Viral immunity: cross-priming with the help of TLR3.
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Cross-presentation is important for regulating T-cell responses to exogenous antigens and can maintain tolerance (cross-tolerance) or induce immune responses (cross-priming). Recent exciting results on the role of the Toll-like receptor TLR3 in promoting cross-priming of viral antigens provide new insights into the mechanisms that allow Toll-like receptor signaling to bridge innate and adaptive immune responses. (+info)
Cutting edge: generation of splenic CD8+ and CD8- dendritic cell equivalents in Fms-like tyrosine kinase 3 ligand bone marrow cultures.
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We demonstrate that functional and phenotypic equivalents of mouse splenic CD8(+) and CD8(-) conventional dendritic cell (cDC) subsets can be generated in vitro when bone marrow is cultured with fms-like tyrosine kinase 3 (flt3) ligand. In addition to CD45RA(high) plasmacytoid DC, two distinct CD24(high) and CD11b(high) cDC subsets were present, and these subsets showed equivalent properties to splenic CD8(+) and CD8(-) cDC, respectively, in the following: 1) surface expression of CD11b, CD24, and signal regulatory protein-alpha; 2) developmental dependence on, and mRNA expression of, IFN regulatory factor-8; 3) mRNA expression of TLRs and chemokine receptors; 4) production of IL-12 p40/70, IFN-alpha, MIP-1alpha, and RANTES in response to TLR ligands; 5) expression of cystatin C; and 6) cross-presentation of exogenous Ag to CD8 T cells. Furthermore, despite lacking surface CD8 expression, the CD24(high) subset contained CD8 mRNA and up-regulated surface expression when transferred into mice. This culture system allows access to bona fide counterparts of the splenic DC subsets. (+info)
Rapid destruction of the tumor microenvironment by CTLs recognizing cancer-specific antigens cross-presented by stromal cells.
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A single tumor contains a heterogeneous population of cancer cells. Some cancer cells express antigens and are susceptible to specific CTLs. However, other cancer cells are antigen-loss variants (ALVs) that escape these CTLs because they express little or no antigen. Here, we show that antigen-specific T cells can eliminate ALVs when the parental population expresses a model gp33 antigen (KAVYNFATM) at a level sufficient to be locally cross-presented by the nonmalignant stromal cells. That is, the ALVs are eliminated as bystanders because the stroma is destroyed. ALVs escape bystander killing when the bone marrow-derived and/or non-bone marrow-derived stroma does not express the appropriate MHC or when the amount of antigen is too low for effective cross-presentation. The rapid destruction of the stroma, including bone marrow-derived as well as sessile components, and of the parental cancer cells, may be essential for the complete rejection of established tumors by preventing variant escape. (+info)
Mouse endothelial cells cross-present lymphocyte-derived antigen on class I MHC via a TAP1- and proteasome-dependent pathway.
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In vivo studies suggest that vascular endothelial cells (ECs) can acquire and cross-present exogenous Ag on MHC-I but the cellular mechanisms underlying this observation remain unknown. We tested whether primary female mouse aortic ECs could cross-present exogenous male Ag to the T cell hybridoma, MHH, specific for HYUty plus D(b). MHC-I-deficient male spleen cells provided a source of male Ag that could not directly stimulate the MHH cells. Addition of male but not female MHC-I-deficient spleen cells to wild-type syngeneic female EC induced MHH stimulation, demonstrating EC cross-presentation. Lactacystin treatment of the donor male MHC-I-deficient spleen cells, to inhibit proteasome function, markedly enhanced EC cross-presentation showing that the process is most efficient for intact proteins rather than degraded peptide fragments. Additional experiments revealed that this EC Ag-processing pathway is both proteasome and TAP1 dependent. These studies demonstrate that cultured murine aortic ECs can process and present MHC-I-restricted Ag derived from a separate, live cell, and they offer insight into the molecular requirements involved in this EC Ag presentation process. Through this pathway, ECs expressing cross-presented peptides can participate in the effector phase of T cell-mediated inflammatory responses such as autoimmunity, anti-tumor immunity, and transplant rejection. (+info)
Inefficient cross-presentation limits the CD8+ T cell response to a subdominant tumor antigen epitope.
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CD8(+) T lymphocytes (T(CD8)) responding to subdominant epitopes provide alternate targets for the immunotherapy of cancer, particularly when self-tolerance limits the response to immunodominant epitopes. However, the mechanisms that promote T(CD8) subdominance to tumor Ags remain obscure. We investigated the basis for the lack of priming against a subdominant tumor epitope following immunization of C57BL/6 (B6) mice with SV40 large tumor Ag (T Ag)-transformed cells. Immunization of B6 mice with wild-type T Ag-transformed cells primes T(CD8) specific for three immunodominant T Ag epitopes (epitopes I, II/III, and IV) but fails to induce T(CD8) specific for the subdominant T Ag epitope V. Using adoptively transferred T(CD8) from epitope V-specific TCR transgenic mice and immunization with T Ag-transformed cells, we demonstrate that the subdominant epitope V is weakly cross-presented relative to immunodominant epitopes derived from the same protein Ag. Priming of naive epitope V-specific TCR transgenic T(CD8) in B6 mice required cross-presentation by host APC. However, robust expansion of these T(CD8) required additional direct presentation of the subdominant epitope by T Ag-transformed cells and was only significant following immunization with T Ag-expressing cells lacking the immunodominant epitopes. These results indicate that limited cross-presentation coupled with competition by immunodominant epitope-specific T(CD8) contributes to the subdominant nature of a tumor-specific epitope. This finding has implications for vaccination strategies targeting T(CD8) responses to cancer. (+info)
Cross-presentation of the long-lived lymphocytic choriomeningitis virus nucleoprotein does not require neosynthesis and is enhanced via heat shock proteins.
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Many viral proteins that contain MHC class I-restricted peptides are long-lived, and it is elusive how they can give rise to class I epitopes. Recently, we showed that direct presentation of an epitope of the long-lived lymphocytic choriomeningitis virus nucleoprotein (LCMV-NP) required neosynthesis in accordance with the defective ribosomal products hypothesis. In this study, we report that LCMV-NP can be cross-primed in mice using either LCMV-NP-transfected human HEK293 or BALB/c-derived B8 cells as Ag donor cells. In addition, we establish that contrary to direct presentation, cross-presentation required accumulation of the mature LCMV-NP and could not be sustained by the newly synthesized LCMV-NP protein, intermediate proteasomal degradation products, or the minimal NP396 epitope. Nevertheless, NP cross-presentation was enhanced by heat shock and was blunted by inhibitors of heat shock protein 90 and gp96. We propose that cross-presentation has evolved to sustain the presentation of stable viral proteins when their neosynthesis has ceased in infected donor cells. (+info)
Dendritic cell-mediated cross-presentation of antigens derived from colon carcinoma cells exposed to a highly cytotoxic multidrug regimen with gemcitabine, oxaliplatin, 5-fluorouracil, and leucovorin, elicits a powerful human antigen-specific CTL response with antitumor activity in vitro.
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Gemcitabine, oxaliplatin, leucovorin, and 5-fluorouracil (GOLF) is a novel multidrug regimen inducing high levels of necrosis and apoptosis in colon carcinoma cells. This regimen is also able to promote a process of Ag remodeling including up-regulation of immunotherapy targets like carcinoembryonic Ag (CEA), thymidylate synthase (TS). We have conducted a preclinical study aimed to investigate whether these drug-induced modifications would also enhance colon cancer cell immunogenicity. Several CTL lines were thus generated by in vitro stimulating human HLA-A(*)02.01(+) PBMCs, from normal donors and colon cancer patients, with autologous dendritic cells cross-primed with cell lysates of colon cancer cells untreated, irradiated, or previously exposed to different drug treatments including the GOLF regimen. Class I HLA-restricted cytolytic activity of these CTL lines was tested against colon cancer cells and CEA and TS gene transfected target cells. These experiments revealed that CTLs sensitized with GOLF-treated cancer cells were much more effective than those sensitized with the untreated colon carcinoma cells or those exposed to the other treatments. CTL lines sensitized against the GOLF-treated colon cancer cells, also expressed a greater percentage of T-lymphocyte precursors able to recognize TS- and CEA-derived peptides. These results suggest that GOLF regimen is a powerful antitumor and immunomodulating regimen that can make the tumor cells a suitable means to induce an Ag-specific CTL response. These results suggest that a rationale combination of GOLF chemotherapy with cytokine-based immunotherapy could generate a chemotherapy-modulated Ag-specific T-lymphocyte response in cancer patients able to destroy the residual disease survived to the cytotoxic drugs. (+info)
Enhancement of dendritic cell antigen cross-presentation by CpG DNA involves type I IFN and stabilization of class I MHC mRNA.
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Dendritic cells (DCs) internalize exogenous Ags and process them for cross-presentation by class I MHC (MHC-I) to CD8+ T cells. This processing can occur by transporter for Ag presentation (TAP)-dependent or TAP-independent mechanisms. We observed that CpG DNA enhanced cross-presentation of Ags by Flt-3L-cultured bone marrow-derived murine DCs by a type I IFN (IFN-alphabeta)-dependent mechanism. Myeloid DCs provided cross-presentation function in this system. Both TAP1 knockout and wild-type DCs showed enhanced cross-presentation when treated with CpG DNA at 26 degrees C, demonstrating that TAP is not essential to this regulatory mechanism, although TAP is an important determinant of MHC-I expression. Enhancement of cross-processing by CpG DNA did not involve increased Ag uptake or proteolysis but did correlate with IFN-alphabeta-dependent increases in expression of MHC-I mRNA and protein. Increased MHC-I mRNA levels resulted in part from stabilization of MHC-I mRNA, a novel posttranscriptional mechanism for regulation of MHC-I expression. Thus, a major mechanism by which CpG oligodeoxynucleotide increase cross presentation by DCs appears to be an IFN-alphabeta-mediated increase in MHC-I synthesis. (+info)