Adaptive regulatory T cells that develop from naive CD4 cells in response to exposure to Ag can act as immunotherapeutic agents to control immune responses. We show that effectors generated from murine islet-specific CD4 cells by TCR stimulation with IL-2 and TGF-β1 have potent suppressive activity. They prevent spontaneous development of type 1 diabetes in NOD mice and inhibit development of pancreatic infiltrates and disease onset orchestrated by Th1 effectors. These regulatory T cells do not require innate CD25+ regulatory cells for generation or function, nor do they share some characteristics typically associated with them, including expression of CD25. However, the adaptive population does acquire the X-linked forkhead/winged helix transcription factor, FoxP3, which is associated with regulatory T cell function and maintains expression in vivo. One mechanism by which they may inhibit Th1 cells is via FasL-dependent cytotoxicity, which occurs in vitro. In vivo, they eliminate Th1 cells in lymphoid tissues, where Fas/FasL interactions potentially play a role because Th1 cells persist when this pathway is blocked. The results suggest that adaptive regulatory CD4 cells may control diabetes in part by impairing the survival of islet-specific Th1 cells, and thereby inhibiting the localization and response of autoaggressive T cells in the pancreatic islets.

Serotype-specific antibodies to pneumococcal capsular polysaccharide (PPS) are a critical component of vaccine-mediated immunity to Streptococcus pneumoniae. In this study, we investigated the in vitro opsonophagocytic activities of three PPS-specific mouse immunoglobulin G1 monoclonal antibodies (MAbs), 1E2, 5F6, and 7A9, and determined their in vivo efficacies against intranasal challenge with WU2, a serotype 3 pneumococcal strain, in normal and immunodeficient mice. The MAbs had different in vitro activities in a pneumococcal killing assay: 7A9 enhanced killing by mouse neutrophils and J774 cells in the presence of a complement source, whereas 5F6 promoted killing in the absence, but not the presence, of complement, and 1E2 did not promote killing under any conditions. Nonetheless, all three MAbs protected normal and complement component 3-deficient mice from a lethal intranasal challenge with WU2 in passive-immunization experiments in which 10 g of the MAbs were administered intraperitoneally before intranasal challenge. In contrast, only 1E2 protected Fc␥ receptor IIB knockout (Fc␥RIIB KO) mice and mice that were depleted of neutrophils with the MAb RB6, whereas 7A9 and 5F6 required neutrophils and Fc␥RIIB to mediate protection. Conversely, 7A9 and 5F6 protected Fc␥R KO mice, but 1E2 did not. Hence, the efficacy of 1E2 required an activating Fc␥R(s), whereas 5F6 and 7A9 required the inhibitory Fc␥R (Fc␥RIIB). Taken together, our data demonstrate that both MAbs that do and do not promote pneumococcal killing in vitro can mediate protection in vivo, although their efficacies depend on different host receptors and/or components.

Filamin A (FlnA) cross-links actin filaments and connects the Von Willebrand factor receptor GPIb-IX-V to the underlying cytoskeleton in platelets. Because FlnA deficiency is embryonic lethal, mice lacking FlnA in platelets were generated by breeding FlnAloxP/loxP females with GATA1-Cre males. FlnAloxP/y GATA1-Cre males have a macrothrombocytopenia and increased tail bleeding times. FlnA-null platelets have decreased expression and altered surface distribution of GPIbα because they lack the normal cytoskeletal linkage of GPIbα to underlying actin filaments. This results in ∼70% less platelet coverage on collagen-coated surfaces at shear rates of 1,500/s, compared with wild-type platelets. Unexpectedly, however, immunoreceptor tyrosine-based activation motif (ITAM)- and ITAM-like–mediated signals are severely compromised in FlnA-null platelets. FlnA-null platelets fail to spread and have decreased α-granule secretion, integrin αIIbβ3 activation, and protein tyrosine phosphorylation, particularly that of the protein tyrosine kinase Syk and phospholipase C–γ2, in response to stimulation through the collagen receptor GPVI and the C-type lectin-like receptor 2. This signaling defect was traced to the loss of a novel FlnA–Syk interaction, as Syk binds to FlnA at immunoglobulin-like repeat 5. Our findings reveal that the interaction between FlnA and Syk regulates ITAM- and ITAM-like–containing receptor signaling and platelet function.

Despite the success of the pneumococcal conjugate vaccine, pneumococcal pneumonia remains a significant clinical problem, and there is still much to learn about natural resistance and cellular immunity to pneumococcus. We investigated the role of T lymphocytes in resistance to serotype (ST) 3 Streptococcus pneumoniae in an intranasal infection model in C57BL/6 (wild-type [Wt]) and CD8+ (CD8−/−)- and CD4+ (MHC class II−/−)-deficient mice. CD8−/− mice exhibited significantly more bacterial dissemination and lung inflammation and a significantly more lethal phenotype than Wt mice. However, there was no difference in the bacterial dissemination, lung inflammation, or survival of Wt and MHC class II−/− mice. Perforin (Pfn)−/− and IFN-γ−/− mice, which were used to dissect the role of CD8+ T cells in our model, also exhibited a more lethal survival phenotype than Wt mice. Comparison of lung chemokine/cytokine levels by Luminex and cellular recruitment by FACS in Wt mice and knockout strains revealed that CD8−/− and IFN-γ−/− mice, which had the most lethal survival phenotype, had more CD4+IL-17+ T (Th17) cells, IL-17, neutrophil chemoattractants, and lung neutrophils, and fewer regulatory T cells than Wt mice. CD4+ T cell depletion improved the survival of ST-infected CD8−/− mice, and survival studies in Th17-deficient mice revealed that the Th17 response was dispensable for ST3 resistance in our model. Taken together, these findings demonstrate that CD8+ cells are required, but CD4+ T cells are dispensable for resistance to ST3 pneumonia in mice and suggest a previously unsuspected role for CD8+ cells in modulating the inflammatory response to ST3.