set enrichment analysis. Dysfunctional, impaired effector differentiation also occurred following immunization of CD4(+) T cell-deficient mice with a poxvirus vector. This study demonstrates that following priming with viral vectors, CD4(+) T cell help is required to promote both the expansion and acquisition of effector functions by CD8(+) T cells, which is accomplished by preventing immediate dysfunction. PMID:27448585

  9. TRAF3 is required for T cell-mediated immunity and T cell receptor/CD28 signaling1

    PubMed Central

    Xie, Ping; Kraus, Zachary J.; Stunz, Laura L.; Liu, Yan; Bishop, Gail A.

    2011-01-01

    We recently reported that TRAF3, a ubiquitously expressed adaptor protein, promotes mature B cell apoptosis. However, the specific function of TRAF3 in T cells has remained unclear. Here we report the generation and characterization of T cell-specific TRAF3−/− mice, in which the TRAF3 gene was deleted from thymocytes and T cells. Ablation of TRAF3 in the T cell-lineage did not affect the numbers or proportions of CD4+,CD8+ or double positive or negative thymocytes, or CD4 or CD8 T cell populations in secondary lymphoid organs except that the T cell specific TRAF3−/− mice had a two-fold increase in FoxP3+ T cells.. In striking contrast to mice lacking TRAF3 in B cells, the T cell TRAF3 deficient mice exhibited defective IgG1 responses to a T dependent antigen, and impaired T cell-mediated immunity to infection with Listeria monocytogenes. Surprisingly, we found that TRAF3 was recruited to the TCR/CD28 signaling complex upon co-stimulation, and that TCR/CD28-mediated proximal and distal signaling events were compromised by TRAF3 deficiency. These findings provide new insights into the roles played by TRAF3 in T cell activation and T cell-mediated immunity. PMID:21084666

  10. NAD+ regulates Treg cell fate and promotes allograft survival via a systemic IL-10 production that is CD4+ CD25+ Foxp3+ T cells independent

    PubMed Central

    Elkhal, Abdallah; Rodriguez Cetina Biefer, Hector; Heinbokel, Timm; Uehara, Hirofumi; Quante, Markus; Seyda, Midas; Schuitenmaker, Jeroen M.; Krenzien, Felix; Camacho, Virginia; de la Fuente, Miguel A.; Ghiran, Ionita; Tullius, Stefan G.

    2016-01-01

    CD4+ CD25+ Foxp3+ Tregs have been shown to play a central role in immune homeostasis while preventing from fatal inflammatory responses, while Th17 cells have traditionally been recognized as pro-inflammatory mediators implicated in a myriad of diseases. Studies have shown the potential of Tregs to convert into Th17 cells, and Th17 cells into Tregs. Increasing evidence have pointed out CD25 as a key molecule during this transdifferentiation process, however molecules that allow such development remain unknown. Here, we investigated the impact of NAD+ on the fate of CD4+ CD25+ Foxp3+ Tregs in-depth, dissected their transcriptional signature profile and explored mechanisms underlying their conversion into IL-17A producing cells. Our results demonstrate that NAD+ promotes Treg conversion into Th17 cells in vitro and in vivo via CD25 cell surface marker. Despite the reduced number of Tregs, known to promote homeostasis, and an increased number of pro-inflammatory Th17 cells, NAD+ was able to promote an impressive allograft survival through a robust systemic IL-10 production that was CD4+ CD25+ Foxp3+ independent. Collectively, our study unravels a novel immunoregulatory mechanism of NAD+ that regulates Tregs fate while promoting allograft survival that may have clinical applications in alloimmunity and in a wide spectrum of inflammatory conditions. PMID:26928119

  11. Bax alpha perturbs T cell development and affects cell cycle entry of T cells.

    PubMed Central

    Brady, H J; Gil-Gómez, G; Kirberg, J; Berns, A J

    1996-01-01

    Bax alpha can heterodimerize with Bcl-2 and Bcl-X(L), countering their effects, as well as promoting apoptosis on overexpression. We show that bax alpha transgenic mice have greatly reduced numbers of mature T cells, which results from an impaired positive selection in the thymus. This perturbation in positive selection is accompanied by an increase in the number of cycling thymocytes. Further to this, mature T cells overexpressing Bax alpha have lower levels of p27Kip1 and enter S phase more rapidly in response to interleukin-2 stimulation than do control T cells, while the converse is true of bcl-2 transgenic T cells. These data indicate that apoptotic regulatory proteins can modulate the level of cell cycle-controlling proteins and thereby directly impact on the cell cycle. Images PMID:9003775

  12. The novel C24D synthetic polypeptide inhibits binding of placenta immunosuppressive ferritin to human T cells and elicits anti-breast cancer immunity in vitro and in vivo.

    PubMed

    Solodeev, Inna; Zahalka, Muayad A; Moroz, Chaya

    2014-09-01

    Immune tolerance mechanisms supporting normal human pregnancy are exploited by breast cancer and other malignancies. We cloned from human placenta and breast cancer cells the novel human immunomodulator named placenta immunosuppressive ferritin (PLIF). PLIF is composed of a ferritin heavy chain-like domain and a novel cytokine-like domain, named C48. Both intact PLIF and C48 inhibit T cell proliferation. Blocking PLIF by specific antibodies in a tolerant breast cancer model in nude mice resulted in tumor cell apoptosis and rejection. This prompted us to study active immune preventive strategies targeting PLIF activity. Currently, we report on the design and synthesis of the novel C24D polypeptide, which inhibits the binding of PLIF to T cells and therefore inhibits the immune suppressive effect of PLIF. The effect of C24D on the generation of anti-breast cancer cytotoxic T lymphocytes (CTLs) was studied in vitro in cultures of MCF-7 (HLA-A2(+)) or T47D (HLA-A2(-)) breast cancer cells incubated with peripheral blood mononuclear cells (PBMCs) from healthy blood donors. We found that C24D treatment exclusively induced development of CTLs. On reactivation by their specific target cells, the CTLs secreted interferon-γ and induced target apoptosis. Anti-MCF-7 CTLs were cross-cytotoxic to MDA-MB-231 (HLA-A2(+)) triple-negative breast cancer but not to T47D. Moreover, C24D treatment in vivo inhibited the growth of MCF-7 tumors engrafted in immune-compromised nude mice transfused with naïve allogeneic human PBMCs. Our results demonstrate that C24D treatment breakdown breast cancer induced tolerance enabling the initiation of effective anti-tumor immune response. PMID:25246274

  13. Chimeric Antigen Receptors Modified T-Cells for Cancer Therapy

    PubMed Central

    Dai, Hanren; Wang, Yao; Lu, Xuechun

    2016-01-01

    The genetic modification and characterization of T-cells with chimeric antigen receptors (CARs) allow functionally distinct T-cell subsets to recognize specific tumor cells. The incorporation of costimulatory molecules or cytokines can enable engineered T-cells to eliminate tumor cells. CARs are generated by fusing the antigen-binding region of a monoclonal antibody (mAb) or other ligand to membrane-spanning and intracellular-signaling domains. They have recently shown clinical benefit in patients treated with CD19-directed autologous T-cells. Recent successes suggest that the modification of T-cells with CARs could be a powerful approach for developing safe and effective cancer therapeutics. Here, we briefly review early studies, consider strategies to improve the therapeutic potential and safety, and discuss the challenges and future prospects for CAR T-cells in cancer therapy. PMID:26819347

  14. Chimeric Antigen Receptors Modified T-Cells for Cancer Therapy.

    PubMed

    Dai, Hanren; Wang, Yao; Lu, Xuechun; Han, Weidong

    2016-07-01

    The genetic modification and characterization of T-cells with chimeric antigen receptors (CARs) allow functionally distinct T-cell subsets to recognize specific tumor cells. The incorporation of costimulatory molecules or cytokines can enable engineered T-cells to eliminate tumor cells. CARs are generated by fusing the antigen-binding region of a monoclonal antibody (mAb) or other ligand to membrane-spanning and intracellular-signaling domains. They have recently shown clinical benefit in patients treated with CD19-directed autologous T-cells. Recent successes suggest that the modification of T-cells with CARs could be a powerful approach for developing safe and effective cancer therapeutics. Here, we briefly review early studies, consider strategies to improve the therapeutic potential and safety, and discuss the challenges and future prospects for CAR T-cells in cancer therapy. PMID:26819347

  15. Chimeric Antigen Receptors Modified T-Cells for Cancer Therapy

    PubMed Central

    Dai, Hanren; Wang, Yao; Lu, Xuechun

    2016-01-01

    The genetic modification and characterization of T-cells with chimeric antigen receptors (CARs) allow functionally distinct T-cell subsets to recognize specific tumor cells. The incorporation of costimulatory molecules or cytokines can enable engineered T-cells to eliminate tumor cells. CARs are generated by fusing the antigen-binding region of a monoclonal antibody (mAb) or other ligand to membrane-spanning and intracellular-signaling domains. They have recently shown clinical benefit in patients treated with CD19-directed autologous T-cells. Recent successes suggest that the modification of T-cells with CARs could be a powerful approach for developing safe and effective cancer therapeutics. Here, we briefly review early studies, consider strategies to improve the therapeutic potential and safety, and discuss the challenges and future prospects for CAR T-cells in cancer therapy.

  16. A "hotspot" for autoimmune T cells in type 1 diabetes.

    PubMed

    Stadinski, Brian D; Obst, Reinhard; Huseby, Eric S

    2016-06-01

    The ability of a single T cell antigen receptor (TCR) to cross-react with multiple antigens allows the finite number of T cells within an organism to respond to the compendium of pathogen challenges faced during a lifetime. Effective immune surveillance, however, comes at a price. TCR cross-reactivity can allow molecular mimics to spuriously activate autoimmune T cells; it also underlies T cell rejection of organ transplants and drives graft-versus-host disease. In this issue of the JCI, Cole and colleagues provide insight into how an insulin-reactive T cell cross-reacts with pathogen-derived antigens by focusing on a limited portion of the peptides to provide a hotspot for binding. These findings dovetail with recent studies of alloreactive and autoimmune TCRs and suggest that the biochemical principles that govern conventional protein-protein interactions may allow the specificity and cross-reactivity profiles of T cells to be predicted. PMID:27183386

  17. Regulation of Lipid Specific and Vitamin Specific Non-MHC Restricted T Cells by Antigen Presenting Cells and Their Therapeutic Potentials

    PubMed Central

    Salio, Mariolina; Cerundolo, Vincenzo

    2015-01-01

    Since initial reports, more than 25 years ago, that T cells recognize lipids in the context on non-polymorphic CD1 molecules, our understanding of antigen presentation to non-peptide-specific T cell populations has deepened. It is now clear that αβ T cells bearing semi-invariant T cell receptor, as well as subsets of γδ T cells, recognize a variety of self and non-self lipids and contribute to shaping immune responses via cross talk with dendritic cells and B cells. Furthermore, it has been demonstrated that small molecules derived from the microbial riboflavin biosynthetic pathway (vitamin B2) bind monomorphic MR1 molecules and activate mucosal-associated invariant T cells, another population of semi-invariant T cells. Novel insights in the biological relevance of non-peptide-specific T cells have emerged with the development of tetrameric CD1 and MR1 molecules, which has allowed accurate enumeration and functional analysis of CD1- and MR1-restricted T cells in humans and discovery of novel populations of semi-invariant T cells. The phenotype and function of non-peptide-specific T cells will be discussed in the context of the known distribution of CD1 and MR1 molecules by different subsets of antigen-presenting cells at steady state and following infection. Concurrent modulation of CD1 transcription and lipid biosynthetic pathways upon TLR stimulation, coupled with efficient lipid antigen processing, result in the increased cell surface expression of antigenic CD1–lipid complexes. Similarly, MR1 expression is almost undetectable in resting APC and it is upregulated following bacterial infection, likely due to stabilization of MR1 molecules by microbial antigens. The tight regulation of CD1 and MR1 expression at steady state and during infection may represent an important mechanism to limit autoreactivity, while promoting T cell responses to foreign antigens. PMID:26284072

  18. The integrin αvβ6: a novel target for CAR T-cell immunotherapy?

    PubMed

    Whilding, Lynsey M; Vallath, Sabari; Maher, John

    2016-04-15

    Immunotherapy of cancer using chimeric antigen receptor (CAR) T-cells is a rapidly expanding field. CARs are fusion molecules that couple the binding of a tumour-associated cell surface target to the delivery of a tailored T-cell activating signal. Re-infusion of such genetically engineered T-cells to patients with haematological disease has demonstrated unprecedented response rates in Phase I clinical trials. However, such successes have not yet been observed using CAR T-cells against solid malignancies and this is, in part, due to a lack of safe tumour-specific targets. The αvβ6 integrin is strongly up-regulated in multiple solid tumours including those derived from colon, lung, breast, cervix, ovaries/fallopian tube, pancreas and head and neck. It is associated with poorer prognosis in several cancers and exerts pro-tumorigenic activities including promotion of tumour growth, migration and invasion. By contrast, physiologic expression of αvβ6 is largely restricted to wound healing. These attributes render this epithelial-specific integrin a highly attractive candidate for targeting using immunotherapeutic strategies such as CAR T-cell adoptive immunotherapy. This mini-review will discuss the role and expression of αvβ6 in cancer, as well as its potential as a therapeutic target. PMID:27068939

  19. Sp1 binds two sites in the CD11c promoter in vivo specifically in myeloid cells and cooperates with AP1 to activate transcription.

    PubMed Central

    Noti, J D; Reinemann, B C; Petrus, M N

    1996-01-01

    The leukocyte integrin gene, CD11c, is transcriptionally regulated and is expressed predominantly on differentiated cells of the myelomonocytic lineage. In this study we have demonstrated that the regions -72 to -63 and -132 to -104 of the CD11c promoter contain elements responsible for phorbol ester-induced differentiation of the myeloid cell line HL60. DNase I footprinting analysis revealed that these regions can bind purified Sp1, and supershift analysis with Sp1 antibody confirmed that Sp1 in HL60 nuclear extracts could bind these regions. Transfection analysis of CD11c promoter-chloramphenicol acetyltransferase constructs containing deletions of these Sp1-binding sites revealed that these sites are essential for expression of the CD11c gene in HL60 cells but not in the T-cell line Molt4 or the cervical carcinoma cell line HeLa. Moreover, cotransfection of pPacSp1 along with these CD11c promoter-chloramphenicol acetyltransferase constructs into Sp1-deficient Drosophila Schneider 2 cells verified that these sites are essential for Sp1-dependent expression of the CD11c promoter. In vivo genomic footprinting revealed that Sp1 contacts the CD11c promoter within the regions -69 to -63 and -116 to -105 in phorbol 12-myristate 13-acetate-differentiated HL60 cells but not in undifferentiated HL60 cells or in Molt4 or HeLa cells. Cotransfection assays in HL60 cells revealed that Sp1 acts synergistically with Ap1 to activate CD11c. Further, both Sp1 sites are capable of cooperating with AP1. In vitro DNase I footprinting analysis with purified Sp1 and c-jun proteins showed that Sp1 binding could facilitate binding of c-jun. We propose that myeloid-specific expression of the CD11c promoter and is facilitated by cooperative interaction between the Sp1- and Ap1-binding sites. PMID:8649405

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

    NASA Astrophysics Data System (ADS)

    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

  1. Genome-wide promoter binding profiling of protein phosphatase-1 and its major nuclear targeting subunits

    PubMed Central

    Verheyen, Toon; Görnemann, Janina; Verbinnen, Iris; Boens, Shannah; Beullens, Monique; Van Eynde, Aleyde; Bollen, Mathieu

    2015-01-01

    Protein phosphatase-1 (PP1) is a key regulator of transcription and is targeted to promoter regions via associated proteins. However, the chromatin binding sites of PP1 have never been studied in a systematic and genome-wide manner. Methylation-based DamID profiling in HeLa cells has enabled us to map hundreds of promoter binding sites of PP1 and three of its major nuclear interactors, i.e. RepoMan, NIPP1 and PNUTS. Our data reveal that the α, β and γ isoforms of PP1 largely bind to distinct subsets of promoters and can also be differentiated by their promoter binding pattern. PP1β emerged as the major promoter-associated isoform and shows an overlapping binding profile with PNUTS at dozens of active promoters. Surprisingly, most promoter binding sites of PP1 are not shared with RepoMan, NIPP1 or PNUTS, hinting at the existence of additional, largely unidentified chromatin-targeting subunits. We also found that PP1 is not required for the global chromatin targeting of RepoMan, NIPP1 and PNUTS, but alters the promoter binding specificity of NIPP1. Our data disclose an unexpected specificity and complexity in the promoter binding of PP1 isoforms and their chromatin-targeting subunits. PMID:25990731

  2. CTLA4Ig inhibits effector T cells through regulatory T cells and TGFβ1

    PubMed Central

    Deppong, Christine M.; Bricker, Traci L.; Rannals, Brandy D.; Van Rooijen, Nico; Hsieh, Chyi-Song; Green, Jonathan M.

    2013-01-01

    The CD28 costimulatory receptor is a critical regulator of T cell function making it an attractive therapeutic target for the treatment of immune mediated diseases. CTLA4Ig, now approved for use in humans, prevents naive T cell activation by binding to B7-proteins and blocking engagement of CD28. However, CTLA4Ig suppresses inflammation even if administered when disease is established, suggesting alternative mechanisms. We identified a novel, CD28-independent mechanism by which CTLA4Ig inhibits activated T cells. We show that in vitro, CTLA4Ig synergizes with nitric oxide from bone marrow derived macrophages to inhibit T cell proliferation. Depletion of Tregs or interference with TGFβ signaling abrogated the inhibitory effect of CTLA4Ig. Parallel in vivo experiments using an allergic airway inflammation model demonstrated that this novel mechanism required both macrophages andTregs. Furthermore, CTLA4Ig was ineffective in SMAD3-deficient mice, supporting a requirement for TGFβ signaling. Thus, in addition to preventing naïve T cells from being fully activated, CTLA4Ig can turn off already activated effector T cells by an NO/Treg/TGFβ-dependent pathway. This mechanism is similar to cell extrinsic effects of endogenous CTLA-4 and may be particularly important in the ability of CTLA4Ig to treat chronic inflammatory disease. PMID:23956428

  3. The human T-cell leukemia virus type 1 Rex regulatory protein exhibits an impaired functionality in human lymphoblastoid Jurkat T cells.

    PubMed Central

    Hamaia, S; Cassé, H; Gazzolo, L; Duc Dodon, M

    1997-01-01

    The Rex protein of human T-cell leukemia virus type 1 (HTLV-1) intervenes in the posttranscriptional regulation of proviral gene expression. Its binding to the Rex response element (XRE) present in the 3' long terminal repeat ensures the coordinate cytoplasmic accumulation of spliced and unspliced forms of viral messengers. Consequently, synthesis of viral structural and enzymatic proteins is strictly dependent on the Rex posttranscriptional activity. Here we report that synthesis of HTLV-1 envelope glycoproteins by Jurkat T cells could be detected only when they were regulated in a Rex-independent manner. Indeed, Jurkat T cells transfected with a Rex-dependent env expression vector (encompassing both the env and pX open reading frames) do not produce significant levels of envelope glycoproteins despite the production of significant amounts of Rex protein. The analysis of levels and distribution patterns of the unspliced env and of the singly spliced tax/rex transcripts suggests that the failure in envelope glycoprotein synthesis may be ascribed to a deficiency of Rex in mediating the nucleocytoplasmic transport of unspliced env RNAs in these cells. Furthermore, despite the synthesis of regulatory proteins, HTLV-1 structural proteins were not detected in Jurkat T cells transfected with an HTLV-1 infectious provirus. Conversely, and as expected, structural proteins were produced by Jurkat cells transfected by a human immunodeficiency virus type 1 (HIV-1) infectious provirus. This phenotype appeared to be linked to a specific dysfunction of Rex, since the functionally equivalent Rev protein of HIV-1 was shown to be fully efficient in promoting the synthesis of HTLV-1 envelope glycoproteins in Jurkat cells. Therefore, it seems likely that the block to Rex function in these lymphoblastoid T cells is determined by inefficient Rex-XRE interactions. These observations suggest that the acquisition of this Rex-deficient phenotype by in vivo-infected HTLV-1 T cells may

  4. The ubiquitin ligase Stub1 negatively modulates regulatory T cell suppressive activity by promoting degradation of the transcription factor Foxp3.

    PubMed

    Chen, Zuojia; Barbi, Joseph; Bu, Shurui; Yang, Huang-Yu; Li, Zhiyuan; Gao, Yayi; Jinasena, Dilini; Fu, Juan; Lin, Fang; Chen, Chen; Zhang, Jing; Yu, Ning; Li, Xiangpei; Shan, Zhao; Nie, Jia; Gao, Zhimei; Tian, Hong; Li, Yangyang; Yao, Zhengju; Zheng, Ying; Park, Benjamin V; Pan, Ziyi; Zhang, Jing; Dang, Eric; Li, Zhiguang; Wang, Honglin; Luo, Weibo; Li, Liwu; Semenza, Gregg L; Zheng, Song-Guo; Loser, Karin; Tsun, Andy; Greene, Mark I; Pardoll, Drew M; Pan, Fan; Li, Bin

    2013-08-22

    Regulatory T (Treg) cells suppress inflammatory immune responses and autoimmunity caused by self-reactive T cells. The key Treg cell transcription factor Foxp3 is downregulated during inflammation to allow for the acquisition of effector T cell-like functions. Here, we demonstrate that stress signals elicited by proinflammatory cytokines and lipopolysaccharides lead to the degradation of Foxp3 through the action of the E3 ubiquitin ligase Stub1. Stub1 interacted with Foxp3 to promote its K48-linked polyubiquitination in an Hsp70-dependent manner. Knockdown of endogenous Stub1 or Hsp70 prevented Foxp3 degradation. Furthermore, the overexpression of Stub1 in Treg cells abrogated their ability to suppress inflammatory immune responses in vitro and in vivo and conferred a T-helper-1-cell-like phenotype. Our results demonstrate the critical role of the stress-activated Stub1-Hsp70 complex in promoting Treg cell inactivation, thus providing a potential therapeutic target for the intervention against autoimmune disease, infection, and cancer. PMID:23973223

  5. Immunity Related GTPase Irgm1 promotes expansion of activated CD4+ T cell populations by preventing interferon-γ-induced cell death

    PubMed Central

    Feng, Carl G.; Zheng, Lixin; Jankovic, Dragana; Báfica, André; Cannons, Jennifer L.; Watford, Wendy T.; Chaussabel, Damien; Hieny, Sara; Caspar, Patricia; Schwartzberg, Pamela L.; Lenardo, Michael J.; Sher, Alan

    2008-01-01

    Summary Mice deficient in interferon-γ (IFN-γ) inducible immunity-related GTPase, Irgm1, display defective host resistance to a variety of intracellular pathogens. This increased susceptibility to infection is associated with impaired IFN-γ-dependent macrophage microbicidal activity in vitro. Here, we show that Irgm1 also regulated the survival of mature effector CD4+ T lymphocytes by protecting them from IFN-γ-induced autophagic cell death. Mice deficient in both IFN-γ and Irgm1 were rescued from the lymphocyte depletion and increased mortality that occurs in single Irgm1–/– animals following mycobacterial infection. These studies reveal a feedback mechanism in the TH1 response that limits the detrimental effects of IFN-γ on effector T lymphocyte survival while promoting the anti-microbial functions of IFN-γ. PMID:18806793

  6. The Lupus Susceptibility Gene Pbx1 Regulates the Balance between Follicular Helper T Cell and Regulatory T Cell Differentiation.

    PubMed

    Choi, Seung-Chul; Hutchinson, Tarun E; Titov, Anton A; Seay, Howard R; Li, Shiwu; Brusko, Todd M; Croker, Byron P; Salek-Ardakani, Shahram; Morel, Laurence

    2016-07-15

    Pbx1 controls chromatin accessibility to a large number of genes and is entirely conserved between mice and humans. The Pbx1-d dominant-negative isoform is more frequent in CD4(+) T cells from lupus patients than from healthy controls. Pbx1-d is associated with the production of autoreactive T cells in mice carrying the Sle1a1 lupus-susceptibility locus. Transgenic (Tg) expression of Pbx1-d in CD4(+) T cells reproduced the phenotypes of Sle1a1 mice, with increased inflammatory functions of CD4(+) T cells and impaired Foxp3(+) regulatory T cell (Treg) homeostasis. Pbx1-d-Tg expression also expanded the number of follicular helper T cells (TFHs) in a cell-intrinsic and Ag-specific manner, which was enhanced in recall responses and resulted in Th1-biased Abs. Moreover, Pbx1-d-Tg CD4(+) T cells upregulated the expression of miR-10a, miR-21, and miR-155, which were implicated in Treg and follicular helper T cell homeostasis. Our results suggest that Pbx1-d impacts lupus development by regulating effector T cell differentiation and promoting TFHs at the expense of Tregs. In addition, our results identify Pbx1 as a novel regulator of CD4(+) T cell effector function. PMID:27296664

  7. Immediate Dysfunction of Vaccine-Elicited CD8+ T Cells Primed in the Absence of CD4+ T Cells

    PubMed Central

    Provine, Nicholas M.; Larocca, Rafael A.; Aid, Malika; Penaloza-MacMaster, Pablo; Badamchi-Zadeh, Alexander; Borducchi, Erica N.; Yates, Kathleen B.; Abbink, Peter; Kirilova, Marinela; Ng’ang’a, David; Bramson, Jonathan; Haining, W. Nicholas

    2016-01-01

    CD4+ T cell help is critical for optimal CD8+ T cell memory differentiation and maintenance in many experimental systems. In addition, many reports have identified reduced primary CD8+ T cell responses in the absence of CD4+ T cell help, which often coincides with reduced Ag or pathogen clearance. In this study, we demonstrate that absence of CD4+ T cells at the time of adenovirus vector immunization of mice led to immediate impairments in early CD8+ T cell functionality and differentiation. Unhelped CD8+ T cells exhibited a reduced effector phenotype, decreased ex vivo cytotoxicity, and decreased capacity to produce cytokines. This dysfunctional state was imprinted within 3 d of immunization. Unhelped CD8+ T cells expressed elevated levels of inhibitory receptors and exhibited transcriptomic exhaustion and anergy profiles by gene set enrichment analysis. Dysfunctional, impaired effector differentiation also occurred following immunization of CD4+ T cell–deficient mice with a poxvirus vector. This study demonstrates that following priming with viral vectors, CD4+ T cell help is required to promote both the expansion and acquisition of effector functions by CD8+ T cells, which is accomplished by preventing immediate dysfunction. PMID:27448585

  8. Inducible T-cell receptor expression in precursor T cells for leukemia control.

    PubMed

    Hoseini, S S; Hapke, M; Herbst, J; Wedekind, D; Baumann, R; Heinz, N; Schiedlmeier, B; Vignali, D A A; van den Brink, M R M; Schambach, A; Blazar, B R; Sauer, M G

    2015-07-01

    Co-transplantation of hematopoietic stem cells with those engineered to express leukemia-reactive T-cell receptors (TCRs) and differentiated ex vivo into precursor T cells (preTs) may reduce the risk of leukemia relapse. As expression of potentially self-(leukemia-) reactive TCRs will lead to negative selection or provoke autoimmunity upon thymic maturation, we investigated a novel concept whereby TCR expression set under the control of an inducible promoter would allow timely controlled TCR expression. After in vivo maturation and gene induction, preTs developed potent anti-leukemia effects. Engineered preTs provided protection even after repeated leukemia challenges by giving rise to effector and central memory cells. Importantly, adoptive transfer of TCR-transduced allogeneic preTs mediated anti-leukemia effect without evoking graft-versus-host disease (GVHD). Earlier transgene induction forced CD8(+) T-cell development was required to obtain a mature T-cell subset of targeted specificity, allowed engineered T cells to efficiently pass positive selection and abrogated the endogenous T-cell repertoire. Later induction favored CD4 differentiation and failed to produce a leukemia-reactive population emphasizing the dominant role of positive selection. Taken together, we provide new functional insights for the employment of TCR-engineered precursor cells as a controllable immunotherapeutic modality with significant anti-leukemia activity. PMID:25652739

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

    SciTech Connect

    Lacoste, J.; Cohen, L.; Hiscott, J. )

    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.

  10. Co-administration of plasmid-encoded granulocyte-macrophage colony-stimulating factor increases human immunodeficiency virus-1 DNA vaccine-induced polyfunctional CD4+ T-cell responses

    PubMed Central

    Santana, Vinicius Canato; Almeida, Rafael Ribeiro; Ribeiro, Susan Pereira; Ferreira, Luís Carlos de Souza; Kalil, Jorge; Rosa, Daniela Santoro; Cunha-Neto, Edecio

    2015-01-01

    T-cell based vaccines against human immunodeficiency virus (HIV) generate specific responses that may limit both transmission and disease progression by controlling viral load. Broad, polyfunctional, and cytotoxic CD4+T-cell responses have been associated with control of simian immunodeficiency virus/HIV-1 replication, supporting the inclusion of CD4+ T-cell epitopes in vaccine formulations. Plasmid-encoded granulocyte-macrophage colony-stimulating factor (pGM-CSF) co-administration has been shown to induce potent CD4+ T-cell responses and to promote accelerated priming and increased migration of antigen-specific CD4+ T-cells. However, no study has shown whether co-immunisation with pGM-CSF enhances the number of vaccine-induced polyfunctional CD4+ T-cells. Our group has previously developed a DNA vaccine encoding conserved, multiple human leukocyte antigen (HLA)-DR binding HIV-1 subtype B peptides, which elicited broad, polyfunctional and long-lived CD4+ T-cell responses. Here, we show that pGM-CSF co-immunisation improved both magnitude and quality of vaccine-induced T-cell responses, particularly by increasing proliferating CD4+ T-cells that produce simultaneously interferon-γ, tumour necrosis factor-α and interleukin-2. Thus, we believe that the use of pGM-CSF may be helpful for vaccine strategies focused on the activation of anti-HIV CD4+ T-cell immunity. PMID:26602876

  11. Co-administration of plasmid-encoded granulocyte-macrophage colony-stimulating factor increases human immunodeficiency virus-1 DNA vaccine-induced polyfunctional CD4+ T-cell responses.

    PubMed

    Santana, Vinicius Canato; Almeida, Rafael Ribeiro; Ribeiro, Susan Pereira; Ferreira, Luís Carlos de Souza; Kalil, Jorge; Rosa, Daniela Santoro; Cunha-Neto, Edecio

    2015-12-01

    T-cell based vaccines against human immunodeficiency virus (HIV) generate specific responses that may limit both transmission and disease progression by controlling viral load. Broad, polyfunctional, and cytotoxic CD4+T-cell responses have been associated with control of simian immunodeficiency virus/HIV-1 replication, supporting the inclusion of CD4+ T-cell epitopes in vaccine formulations. Plasmid-encoded granulocyte-macrophage colony-stimulating factor (pGM-CSF) co-administration has been shown to induce potent CD4+ T-cell responses and to promote accelerated priming and increased migration of antigen-specific CD4+ T-cells. However, no study has shown whether co-immunisation with pGM-CSF enhances the number of vaccine-induced polyfunctional CD4+ T-cells. Our group has previously developed a DNA vaccine encoding conserved, multiple human leukocyte antigen (HLA)-DR binding HIV-1 subtype B peptides, which elicited broad, polyfunctional and long-lived CD4+ T-cell responses. Here, we show that pGM-CSF co-immunisation improved both magnitude and quality of vaccine-induced T-cell responses, particularly by increasing proliferating CD4+ T-cells that produce simultaneously interferon-γ, tumour necrosis factor-α and interleukin-2. Thus, we believe that the use of pGM-CSF may be helpful for vaccine strategies focused on the activation of anti-HIV CD4+ T-cell immunity. PMID:26602876

  12. An Interleukin-33-Mast Cell-Interleukin-2 Axis Suppresses Papain-Induced Allergic Inflammation by Promoting Regulatory T Cell Numbers

    PubMed Central

    Morita, Hideaki; Arae, Ken; Unno, Hirotoshi; Miyauchi, Kousuke; Toyama, Sumika; Nambu, Aya; Oboki, Keisuke; Ohno, Tatsukuni; Motomura, Kenichiro; Matsuda, Akira; Yamaguchi, Sachiko; Narushima, Seiko; Kajiwara, Naoki; Iikura, Motoyasu; Suto, Hajime; McKenzie, Andrew N.J.; Takahashi, Takao; Karasuyama, Hajime; Okumura, Ko; Azuma, Miyuki; Moro, Kazuyo; Akdis, Cezmi A.; Galli, Stephen J.; Koyasu, Shigeo; Kubo, Masato; Sudo, Katsuko; Saito, Hirohisa; Matsumoto, Kenji; Nakae, Susumu

    2015-01-01

    SUMMARY House dust mite-derived proteases contribute to allergic disorders in part by disrupting epithelial barrier function. Interleukin-33 (IL-33), produced by lung cells after exposure to protease allergens, can induce innate-type airway eosinophilia by activating natural helper (NH) cells, a member of group 2 innate lymphoid cells (ILC2), to secrete Th2 type-cytokines. Because IL-33 also can induce mast cells (MCs) to secrete Th2 type-cytokines, MCs are thought to cooperate with NH cells in enhancing protease or IL-33-mediated innate-type airway eosinophilia. However, we found that MC-deficient KitW-sh/W-sh mice exhibited exacerbated protease-induced lung inflammation associated with reduced numbers of regulatory T (Treg) cells. Moreover, IL-2 produced by IL-33-stimulated MCs promoted expansion of numbers of Treg cells, thereby suppressing development of papain- or IL-33-induced airway eosinophilia. We have thus identified a unique anti-inflammatory pathway that can limit induction of innate-type allergic airway inflammation mediated by NH cells. PMID:26200013

  13. Glycogen synthase kinase-3 controls IL-10 expression in CD4+ effector T-cell subsets through epigenetic modification of the IL-10 promoter

    PubMed Central

    Hill, Elaine V; Ng, T H Sky; Burton, Bronwen R; Oakley, Charly M; Malik, Karim; Wraith, David C

    2015-01-01

    The serine/threonine kinase glycogen synthase kinase-3 (GSK3) plays an important role in balancing pro- and anti-inflammatory cytokines. We have examined the role of GSK3 in production of IL-10 by subsets of CD4+ T helper cells. Treatment of naive murine CD4+ T cells with GSK3 inhibitors did not affect their production of IL-10. However, treatment of Th1 and Th2 cells with GSK3 inhibitors dramatically increased production of IL-10. GSK3 inhibition also led to upregulation of IL-10 among Th1, Th2, and Th17 subsets isolated from human blood. The encephalitogenic potential of GSK3 inhibitor treated murine Th1 cells was significantly reduced in adoptive transfer experiments by an IL-10-dependent mechanism. Analysis of the murine IL-10 promoter in response to inhibition of GSK3 in Th1 cells showed modification to a transcriptionally active state indicated by changes in histone H3 acetylation and methylation. Additionally, GSK3 inhibition increased expression of the transcription factors c-Maf, Nfil3, and GATA3, correlating with the increase in IL-10. These findings are important in the context of autoimmune disease since they show that it is possible to reprogram disease-causing cells through GSK3 inhibition. PMID:25627813

  14. An asthma-associated IL4R variant exacerbates airway inflammation by promoting conversion of regulatory T cells to TH17-like cells.

    PubMed

    Massoud, Amir Hossein; Charbonnier, Louis-Marie; Lopez, David; Pellegrini, Matteo; Phipatanakul, Wanda; Chatila, Talal A

    2016-09-01

    Mechanisms by which regulatory T (Treg) cells fail to control inflammation in asthma remain poorly understood. We show that a severe asthma-associated polymorphism in the gene encoding the interleukin (IL)-4 receptor alpha chain (Il4ra(R576)) promotes conversion of induced Treg (iTreg) cells toward a T helper 17 (TH17) cell fate. This skewing is mediated by the recruitment by IL-4Rα(R576) of the growth-factor-receptor-bound protein 2 (GRB2) adaptor protein, which drives IL-17 expression by activating a pathway that involves extracellular-signal-regulated kinase, IL-6 and the transcription factor STAT3. Treg cell-specific deletion of genes that regulate TH17 cell differentiation, including Il6ra and RAR-related orphan receptor gamma (Rorc), but not of Il4 or Il13, prevented exacerbated airway inflammation in mice expressing Il4ra(R576) (hereafter referred to as Il4ra(R576) mice). Furthermore, treatment of Il4ra(R576) mice with a neutralizing IL-6-specific antibody prevented iTreg cell reprogramming into TH17-like cells and protected against severe airway inflammation. These findings identify a previously unknown mechanism for the development of mixed TH2-TH17 cell inflammation in genetically prone individuals and point to interventions that stabilize iTreg cells as potentially effective therapeutic strategies. PMID:27479084

  15. An Interleukin-33-Mast Cell-Interleukin-2 Axis Suppresses Papain-Induced Allergic Inflammation by Promoting Regulatory T Cell Numbers.

    PubMed

    Morita, Hideaki; Arae, Ken; Unno, Hirotoshi; Miyauchi, Kousuke; Toyama, Sumika; Nambu, Aya; Oboki, Keisuke; Ohno, Tatsukuni; Motomura, Kenichiro; Matsuda, Akira; Yamaguchi, Sachiko; Narushima, Seiko; Kajiwara, Naoki; Iikura, Motoyasu; Suto, Hajime; McKenzie, Andrew N J; Takahashi, Takao; Karasuyama, Hajime; Okumura, Ko; Azuma, Miyuki; Moro, Kazuyo; Akdis, Cezmi A; Galli, Stephen J; Koyasu, Shigeo; Kubo, Masato; Sudo, Katsuko; Saito, Hirohisa; Matsumoto, Kenji; Nakae, Susumu

    2015-07-21

    House dust mite-derived proteases contribute to allergic disorders in part by disrupting epithelial barrier function. Interleukin-33 (IL-33), produced by lung cells after exposure to protease allergens, can induce innate-type airway eosinophilia by activating natural helper (NH) cells, a member of group 2 innate lymphoid cells (ILC2), to secrete Th2 type-cytokines. Because IL-33 also can induce mast cells (MCs) to secrete Th2 type-cytokines, MCs are thought to cooperate with NH cells in enhancing protease or IL-33-mediated innate-type airway eosinophilia. However, we found that MC-deficient Kit(W-sh/W-sh) mice exhibited exacerbated protease-induced lung inflammation associated with reduced numbers of regulatory T (Treg) cells. Moreover, IL-2 produced by IL-33-stimulated MCs promoted expansion of numbers of Treg cells, thereby suppressing development of papain- or IL-33-induced airway eosinophilia. We have thus identified a unique anti-inflammatory pathway that can limit induction of innate-type allergic airway inflammation mediated by NH cells. PMID:26200013

  16. A clinical grade poly I:C-analogue (Ampligen) promotes optimal DC maturation and Th1-type T cell responses of healthy donors and cancer patients in vitro.

    PubMed

    Navabi, H; Jasani, B; Reece, A; Clayton, A; Tabi, Z; Donninger, C; Mason, M; Adams, M

    2009-01-01

    Maturation of dendritic cells (DC) can be triggered in vitro by inflammatory cytokines or Toll-like receptor (TLR) ligands such as CpG or polyI:C. Corresponding, well-characterized agents which can be applied in clinical settings are sparse. We have evaluated a clinical grade, non-toxic analogue of polyI:C, poly(I:C12U) (Ampligen), as a potential adjuvant for cancer immunotherapy, for its ability to drive maturation of human myeloid DC. Our results provide evidence that poly(I:C12U) is effective in inducing optimal phenotypic (elevated levels of MHC-Class I/Class II, CD83, CCR7, CD86 and CD40 molecules) and functional maturation of human DC in vitro, capable of promoting the production of the inflammatory (Th1-type) cytokine IL-12, with significantly lower levels of IL-10 production, compared to that induced by the parent compound polyI:C. Importantly, poly(I:C12U) has a comparable effect on the maturation and function of DC derived either from healthy donors or cancer patients indicating that it is able to overcome any immune suppressive factors associated with the tumour bearing state. These characteristics make poly(I:C12U) a suitable agent for use as an adjuvant in cancer directed immunotherapeutic regimes. PMID:18977262

  17. APRIL modulates B and T cell immunity

    PubMed Central

    Stein, Jens V.; López-Fraga, Marta; Elustondo, Fernando A.; Carvalho-Pinto, Carla E.; Rodríguez, Dolores; Gómez-Caro, Ruth; de Jong, Joan; Martínez-A, Carlos; Medema, Jan Paul; Hahne, Michael

    2002-01-01

    The TNF-like ligands APRIL and BLyS are close relatives and share the capacity to bind the receptors TACI and BCMA. BLyS has been shown to play an important role in B cell homeostasis and autoimmunity, but the biological role of APRIL remains less well defined. Analysis of T cells revealed an activation-dependent increase in APRIL mRNA expression. We therefore generated mice expressing APRIL as a transgene in T cells. These mice appeared normal and showed no signs of B cell hyperplasia. Transgenic T cells revealed a greatly enhanced survival in vitro as well as enhanced survival of staphylococcal enterotoxin B–reactive CD4+ T cells in vivo, which both directly correlate with elevated Bcl-2 levels. Analysis of humoral responses to T cell–dependent antigens in the transgenic mice indicated that APRIL affects only IgM but not IgG responses. In contrast, T cell–independent type 2 (TI-2) humoral response was enhanced in APRIL transgenic mice. As TACI was previously reported to be indispensable for TI-2 antibody formation, these results suggest a role for APRIL/TACI interactions in the generation of this response. Taken together, our data indicate that APRIL is involved in the induction and/or maintenance of T and B cell responses. PMID:12070306

  18. Purification of melanoma reactive T cell by using a monocyte-based solid phase T-cell selection system for adoptive therapy.

    PubMed

    Li, Jongming; Mookerjee, Bijoyesh; Wagner, John

    2008-01-01

    The generation of melanoma-reactive T cells with the characteristics necessary for in vivo effectiveness remains a considerable obstacle to the application of adoptive cell therapy. Recent clinical success with adoptive cell therapy for melanoma is motivating additional investigation to improve the technology of generating such tumor reactive lymphocytes. Here we describe a novel solid phase T-cell selection system, in which monocytes are immobilized on solid support for antigen-specific T-cell purification. We hypothesized and proved that antigen-specific T cells recognize their cognate antigens and bind to them faster than nonantigen-specific T cells and are concentrated on the surface after removing the nonadherent cells by washing. Moreover, activated antigen-specific T cells proliferated more rapidly than nonspecific T cells, further increasing the frequency and purity of antigen-specific T cells. Optimal selection times for Melan-A-specific T cells are studied. Our data demonstrated that T-cell selection can usually increase the frequency of tumor antigen-specific T cells by >10-fold, whereas T-cell expansion after the selection boost the frequency of tumor antigen-specific T cells by another approximately 10-fold. More importantly, these T cells are generated under more physiologic conditions. This new T-cell selection system is superior to traditional repeated stimulation methods in generating tumor antigen-specific T cells for adoptive cell immunotherapy. This inexpensive and simple T-cell selection system can produce large quantity of highly purified Melan-A-specific T cells within 2 weeks after T-cell activation. PMID:18157015

  19. Mechanisms of T cell organotropism.

    PubMed

    Fu, Hongmei; Ward, Eleanor Jayne; Marelli-Berg, Federica M

    2016-08-01

    Protective immunity relies upon T cell differentiation and subsequent migration to target tissues. Similarly, immune homeostasis requires the localization of regulatory T cells (Tregs) to the sites where immunity takes place. While naïve T lymphocytes recirculate predominantly in secondary lymphoid tissue, primed T cells and activated Tregs must traffic to the antigen rich non-lymphoid tissue to exert effector and regulatory responses, respectively. Following priming in draining lymph nodes, T cells acquire the 'homing receptors' to facilitate their access to specific tissues and organs. An additional level of topographic specificity is provided by T cells receptor recognition of antigen displayed by the endothelium. Furthermore, co-stimulatory signals (such as those induced by CD28) have been shown not only to regulate T cell activation and differentiation, but also to orchestrate the anatomy of the ensuing T cell response. We here review the molecular mechanisms supporting trafficking of both effector and regulatory T cells to specific antigen-rich tissues. PMID:27038487

  20. Identification of procollagen promoter DNA-binding proteins: effects of dexamethasone

    SciTech Connect

    Sweeney, C.; Cutroneo, K.R.

    1987-05-01

    Glucocorticoids selectively decrease procollagen synthesis by decreasing procollagen mRNA transcription. Dexamethasone coordinately decreased total cellular type I and type III procollagen mRNAs in mouse embryonic skin fibroblasts. Since sequence specific DNA-binding proteins are known to modulate eukaryotic gene expression the authors identified in mouse fibroblasts nuclear proteins which bind to types I and III procollagen promoter DNAs. Nuclear proteins were electrophoresed, blotted onto nitrocellulose and probed with /sup 32/P-end-labeled type I and type III procollagen promoter DNAs in the presence of equimolar amounts of /sup 32/P-end-labeled vector DNA. Differences in total DNA binding were noted by the densitometric scans of the nuclear proteins. Dexamethasone treatment enhanced total DNA binding. Increasing the NaCl concentration decreased the number of promoter DNA-binding proteins without altering the relative specificity for the promoter DNAs. Promoter DNA binding to nuclear proteins was also inhibited by increasing concentrations of E. coli DNA. The number of DNA-binding proteins was greater for type III procollagen promoter DNA. The effect of dexamethasone treatment on promoter DNA binding to nuclear proteins was determined.

  1. The Effects of TLR Activation on T-Cell Development and Differentiation

    PubMed Central

    Jin, Bo; Sun, Tao; Yu, Xiao-Hong; Yang, Ying-Xiang; Yeo, Anthony E. T.

    2012-01-01

    Invading pathogens have unique molecular signatures that are recognized by Toll-like receptors (TLRs) resulting in either activation of antigen-presenting cells (APCs) and/or costimulation of T cells inducing both innate and adaptive immunity. TLRs are also involved in T-cell development and can reprogram Treg cells to become helper cells. T cells consist of various subsets, that is, Th1, Th2, Th17, T follicular helper (Tfh), cytotoxic T lymphocytes (CTLs), regulatory T cells (Treg) and these originate from thymic progenitor thymocytes. T-cell receptor (TCR) activation in distinct T-cell subsets with different TLRs results in differing outcomes, for example, activation of TLR4 expressed in T cells promotes suppressive function of regulatory T cells (Treg), while activation of TLR6 expressed in T cells abrogates Treg function. The current state of knowledge of regarding TLR-mediated T-cell development and differentiation is reviewed. PMID:22737174

  2. Expression of the human T-cell receptor V beta 5.3 in Escherichia coli by thermal induction of the trc promoter: nucleotide sequence of the lacIts gene.

    PubMed

    Adari, H; Andrews, B; Ford, P J; Hannig, G; Brosius, J; Makrides, S C

    1995-11-01

    We have constructed a vector, pKBi, for the high-level expression of the variable beta chain 5.3 (V beta 5.3) of the human T-cell receptor in Escherichia coli. This vector incorporates the trc promoter, a polylinker, two transcription terminators, and the tetracycline resistance gene. Furthermore, the vector contains the lacIts gene that encodes a temperature-sensitive (ts) lac repressor, thus obviating both the need to use IPTG as a transcriptional inducer, and bacterial strains that harbor either the lacI or lacIq genes. The sequence of the lacIts gene shows an open reading frame of 1,080 nucleotides encoding 360 amino acids, and differs from the lacI gene at nucleotide 559 (with reference to the first nucleotide of the start codon). This nucleotide changes from G to A, causing amino acid residue 187 to change from glycine (GGC) to serine (AGC). This mutation imparts thermal sensitivity to the lac repressor protein. This is the first time that a TCR V beta region has been expressed at high levels (up to 28 mg/liter of culture) without fusion partners. The availability of the lacIts gene for thermal induction of the trc promoter, and the presence of the tetracycline resistance gene should make the expression vector pKBi particularly attractive for the efficient production of human therapeutic proteins in bacteria. PMID:7576181

  3. The role of bovine γδ T cells and their WC1 co-receptor in response to bacterial pathogens and promoting vaccine efficacy: a model for cattle and humans.

    PubMed

    Baldwin, Cynthia L; Hsu, Haoting; Chen, Chuang; Palmer, Mitchell; McGill, Jodi; Waters, W Ray; Telfer, Janice C

    2014-06-15

    γδ T cells are critical to immune surveillance and protection since they are found as resident cells in many organs and tissues, including in humans and ruminants, and circulate at substantial numbers in the blood. It is known that γδ T cells contribute to cellular immunity and protection against important pathogens including organizing granulomas in response to Mycobacteria. We have shown that IFNγ-producing bovine γδ T cells bearing the WC1 co-receptor are the major cell population responding in recall responses to Leptospira during the first month following priming by vaccination against serovar Hardjo. To date, successful vaccines largely include those to diseases that only require antibody responses for protection and attempts at creating subunit peptide vaccines to stimulate conventional αβ T cells for cellular immune responses have been mostly unsuccessful. However, activation of nonconventional T cells, such as γδ T cells that direct adaptive T cell responses, has received little attention for improving vaccines because it is not clear how best to prime γδ T cells for recall responses. Annotation of the bovine genome showed there were 13 WC1 molecules coded for by individual genes. This gene number is conserved among breeds and individuals and expression of the WC1 molecules are distributed among cells to form a number of γδ T cell subsets. Using RNA silencing, we have shown that the WC1 co-receptor contributes to the ability of γδ T cells to respond to Leptospira spp. The Leptospira-responsive γδ T cells are found within a subset of the serologically defined WC1.1(+) γδ T cell subpopulation and our data indicate that the WC1 molecules expressed act as pattern recognition receptors interacting directly with bacterial components. We are now extending this work to Mycobacteria bovis. PMID:24726858