Neutrophil-dependent tumor rejection and priming of tumoricidal CD8+ T cell response induced by dendritic cells overexpressing CD95L.

Overexpression of CD95 (Fas/Apo-1) ligand (CD95L) has been shown to induce T cell tolerance but also, neutrophilic inflammation and rejection of allogeneic tissue. We explored the capacity of dendritic cells (DCs) genetically engineered to overexpress CD95L to induce an antitumor response. We first found that DCs overexpressing CD95L, in addition to MHC class I-restricted OVA peptides (CD95L-OVA-DCs), induced increased antigen-specific CD8(+) T cell responses as compared with DCs overexpressing OVA peptides alone. The enhanced T cell responses were associated with improved regression of a tumor expressing OVA, allowing survival of all animals. When DCs overexpressing CD95L (CD95L-DCs) were injected with the tumor expressing OVA, in vivo tumor proliferation was strikingly inhibited. A strong cellular apoptosis and a massive neutrophilic infiltrate developed in this setting. Neutrophil depletion prevented tumor regression as well as enhanced IFN-gamma production induced by CD95L-OVA-DCs. Furthermore, the CD8(+) T cell response induced by the coadministration of tumor cells and CD95L-DCs led to rejection of a tumor implanted at a distance from the DC injection site. In summary, DCs expressing CD95L promote tumor rejection involving neutrophil-mediated innate immunity and CD8(+) T cell-dependent adaptative immune responses.

Interferon regulatory factor 3 is involved in Toll-like receptor 4 (TLR4)- and TLR3-induced IL-12p35 gene activation.

Interleukin-12 (IL-12) is a heterodimeric cytokine produced by dendritic cells (DCs) in response to Toll-like receptor (TLR) ligation. While the mechanisms regulating IL-12p40 chain gene expression are well characterized, molecular events involved in IL-12p35 chain gene activation remain to be clarified. Since IL-12p35 mRNA was induced in human DCs activated through TLR3 or TLR4 but not TLR2, we investigated the potential role of interferon regulatory factor 3 (IRF-3) in IL-12p35 gene transactivation. First, a binding site for IRF-3 named interferon-stimulated response element-1 (ISRE-1) was identified in the human IL-12p35 promoter region between nucleotides -251 and -240. The ISRE-1 site was required for IL-12p35 gene activation in RAW 264.7 cells stimulated by lipopolysaccharide (LPS) or PolyI:C. Ectopic expression of IRF-3 was found to up-regulate IL-12p35 gene activation in the same system. Furthermore, chromatin immunoprecipitation (ChIP) studies demonstrated that IRF-3 is recruited to ISRE-1 site in TLR4- or TLR3-stimulated human DCs. Finally, experiments on DCs from IRF-3-deficient mice established that TLR4-induced IL-12p35 mRNA and IL-12p70 synthesis are impaired in absence of IRF-3. We conclude that IRF-3 binds to a critical cis-acting element in the IL-12p35 gene promoter and thereby represents a key factor for the induction of IL-12p70 synthesis in DCs.

Regulatory role of host CD8+ T lymphocytes in experimental graft-versus-host disease across a single major histocompatibility complex class II incompatibility.

BACKGROUND: CD8+ T cells are known to regulate type 2 helper T cell (Th2) alloreactive immune responses but their mode of activation is unclear. We investigated the role of host CD8+ T cells in experimental Th2-type graft-versus-host disease (GVHD) where donor/recipient disparity is restricted to a single major histocompatibility complex (MHC) class II antigen. METHODS: Immunoglobulin (Ig) E serum levels, eosinophilia and lymphoid tissue hyperplasia were compared after injection of bm12 CD4+ T cells in either wild-type or CD8+ T cell-deficient (CD8-/-) C57BL/6 mice. In vitro, we explored effects of the addition of CD8+ T cells from wild-type or IFN-gamma-/- mice in mixed leukocyte cultures prepared with beta2 microglobulin-deficient (beta2m-/-) CD4+ T cells as responders or beta2m dendritic cells as stimulators. RESULTS: HyperIgE resolved after 3 weeks in wild-type hosts whereas it persisted for 6 weeks in CD8-/- hosts. Eosinophil infiltrates in lymph nodes were significantly enhanced in CD8-/- hosts. Increased serum levels of IL-5 and IL-13 in CD8-/- hosts confirmed the enhancement of Th2-type responses in the context of recipient CD8+ T cell deficiency. Hyperplasia of lymph nodes and spleen were similar in both groups, as well as in vivo proliferation of donor CD4+ T cells. In vitro, CD8+ T cell regulation of the alloreactive Th2 response depended on their production of IFN-gamma and did not require expression of beta2m on CD4+ T cells or antigen-presenting cells. CONCLUSIONS: Host CD8+ T cells regulate alloreactive Th2 responses during graft-versus-host disease through an IFN-gamma dependent pathway, independently of the recognition of beta2m-associated MHC class I molecules.