Dharmesh D. Desai

RAGE Ligation Affects T Cell Activation and Controls T Cell Differentiation

The pattern recognition receptor, RAGE, has been shown to be involved in adaptive immune responses but its role on the components of these responses is not well understood. We have studied the effects of a small molecule inhibitor of RAGE and the deletion of the receptor (RAGE−/− mice) on T cell responses involved in autoimmunity and allograft rejection. Syngeneic islet graft and islet allograft rejection was reduced in NOD and B6 mice treated with TTP488, a small molecule RAGE inhibitor (p < 0.001). RAGE−/− mice with streptozotocin-induced diabetes showed delayed rejection of islet allografts compared with wild type (WT) mice (p < 0.02). This response in vivo correlated with reduced proliferative responses of RAGE−/− T cells in MLRs and in WT T cells cultured with TTP488. Overall T cell proliferation following activation with anti-CD3 and anti-CD28 mAbs were similar in RAGE−/− and WT cells, but RAGE−/− T cells did not respond to costimulation with anti-CD28 mAb. Furthermore, culture supernatants from cultures with anti-CD3 and anti-CD28 mAbs showed higher levels of IL-10, IL-5, and TNF-α with RAGE−/− compared with WT T cells, and WT T cells showed reduced production of IFN-γ in the presence of TTP488, suggesting that RAGE may be important in the differentiation of T cell subjects. Indeed, by real-time PCR, we found higher levels of RAGE mRNA expression on clonal T cells activated under Th1 differentiating conditions. We conclude that activation of RAGE on T cells is involved in early events that lead to differentiation of Th1+ T cells.

Fcγ Receptor IIB on Dendritic Cells Enforces Peripheral Tolerance by Inhibiting Effector T Cell Responses

The uptake of immune complexes by FcRs on APCs augments humoral and cellular responses to exogenous Ag. In this study, CD11c+ dendritic cells are shown to be responsible in vivo for immune complex-triggered priming of T cells. We examine the consequence of Ab-mediated uptake of self Ag by dendritic cells in the rat insulin promoter-membrane OVA model and identify a role for the inhibitory FcγRIIB in the maintenance of peripheral CD8 T cell tolerance. Effector differentiation of diabetogenic OT-I CD8+ T cells is enhanced in rat insulin promoter-membrane OVA mice lacking FcγRIIB, resulting in a high incidence of diabetes. FcγRIIB-mediated inhibition of CD8 T cell priming results from suppression of both DC activation and cross-presentation through activating FcγRs. Further FcγRIIB on DCs inhibited the induction of OVA-specific Th1 effectors, limiting Th1-type differentiation and memory T cell accumulation. In these MHC II-restricted responses, the presence of FcγRIIB only modestly affected initial CD4 T cell proliferative responses, suggesting that FcγRIIB limited effector cell differentiation primarily by inhibiting DC activation. Thus, FcγRIIB can contribute to peripheral tolerance maintenance by inhibiting DC activation alone or by also limiting processing of exogenously acquired Ag.

Antibody-enhanced cross-presentation of self antigen breaks T cell tolerance.

We have developed a model of autoimmunity to investigate autoantibody-mediated cross-presentation of self antigen. RIP-mOVA mice, expressing OVA in pancreatic beta cells, develop severe autoimmune diabetes when given OT-I cells (OVA-specific CD8(+) T cells) and anti-OVA IgG but not when given T cells alone. Anti-OVA IgG is not directly injurious to the islets but rather enhances cross-presentation of apoptotic islet antigen to the OT-I cells, leading to their differentiation into potent effector cells. Antibody-driven effector T cell activation is dependent on the presence of activating Fc receptors for IgG (FcgammaRs) and cross-priming DCs. As a consequence, diabetes incidence and severity was reduced in mice lacking activating FcgammaRs. An intact complement pathway was also required for disease development, as C3 deficiency was also partially protective. C3-deficient animals exhibited augmented T cell priming overall, indicating a proinflammatory role for complement activation after the T cell priming phase. Thus, we show that autoreactive antibody can potently enhance the activation of effector T cells in response to cross-presented self antigen, thereby contributing to T cell-mediated autoimmunity.

Dominant Expression of the Inhibitory FcγRIIB Prevents Antigen Presentation by Murine Plasmacytoid Dendritic Cells

Plasmacytoid dendritic cells (pDCs) are key regulators of the innate immune response, yet their direct role as APCs in the adaptive immune response is unclear. We found that unlike conventional DCs, immune complex (IC) exposed murine pDCs neither up-regulated costimulatory molecules nor activated Ag-specific CD4+ and CD8+ T cells. The inability of murine pDCs to promote T cell activation was due to inefficient proteolytic processing of internalized ICs. This defect in the IC processing capacity of pDCs results from a lack of activating FcγR expression (FcγRI, III, IV) and the dominant expression of the inhibitory receptor FcγRIIB. Consistent with this idea, transgenic expression of the activating human FcγRIIA gene, not present in the mouse genome, recapitulated the human situation and rescued IC antigenic presentation capacity by murine pDCs. The selective expression of FcγRIIB by murine pDCs was not strain dependent and was maintained even following stimulation with TLR ligands and inflammatory cytokines. The unexpected difference between the mouse and human in the expression of activating/inhibitory FcγRs has implications for the role of pDCs in Ab-modulated autoimmunity and anti-viral immunity.

T CELLS SPECIFIC FOR DNA-BINDING PEPTIDES

The progressive selection for native DNA (dsDNA) specificity as the autoimmune antibody response matures suggests that dsDNA or more likely dsDNA-protein complexes may be the immune stimulus for autoimmune DNA antibody production. In more recent experiments we have been able to confirm that complexes of DNA and a DNA-binding peptide Fusl will stimulate the production of antiDNA antibodies in mice not genetically prone to autoimmune disease.2,3 The DNA-specific antibody induced by immunisation with DNA-Fusl complexes has serological, structural, and immunopathogenic potential of autoimmune anti-DNA antibody. The Fus 1 peptide is 27 amino acids long, highly basic, and has the structural characteristics of a ’zinc-finger’ DNA-binding motif. The sequence for Fus 1 represents an internal amino acid sequence from the 52 amino acid carboxyl-extension protein (CEP) of ubiquitin in Trypanosoma cruzi.4 The CEP and the Fus peptide derived from CEP are highly conserved in all eukaryotes including mice and humans. Datta and colleagueS5 ’ have generated cloned helper T-cell lines from autoimmune mice specific for nucleosomal proteins. Such T cell lines are able to provide the necessary help for antibody production by DNAspecific B cells in vitro. In vivo such T cells stimulated