During viral infection of the respiratory tract, CD27, 4-1BB, and OX40 collectively determine formation of CD8+ memory T cells and their capacity for secondary expansion

Independent studies have shown that CD27, 4-1BB, and OX40 can all promote survival of activated CD8+ T cells. We have therefore compared their impact on CD8+ memory T cell formation and responsiveness within one, physiologically relevant model system. Recombinant mice, selectively lacking input of one or two receptors, were challenged intranasally with influenza virus, and the immunodominant virus-specific CD8+ T cell response was quantified at priming and effector sites. Upon primary infection, CD27 and (to a lesser extent) 4-1BB made nonredundant contributions to accumulation of CD8+ virus-specific T cells in draining lymph nodes and lung, while OX40 had no effect. Interestingly though, in the memory response, accumulation of virus-specific CD8+ T cells in spleen and lung critically depended on all three receptor systems. This was explained by two observations: 1) CD27, 4-1BB, and OX40 were collectively responsible for generation of the same memory CD8+ T cell pool; 2) CD27, 4-1BB, and OX40 collectively determined the extent of secondary expansion, as shown by adoptive transfers with standardized numbers of memory cells. Surprisingly, wild-type CD8+ memory T cells expanded normally in primed OX40 ligand- or 4-1BB ligand-deficient mice. However, when wild-type memory cells were generated in OX40 ligand- or 4-1BB ligand-deficient mice, their secondary expansion was impaired. This provides the novel concept that stimulation of CD8+ T cells by OX40 and 4-1BB ligand during priming imprints into them the capacity for secondary expansion. Our data argue that ligand on dendritic cells and/or B cells may be critical for this.     

Characterization of the CD154-positive and CD40-positive cellular subsets required for pathogenesis in retrovirus-induced murine immunodeficiency

Genetically susceptible C57BL/6 (B6) mice that are infected with the LP-BM5 isolate of murine retroviruses develop profound splenomegaly, lymphadenopathy, hypergammaglobulinemia, terminal B-cell lymphomas, and an immunodeficiency state bearing many similarities to the pathologies seen in AIDS. Because of these similarities, this syndrome has been called murine AIDS (MAIDS). We have previously shown that CD154 (CD40 ligand)-CD40 molecular interactions are required both for the initiation and progression of MAIDS. Thus, in vivo anti-CD154 monoclonal antibody (MAb) treatment inhibited MAIDS symptoms in LP-BM5-infected wild-type mice when either a short course of anti-CD154 MAb treatment was started on the day of infection or a course was initiated 3 to 4 weeks after LP-BM5 administration, after disease was established. Here, we further characterize this required CD154-CD40 interaction by a series of adoptive transfer experiments designed to elucidate which cellular subsets must express CD154 or CD40 for LP-BM5 to induce MAIDS. Specifically with regard to CD154 expression, MAIDS-insusceptible B6 nude mice reconstituted with highly purified CD4+ T cells from wild-type, but not from CD154 knockout, B6 donors displayed clear MAIDS after LP-BM5 infection. In contrast, nude B6 recipients that received CD8+ T cells from wild-type B6 donors did not develop MAIDS after LP-BM5 infection. B6 CD40 knockout mice, which are also relatively resistant to LP-BM5-induced MAIDS, became susceptible to LP-BM5-induced disease after reconstitution with highly purified wild-type B cells but not after receiving purified wild-type dendritic cells (DC) or a combined CD40+ population composed of DC and macrophages obtained from B6 SCID mouse donors. Based on these and other experiments, we thus conclude that the cellular basis for the requirement for CD154-CD40 interactions for MAIDS induction and progression can be accounted for by CD154 expression on CD4+ T cells and CD40 expression on B cells.     

Ectopic CD40 ligand expression on B cells triggers intestinal inflammation

Several studies indicate that CD4(+) T cells, macrophages, and dendritic cells initially mediate intestinal inflammation in murine models of human inflammatory bowel disease. However, the initial role of B cells in the development of intestinal inflammation remains unclear. In this study we present evidence that B cells can trigger intestinal inflammation using transgenic (Tg) mice expressing CD40 ligand (CD40L) ectopically on B cells (CD40L/B Tg). We demonstrated that CD40L/B Tg mice spontaneously developed severe transmural intestinal inflammation in both colon and ileum at 8-15 wk of age. In contrast, CD40L/B TgxCD40(-/-) double-mutant mice did not develop colitis, indicating the direct involvement of CD40-CD40L interaction in the development of intestinal inflammation. The inflammatory infiltrates consisted predominantly of massive aggregated, IgM-positive B cells. These mice were also characterized by the presence of anti-colon autoantibodies and elevated IFN-gamma production. Furthermore, although mice transferred with CD4(+) T cells alone or with both CD4(+) T and B220(+) B cells, but not B220(+) cells alone, from diseased CD40L/B Tg mice, develop colitis, mice transferred with B220(+) B cells from diseased CD40L/B Tg mice and CD4(+) T cells from wild-type mice also develop colitis, indicating that the Tg B cells should be a trigger for this colitis model, whereas T cells are involved as effectors. As it has been demonstrated that CD40L is ectopically expressed on B cells in some autoimmune diseases, the present study suggests the possible contribution of B cells in triggering intestinal inflammation in human inflammatory bowel disease.     

CD40, but not CD154, expression on B cells is necessary for optimal primary B cell responses

CD40 is an important costimulatory molecule for B cells as well as dendritic cells, monocytes, and other APCs. The ligand for CD40, CD154, is expressed on activated T cells, NK cells, mast cells, basophils, and even activated B cells. Although both CD40(-/-) and CD154(-/-) mice have impaired ability to isotype switch, form germinal centers, make memory B cells, and produce Ab, it is not entirely clear whether these defects are intrinsic to B cells, to other APCs, or to T cells. Using bone marrow chimeric mice, we investigated whether CD40 or CD154 must be expressed on B cells for optimal B cell responses in vivo. We demonstrate that CD40 expression on B cells is required for the generation of germinal centers, isotype switching, and sustained Ab production, even when other APCs express CD40. In contrast, the expression of CD154 on B cells is not required for the generation of germinal centers, isotype switching, or sustained Ab production. In fact, B cell responses are completely normal when CD154 expression is limited exclusively to Ag-specific T cells. These results suggest that the interaction of CD154 expressed by activated CD4 T cells with CD40 expressed by B cells is the primary pathway necessary to achieve B cell activation and differentiation and that CD154 expression on B cells does not noticeably facilitate B cell activation and differentiation.     

CD4+ T lymphocytes expressing CD40 ligand help the IgM antibody response to soluble pneumococcal polysaccharides via an intermediate cell type

Streptococcus pneumoniae causes serious infections in children, the elderly, and immunocompromised patients. Protection against infections with S. pneumoniae is mediated through Abs against the capsular polysaccharides (caps-PS). We previously showed that the murine Ab response to caps-PS is dependent on CD40-CD40L interaction. In the present paper, we addressed the question of whether the CD40-CD40L-mediated modulation of the anti-caps-PS immune reaction is the result of a direct interaction between B lymphocytes and T lymphocytes or of an indirect interaction. SCID/SCID mice reconstituted with B lymphocytes from wild-type mice did not mount anti-caps-PS Abs. SCID/SCID mice reconstituted with B lymphocytes from wild-type mice and CD4+ T lymphocytes from wild-type mice but not CD4+ T lymphocytes from CD40L knockout mice stimulated the anti-caps-PS Ab response. This indicated that CD4+ T lymphocytes stimulated the anti-caps-PS Ab response in a CD40L-dependent manner. SCID/SCID mice reconstituted with B lymphocytes from CD40 knockout mice and CD4+ T lymphocytes from wild-type mice generated an anti-caps-PS Ab response that could be inhibited by MR1, a blocking anti-CD40L Ab. These data indicated that CD4+ T lymphocytes stimulated the anti-caps-PS Ab response in an indirect way. Finally, lethally irradiated CD40 knockout mice reconstituted with bone marrow from wild-type mice mounted an anti-caps-PS Ab response that was comparable to the Ab response in wild-type mice, revealing that the required CD40 was on hemopoietic cells. In conclusion, we provide evidence that CD4+ T lymphocytes expressing CD40L stimulate the Ab response to soluble caps-PS by interacting with CD40-expressing non-B cells.     

CD40 engagement on dendritic cells, but not on B or T cells, is required for long-term control of murine gammaherpesvirus 68

CD4 T cells are not essential for primary clearance of replicating murine gammaherpesvirus 68 (MHV-68) but are required for effective long-term control. The virus reactivates in the lungs of major histocompatibility complex class II-deficient (CII-/-) mice that lack functional CD4 T cells. CD40 ligand (CD40L) is upregulated on activated CD4 T cells, and it is thought that CD40-CD40L interactions are an important component of CD4 T-cell help. Our previous studies have shown that agonistic antibodies to CD40 can substitute for CD4 T-cell function in the long-term control of MHV-68. In the present study, we sought to identify the CD40-positive cell type mediating this effect. To address this question, we adoptively transferred MHV-68 peptide-pulsed CII(-/-) dendritic cells (DC) that had been treated with an agonistic antibody to CD40 into MHV-68-infected CII(-/-) recipients. Viral reactivation was significantly lower in mice injected with anti-CD40-treated DC than in those injected with control DC or in mice that did not receive any DC. However, in similar experiments with B cells, anti-CD40 treatment had no effect. We also investigated the requirement for CD40 expression on T cells by adoptive transfer of T cells from CD40(+/+) or CD40(-/-) mice into T-cell-deficient recipients that were subsequently infected with MHV-68. The results showed that CD40 expression on T cells is not necessary for preventing viral reactivation. Taken together, our data suggest that CD40 engagement on DC, but not on T or B cells, is essential for effective long-term control of MHV-68.     

Impaired ability of MHC class II-/- dendritic cells to provide tumor protection is rescued by CD40 ligation.

The contribution of CD4+ T cells to dendritic cell (DC) activation and to the induction of CD8+ T cell responses in vivo was investigated using a model of antitumor immune responses. Immunization with peptide-loaded MHC class II-deficient (MHC class II-/-) DC induced the activation of Ag-specific CD8+ T cells and their accumulation in the lymph nodes and spleens of immunized mice. The accumulation induced by MHC class II-/- DC immunization was lower than the accumulation observed after immunization with MHC class II+/+ DC. Similarly, immunization with peptide-loaded, MHC class II-/- DC induced some degree of protection against tumor challenge, but this protection was lower than the protection achieved after immunization with MHC class II+/+ DC. Incubation with a membrane-associated form of CD40 ligand resulted in the up-regulation of costimulatory molecules on MHC class II-/- DC and fully rescued their ability to induce antitumor immunity. We conclude that CD4+ T cells play a critical role in the generation of antitumor immune responses through their capacity to induce the activation of DC via CD40/CD40 ligand interaction, and thus maximize CD8+ T cell responses.     

Induction of anti-tumor immunity by vaccination with dendritic cells pulsed with anti-CD44 IgG opsonized tumor cells

Due to the pivotal role that dendritic cells (DC) play in eliciting and maintaining functional anti-tumor T cell responses, these APC have been exploited against tumors. DC express several receptors for the Fc portion of IgG (Fcγ receptors) that mediate the internalization of antigen-IgG complexes and promote efficient MHC class I and II restricted antigen presentation. In this study, the efficacy of vaccination with DC pulsed with apoptotic B16 melanoma cells opsonized with an anti-CD44 IgG (B16-CD44) was explored. Immature bone marrow derived DC grown in vitro with IL-4 and GM-CSF were pulsed with B16-CD44. After 48 h of pulsing, maturation of DC was demonstrated by production of IL-12 and upregulation of CD80 and CD40 expression. To test the efficacy of vaccination with DC+B16-CD44, mice were vaccinated subcutaneously Lymphocytes from mice vaccinated with DC+B16-CD44 produced IFN-γ in response to B16 melanoma lysates as well as an MHC class I restricted B16 melanoma-associated peptide, indicating B16 specific CD8 T cell activation. Upon challenge with viable B16 cells, all mice vaccinated with DC alone developed tumor compared to 40% of mice vaccinated with DC+B16-CD44; 60% of the latter mice remained tumor free for at least 8 months. In addition, established lung tumors and distant metastases were significantly reduced in mice treated with DC+B16-CD44. Lastly, delayed growth of established subcutaneous tumors was induced by combination therapy with anti-CD44 antibodies followed by DC injection. This study demonstrates the efficacy of targeting tumor antigens to DC via Fcγ receptors.     

CD40-CD40 ligand interactions promote trafficking of CD8+ T cells into the brain and protection against West Nile virus encephalitis

Recent studies have established a protective role for T cells during primary West Nile virus (WNV) infection. Binding of CD40 by CD40 ligand (CD40L) on activated CD4+ T cells provides an important costimulatory signal for immunoglobulin class switching, antibody affinity maturation, and priming of CD8+ T-cell responses. We examined here the function of CD40-dependent interactions in limiting primary WNV infection. Compared to congenic wild-type mice, CD40(-/-) mice uniformly succumbed to WNV infection. Although CD40(-/-) mice produced low levels of WNV-specific immunoglobulin M (IgM) and IgG, viral clearance from the spleen and serum was not altered, and CD8+ T-cell priming in peripheral lymphoid tissues was normal. Unexpectedly, CD8+ T-cell trafficking to the central nervous system (CNS) was markedly impaired in CD40(-/-) mice, and this correlated with elevated WNV titers in the CNS and death. In the brains of CD40(-/-) mice, T cells were retained in the perivascular space and did not migrate into the parenchyma, the predominant site of WNV infection. In contrast, in wild-type mice, T cells trafficked to the site of infection in neurons. Beside its role in maturation of antibody responses, our experiments suggest a novel function of CD40-CD40L interactions: to facilitate T-cell migration across the blood-brain barrier to control WNV infection.     

The roles of MHC class II, CD40, and B7 costimulation in CTL induction by plasmid DNA

DNA-based vaccines generate potent CTL responses. The mechanism of T cell stimulation has been attributed to plasmid-transfected dendritic cells. These cells have also been shown to express plasmid-encoded proteins and to become activated by surface marker up-regulation. However, the increased surface expression of CD40 and B7 on these dendritic cells is insufficient to overcome the need for MHC class II-restricted CD4(+) T cell help in the priming of a CTL response. In this study, MHC class II(-/-) mice were unable to generate a CTL response following DNA immunization. This deficit in CTL stimulation by MHC class II-deficient mice was only modestly restored with CD40-activating Ab, suggesting that there were other elements provided by MHC class II-restricted T cell help for CTL induction. CTL activity was also augmented by coinjection with a vector encoding the costimulatory ligand B7.1, but not B7.2. These data indicate that dendritic cells in plasmid DNA-injected mice require conditioning signals from MHC class II-restricted T cells that are both CD40 dependent and independent and that there are different roles for costimulatory molecules that may be involved in inducing optimal CTL activity.     

Protection from type 1 diabetes by invariant NK T cells requires the activity of CD4+CD25+ regulatory T cells

Invariant NK T (iNKT) cells regulate immune responses, express NK cell markers and an invariant TCR, and recognize lipid Ags in a CD1d-restricted manner. Previously, we reported that activation of iNKT cells by alpha-galactosylceramide (alpha-GalCer) protects against type 1 diabetes (T1D) in NOD mice via an IL-4-dependent mechanism. To further investigate how iNKT cells protect from T1D, we analyzed whether iNKT cells require the presence of another subset(s) of regulatory T cells (Treg), such as CD4+ CD25+ Treg, for this protection. We found that CD4+ CD25+ T cells from NOD.CD1d(-/-) mice deficient in iNKT cell function similarly in vitro to CD4+ CD25+ T cells from wild-type NOD mice and suppress the proliferation of NOD T responder cells upon alpha-GalCer stimulation. Cotransfer of NOD diabetogenic T cells with CD4+ CD25+ Tregs from NOD mice pretreated with alpha-GalCer demonstrated that activated iNKT cells do not influence the ability of T(regs) to inhibit the transfer of T1D. In contrast, protection from T1D mediated by transfer of activated iNKT cells requires the activity of CD4+ CD25+ T cells, because splenocytes pretreated with alpha-GalCer and then inactivated by anti-CD25 of CD25+ cells did not protect from T1D. Similarly, mice inactivated of CD4+ CD25+ T cells before alpha-GalCer treatment were also not protected from T1D. Our data suggest that CD4+ CD25+ T cells retain their function during iNKT cell activation, and that the activity of CD4+ CD25+ Tregs is required for iNKT cells to transfer protection from T1D.     

Murine Model of CD40-activation of B cells

Research on B cells has shown that CD40 activation improves their antigen presentation capacity. When stimulated with interleukin-4 and CD40 ligand (CD40L), human B cells can be expanded without difficulties from small amounts of peripheral blood within 14 days to very large amounts of highly-pure CD40-B cells (>10⁹ cells per patient) from healthy donors as well as cancer patients^1-4. CD40-B cells express important lymph node homing molecules and can attract T cells in vitro⁵. Furthermore they efficiently take up, process and present antigens to T cells^6,7. CD40-B cells were shown to not only prime naíve, but also expand memory T cells^8,9. Therefore CD40-activated B cells (CD40-B cells) have been studied as an alternative source of immuno-stimulatory antigen-presenting cells (APC) for cell-based immunotherapy1,5,10. In order to further study whether CD40-B cells induce effective T cell responses in vivo and to study the underlying mechanism we established a cell culture system for the generation of murine CD40-activated B cells. Using splenocytes or purified B cells from C57BL/6 mice for CD40-activation, optimal conditions were identified as follows: Starting from splenocytes of C57BL/6 mice (haplotype H-2b) lymphocytes are purified by density gradient centrifugation and co-cultured with HeLa cells expressing recombinant murine CD40 ligand (tmuCD40L HeLa)¹¹. Cells are recultured every 3-4 days and key components such as CD40L, interleukin-4, -Mercaptoethanol and cyclosporin A are replenished. In this protocol we demonstrate how to obtain fully activated murine CD40-B cells (mCD40B) with similar APC-phenotype to human CD40-B cells (Fig 1a,b). CD40-stimulation leads to a rapid outgrowth and expansion of highly pure (>90%) CD19+ B cells within 14 days of cell culture (Fig 1c,d). To avoid contamination with non-transfected cells, expression of the murine CD40 ligand on the transfectants has to be controlled regularly (Fig 2). Murine CD40-activated B cells can be used to study B-cell activation and differentiation as well as to investigate their potential to function as APC in vitro and in vivo. Moreover, they represent a promising tool for establishing therapeutic or preventive vaccination against tumors and will help to answer questions regarding safety and immunogenicity of this approach¹².Download video file.^{(141M, mp4)}     

CD83 expression influences CD4+ T cell development in the thymus

T lymphocyte selection and lineage commitment in the thymus requires multiple signals. Herein, CD4+ T cell generation required engagement of CD83, a surface molecule expressed by thymic epithelial and dendritic cells. CD83-deficient (CD83-/-) mice had a specific block in CD4+ single-positive thymocyte development without increased CD4+CD8+ double- or CD8+ single-positive thymocytes. This resulted in a selective 75%-90% reduction in peripheral CD4+ T cells, predominantly within the naive subset. Wild-type thymocytes and bone marrow stem cells failed to differentiate into mature CD4+ T cells when transferred into CD83-/- mice, while CD83-/- thymocytes and stem cells developed normally in wild-type mice. Thereby, CD83 expression represents an additional regulatory component for CD4+ T cell development in the thymus.     

Role of CD28 in the generation of effector and memory responses required for resistance to Toxoplasma gondii.

CD28 deficient (CD28-/-) mice were used to study the role of costimulation in the T cell-mediated, IFN-gamma-dependent mechanism of resistance to Toxoplasma gondii. These mice were resistant to infection with the ME49 strain of T. gondii. Analysis of the immune response of acutely infected CD28-/- mice revealed that IL-12 was required for T cell production of IFN-gamma and this was independent of the CD40/CD40 ligand interaction. A similar mechanism of IL-12-dependent, CD28/B7 independent production of IFN-gamma by T cells was also observed in wild-type mice. Interestingly, although chronically infected wild-type mice were resistant to rechallenge with the virulent RH strain of T. gondii, chronically infected CD28-/- mice were susceptible to rechallenge with the RH strain. This deficiency in the protective memory response by CD28-/- mice correlated with a lack of IL-2 and IFN-gamma in recall responses and reduced numbers of CD4+ T cells expressing a memory phenotype. Together, our findings demonstrate that CD28 is not required for the development of a protective T cell response to T. gondii, but CD28 is required for an optimal secondary immune response.     

CD28 is required for T cell activation and IFN-gamma production by CD4+ and CD8+ T cells in response to Trypanosoma cruzi infection

In the present study we evaluated the mechanisms behind the implication of the costimulatory molecule CD28 for the immune response against the intracellular protozoan parasite Trypanosma cruzi. Our results reveal a critical role for CD28 in the activation of both CD4+ and CD8+ T cells and induction of the effector mechanisms that ultimately mediate the control of parasite growth and pathogenesis in infected mice. CD28-deficient (CD28-/-) mice are highly susceptible to T. cruzi infection, presenting higher parasitemia and tissue parasitism, but less inflammatory cell infiltrate in the heart than C57Bl/6 wild-type (WT) mice. All the infected WT mice survived acute infection, whereas 100% of CD28-/- mice succumbed to it. The increased susceptibility of the CD28-/- mice was associated with a dramatic decrease in the production of IFN-gamma by both CD4+ and CD8+ T cells resulting in a diminished capacity to produce nitric oxide (NO) and mediate parasite killing. T cell activation was also profoundly impaired in CD28-/- mice, which presented decreased lymphoproliferative response after the infection compared to WT mice. Together, these data represent the first evidence that CD28 is critical for efficient CD4+ T cell activation in response to T. cruzi infection in mice.     

Restoration of the antibody response to IgE/antigen complexes in CD23-deficient mice by CD23+ spleen or bone marrow cells

Mice immunized with IgE/Ag complexes produce significantly more Ag-specific Abs than mice immunized with Ag alone. The enhancement is mediated via the low-affinity receptor for IgE (FcepsilonRII or CD23), as shown by its complete absence in mice pretreated with mAbs specific for CD23 and in CD23-deficient mice. Because the constitutive expression of murine CD23 is limited to B cells and follicular dendritic cells (FDCs), one of these cell types is likely to be involved. One of the suggested modes of action of IgE/CD23 is to increase the ability of B cells to present Ag to T cells, as demonstrated to take place in vitro. Another possibility is that FDCs capture the IgE/Ag complexes and present these directly to B cells. The purpose of the present study was to determine whether CD23+ B cells or FDCs are responsible for the IgE/CD23-mediated enhancement of specific Ab responses in vivo. We show that the enhancement is completely restored in irradiated CD23-deficient mice reconstituted with CD23+ spleen or bone marrow cells. In these mice, the B cells are CD23+ and the FDCs are presumably CD23- because the FDCs are radiation resistant and are reported not to be replaced by donor cells after this type of cell transfer. In contrast, enhancement was not restored in irradiated wild-type mice reconstituted with CD23- cells. These results indicate that CD23+ B cells, and not FDCs, are the cells that capture IgE/Ag complexes and induce enhancement of Ab responses in vivo.     

Bradykinin B2 Receptors of dendritic cells, acting as sensors of kinins proteolytically released by Trypanosoma cruzi, are critical for the development of protective type-1 responses

Although the concept that dendritic cells (DCs) recognize pathogens through the engagement of Toll-like receptors is widely accepted, we recently suggested that immature DCs might sense kinin-releasing strains of Trypanosoma cruzi through the triggering of G-protein-coupled bradykinin B2 receptors (B2R). Here we report that C57BL/6.B2R-/- mice infected intraperitoneally with T. cruzi display higher parasitemia and mortality rates as compared to B2R+/+ mice. qRT-PCR revealed a 5-fold increase in T. cruzi DNA (14 d post-infection [p.i.]) in B2R-/- heart, while spleen parasitism was negligible in both mice strains. Analysis of recall responses (14 d p.i.) showed high and comparable frequencies of IFN-gamma-producing CD4+ and CD8+ T cells in the spleen of B2R-/- and wild-type mice. However, production of IFN-gamma by effector T cells isolated from B2R-/- heart was significantly reduced as compared with wild-type mice. As the infection continued, wild-type mice presented IFN-gamma-producing (CD4+CD44+ and CD8+CD44+) T cells both in the spleen and heart while B2R-/- mice showed negligible frequencies of such activated T cells. Furthermore, the collapse of type-1 immune responses in B2R-/- mice was linked to upregulated secretion of IL-17 and TNF-alpha by antigen-responsive CD4+ T cells. In vitro analysis of tissue culture trypomastigote interaction with splenic CD11c+ DCs indicated that DC maturation (IL-12, CD40, and CD86) is controlled by the kinin/B2R pathway. Further, systemic injection of trypomastigotes induced IL-12 production by CD11c+ DCs isolated from B2R+/+ spleen, but not by DCs from B2R-/- mice. Notably, adoptive transfer of B2R+/+ CD11c+ DCs (intravenously) into B2R-/- mice rendered them resistant to acute challenge, rescued development of type-1 immunity, and repressed TH17 responses. Collectively, our results demonstrate that activation of B2R, a DC sensor of endogenous maturation signals, is critically required for development of acquired resistance to T. cruzi infection.     

Alpha beta TCR+ cells are a minimal fraction of peripheral CD8+ pool in MHC class I-deficient mice

MHC class I molecules play a role in the maintenance of the naive peripheral CD8+ T cell pool. The mechanisms of the peripheral maintenance and the life span of residual CD8+ cells present in the periphery of beta 2-microglobulin-deficient (beta 2m-/-) mice are unknown. We here show that very few CD8+ cells in beta 2m-/- mice coexpress CD8 beta, a marker of the thymus-derived CD8+ T cells. Most of the CD8 alpha+ cells express CD11c and can be found in beta 2m/RAG-2 double-deficient mice, demonstrating that these cells do not require rearranged Ag receptors for differentiation and survival and may be of dendritic cell lineage. Rare CD8 alpha+CD8 beta+ cells can be detected following in vivo alloantigenic stimulation 2 wk after the adult thymectomy. Selective MHC class I expression by bone marrow-derived cells does not lead to an accumulation of CD8 beta+ cells in beta 2m-/- mice. These findings demonstrate that 1) thymic export of CD8+ T cells in beta 2m-/- mice is reduced more severely than previously thought; 2) non-T cells expressing CD8 alpha become prominent when CD8+ T cells are virtually absent; 3) at least some beta 2m-/- CD8+ T cells have a life span in the periphery comparable to wild-type CD8+ cells; and 4) similar ligands induce positive selection in the thymus and survival of CD8+ T cells in the periphery.     

Mice deficient in OX40 and CD30 signals lack memory antibody responses because of deficient CD4 T cell memory

Recently, we reported that a CD4(+)CD3(-)CD11c(-) accessory cell provided OX40-dependent survival signals to follicular T cells. These accessory cells express both OX40 ligand and CD30 ligand, and the receptors, OX40 and CD30, are both expressed on Th2-primed CD4 T cells. OX40 and CD30 signals share common signaling pathways, suggesting that CD30 signals might substantially compensate in OX40-deficient mice. In this report we have dissected the signaling roles of CD30 alone and in combination with OX40. CD30-deficient mice showed an impaired capacity to sustain follicular germinal center responses, and recall memory Ab responses were substantially reduced. Deficiencies in OX40 and CD30 signals were additive; secondary Ab responses were ablated in double-deficient mice. Although the initial proliferation of OX40/CD30 double-knockout OTII transgenic T cells was comparable to that of their normal counterparts, they failed to survive in vivo, and this was associated with reduced T cell numbers associated with CD4(+)CD3(-) cells in B follicles. Finally, we show that OX40/CD30 double-knockout OTII transgenic T cells fail to survive compared with normal T cells when cocultured with CD4(+)CD3(-) cells in vitro.