Regulation of T cell activation by Notch ligand, DLL4, promotes IL-17 production and Rorc activation.

The activation and differentiation of T cells are dependent upon numerous initiating events that are influenced by the immune environment, nature of the Ag, as well as the activation state of APCs. In the present studies we have investigated the role of a specific notch ligand, delta-like 4 (Dll4). In particular, our data have indicated that Dll4 is inducible by pathogen-associated signals through TLR activation on dendritic cells but not early response inflammatory cytokines, IL-1 and IL-18 that also activate cells via MyD88 adapter pathway. Our observations from in vitro cultures confirmed earlier reports demonstrating that Dll4 inhibits Th2 cytokine production. Furthermore, Dll4 influences the generation of IL-17-producing T cells in the presence of additional skewing cytokines, IL-6 and TGF-beta. In the absence of notch signals, IL-17 production was significantly inhibited even under specific skewing conditions. These studies further demonstrate that Dll4 up-regulates Rorc expression in T cells and that both Rorc and Il17 gene promoters are direct transcriptional notch targets that further enhance the differentiation of Th17 cell populations. Thus, facilitation of efficient T cell differentiation may depend upon the activation of T cells via specific notch ligand stimulation.

MyD88-mediated instructive signals in dendritic cells regulate pulmonary immune responses during respiratory virus infection.

Respiratory syncytial virus (RSV) is the leading cause of respiratory disease in infants worldwide. The induction of innate immunity and the establishment of adaptive immune responses are influenced by the recognition of pathogen-associated molecular patterns by TLRs. One of the primary pathways for TLR activation is by MyD88 adapter protein signaling. The present studies indicate that MyD88 deficiency profoundly impacts the pulmonary environment in RSV-infected mice characterized by the accumulation of eosinophils and augmented mucus production. Although there was little difference in CD4 T cell accumulation, there was also a significant decrease in conventional dendritic cells recruitment to the lungs of MyD88(-/-) mice. The exacerbation of RSV pathophysiology in MyD88(-/-) mice was associated with an enhanced Th2 cytokine profile that contributed to an inappropriate immune response. Furthermore, bone marrow-derived dendritic cells (BMDC) isolated from MyD88(-/-) mice were incapable of producing two important Th1 instructive signals, IL-12 and delta-like4, upon RSV infection. Although MyD88(-/-) BMDCs infected with RSV did up-regulate costimulatory molecules, they did not up-regulate class II as efficiently and stimulated less IFN-gamma from CD4(+) T cells in vitro compared with wild-type BMDCs. Finally, adoptive transfer of C57BL/6 BMDCs into MyD88(-/-) mice reconstituted Th1 immune responses in vivo, whereas transfer of MyD88(-/-) BMDCs into wild-type mice skewed the RSV responses toward a Th2 phenotype. Taken together, our data indicate that MyD88-mediated pathways are essential for the least pathogenic responses to this viral pathogen through the regulation of important Th1-associated instructive signals.

Notch ligand Delta-like 4 regulates disease pathogenesis during respiratory viral infections by modulating Th2 cytokines.

Recent data have indicated that an important instructive class of signals regulating the immune response is Notch ligand-mediated activation. Using quantitative polymerase chain reaction, we observed that only Delta-like 4 (dll4) was up-regulated on bone marrow-derived dendritic cells after respiratory syncytial virus (RSV) infection, and that it was dependent on MyD88-mediated pathways. Using a polyclonal antibody specific for dll4, the development of RSV-induced disease was examined. Animals treated with anti-dll4 had substantially increased airway hyperresponsiveness compared with control antibody-treated animals. When the lymphocytic lung infiltrate was examined, a significant increase in total CD4+ T cells and activated (perforin+) CD8+ T cells was observed. Isolated lung CD4+ T cells demonstrated significant increases in Th2-type cytokines and a decrease in interferon gamma, demonstrating an association with increased disease pathogenesis. Parallel in vitro studies examining the integrated role of dll4 with interleukin-12 demonstrated that, together, both of these instructive signals direct the immune response toward a more competent, less pathogenic antiviral response. These data demonstrate that dll4-mediated Notch activation is one regulator of antiviral immunity.