Transport of Streptococcus pneumoniae Capsular Polysaccharide in MHC Class II Tubules

Bacterial capsular polysaccharides are virulence factors and are considered T cell–independent antigens. However, the capsular polysaccharide Sp1 from Streptococcus pneumoniae serotype 1 has been shown to activate CD4(+) T cells in a major histocompatibility complex (MHC) class II–dependent manner. The mechanism of carbohydrate presentation to CD4(+) T cells is unknown. We show in live murine dendritic cells (DCs) that Sp1 translocates from lysosomal compartments to the plasma membrane in MHCII-positive tubules. Sp1 cell surface presentation results in reduction of self-peptide presentation without alteration of the MHCII self peptide repertoire. In DM-deficient mice, retrograde transport of Sp1/MHCII complexes resulting in T cell–dependent immune responses to the polysaccharide in vitro and in vivo is significantly reduced. The results demonstrate the capacity of a bacterial capsular polysaccharide antigen to use DC tubules as a vehicle for its transport as an MHCII/saccharide complex to the cell surface for the induction of T cell activation. Furthermore, retrograde transport requires the functional role of DM in self peptide–carbohydrate exchange. These observations open new opportunities for the design of vaccines against microbial encapsulated pathogens

Streptococcus pneumoniae Serotype 1 Capsular Polysaccharide Induces CD8+CD28− Regulatory T Lymphocytes by TCR Crosslinking

Zwitterionic capsular polysaccharides (ZPS) of commensal bacteria are characterized by having both positive and negative charged substituents on each repeating unit of a highly repetitive structure that has an α-helix configuration. In this paper we look at the immune response of CD8+ T cells to ZPSs. Intraperitoneal application of the ZPS Sp1 from Streptococcus pneumoniae serotype 1 induces CD8+CD28− T cells in the spleen and peritoneal cavity of WT mice. However, chemically modified Sp1 (mSp1) without the positive charge and resembling common negatively charged polysaccharides fails to induce CD8+CD28− T lymphocytes. The Sp1-induced CD8+CD28− T lymphocytes are CD122lowCTLA-4+CD39+. They synthesize IL-10 and TGF-β. The Sp1-induced CD8+CD28− T cells exhibit immunosuppressive properties on CD4+ T cells in vivo and in vitro. Experimental approaches to elucidate the mechanism of CD8+ T cell activation by Sp1 demonstrate in a dimeric MHC class I-Ig model that Sp1 induces CD8+ T cell activation by enhancing crosslinking of TCR. The expansion of CD8+CD28− T cells is independent, of direct antigen-presenting cell/T cell contact and, to the specificity of the T cell receptor (TCR). In CD8+CD28− T cells, Sp1 enhances Zap-70 phosphorylation and increasingly involves NF-κB which ultimately results in protection versus apoptosis and cell death and promotes survival and accumulation of the CD8+CD28− population. This is the first description of a naturally occurring bacterial antigen that is able to induce suppressive CD8+CD28− T lymphocytes in vivo and in vitro. The underlying mechanism of CD8+ T cell activation appears to rely on enhanced TCR crosslinking. The data provides evidence that ZPS of commensal bacteria play an important role in peripheral tolerance mechanisms and the maintenance of the homeostasis of the immune system

HLA-DR Alpha 2 Mediates Negative Signalling via Binding to Tirc7 Leading to Anti-Inflammatory and Apoptotic Effects in Lymphocytes In Vitro and In Vivo

Classically, HLA-DR expressed on antigen presenting cells (APC) initiates lymphocyte activation via presentation of peptides to TCR bearing CD4+ T-Cells. Here we demonstrate that HLA-DR alpha 2 domain (sHLA-DRα2) also induces negative signals by engaging TIRC7 on lymphocytes. This interaction inhibits proliferation and induces apoptosis in CD4+ and CD8+ T-cells via activation of the intrinsic pathway. Proliferation inhibition is associated with SHP-1 recruitment by TIRC7, decreased phosphorylation of STAT4, TCR-ζ chain & ZAP70, and inhibition of IFN-γ and FasL expression. HLA-DRα2 and TIRC7 co-localize at the APC-T cell interaction site. Triggering HLA-DR - TIRC7 pathway demonstrates that sHLA-DRα2 treatment inhibits proinflammatory-inflammatory cytokine expression in APC & T cells after lipopolysaccaride (LPS) stimulation in vitro and induces apoptosis in vivo. These results suggest a novel antiproliferative role for HLA-DR mediated via TIRC7, revise the notion of an exclusive stimulatory interaction of HLA-DR with CD4+ T cells and highlights a novel physiologically relevant regulatory pathway

Effect of B7-2 and CD40 Signals from Activated Antigen-Presenting Cells on the Ability of Zwitterionic Polysaccharides To Induce T-Cell Stimulation

Carbohydrates have been thought to stimulate immune responses independently of T cells; however, zwitterionic polysaccharides (ZPSs) from the capsules of some bacteria elicit potent CD4(+)-T-cell responses in vivo and in vitro. We demonstrated that HLA-DR on professional antigen-presenting cells (APCs) is required for ZPS-induced T-cell proliferation in vitro (15). Recently, it was shown that ZPSs are processed to low-molecular-weight carbohydrates by a nitric oxide-mediated mechanism in endosomes and locate in the major histocompatibility complex class II pathway (5, 15). The effect of the ZPS-mediated expression of HLA-DR and costimulatory molecules on the APC and T-cell engagement and subsequent T-cell activation has not been elucidated. Herein, we report that ZPS-mediated induction of HLA-DR-surface expression and T-cell proliferation are maximally enhanced after incubation of APCs for 8 h with ZPS. Treatment of APCs with bafilomycin A inhibits the up-regulation of ZPS-mediated HLA-DR surface expression and leads to inhibition of T-cell proliferation. Monoclonal antibodies (MAbs) to the costimulatory molecules B7-2 and CD40L specifically block ZPS-mediated T-cell activation, while a MAb to B7-1 does not. Surface expression of B7-2 and B7-1 but not of CD40 is maximally enhanced at 8 to 16 h of treatment of APCs with ZPS. The results demonstrate that the cellular immune response to ZPS depends on the translocation of HLA-DR to the cell surface and requires costimulation via B7-2 and CD40 on activated APCs. The implication is that activation of ZPS-specific T cells requires an orchestrated arrangement of both presenting and costimulatory molecules to form an immunological synapse

Interleukin-6 is essential for zwitterionic polysaccharide-mediated abscess formation

Abscess formation associated with secondary peritonitis causes severe morbidity and can be fatal. Formation of abscesses requires the presence of CD4(+) T-cells. Zwitterionic polysaccharides (ZPSs) represent a novel class of immunomodulatory bacterial antigens that stimulate CD4(+) T-cells in a major histocompatibility complex (MHC) class II-dependent manner. The capsular polysaccharide Sp1 of Streptococcus pneumoniae serotype 1 possesses a zwitterionic charge with free amino groups and promotes T-cell-dependent abscess formation in an experimental mouse model. So far, nothing is known about the function of Interleukin (IL)-6 in intraperitoneal abscess formation. Here, we demonstrate that macrophages and dendritic cells (DCs), the most prevalent professional antigen-presenting cells involved in the formation of abscesses, secrete Interleukin (IL)-6 and are incorporated in the abscess capsule. Sp1 inhibits apoptosis of CD4(+) T-cells and causes IL-17 expression by CD4(+) T-cells in an IL-6-dependent manner. Abrogation of the Sp1-induced pleiotropic effects of IL-6 in IL-6-deficient mice and mice treated with an IL-6- specific neutralizing antibody results in significant inhibition of abscess formation. The data delineate the essential role of IL-6 in the linkage of innate and adaptive immunity in polysaccharide-mediated abscess formation

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

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