Contribution of 4-1BBL on radioresistant cells in providing survival signals through 4-1BB expressed on CD8⁺ memory T cells in the bone marrow.

The persistence of memory lymphocytes is a critical feature of adaptive immunity. The TNF family ligand 4-1BBL supports the antigen-independent survival of CD8⁺ memory T cells. Here, we show that mice lacking 4-1BB only on αß T cells show a similar defect in CD8⁺ T-cell recall responses, as previously shown in 4-1BBL-deficient mice. We show that 4-1BB is selectively expressed on BM CD8⁺ but not CD4⁺ memory T cells of unimmunized mice. Its ligand, 4-1BBL, is found on VCAM-1⁺ stromal cells, CD11c⁺ cells, and a Gr1(lo) myeloid population in unimmunized mice. Adoptive transfer of in vitro generated memory T cells into mice lacking 4-1BBL only on radioresistant cells recapitulates the defect in CD8⁺ T-cell survival seen in the complete knockout mice, with smaller effects of 4-1BBL on hematopoietic cells. In BM, adoptively transferred DsRed CD8⁺ memory T cells are most often found in proximity to VCAM-1⁺ cells or Gr1⁺ cells, followed by B220⁺ cells and to a much lesser extent near CD11c⁺ cells. Thus, a VCAM-1⁺CD45(-) stromal cell is a plausible candidate for the radioresistant cell that provides 4-1BBL to CD8⁺ memory T cells in the BM.

Approaches to studying costimulation of human antiviral T cell responses: prospects for immunotherapeutic vaccines.

The generation of strong and specific CD8 T cell responses is important in the control of viral infections. Costimulatory molecules provide signals necessary for the development or maintenance of these responses. A major focus of our laboratory is to investigate the role of costimulatory molecules of the TNFR and CD28 families in antiviral responses. Our aim is to translate information obtained using murine models to the study of these molecules using human cells. We have devised an in vitro system using recombinant replication- deficient adenovirus to deliver costimulatory molecules to antigen-presenting cells that are then used to stimulate autologous T cells from both healthy and HIV-infected individuals. Here we describe our findings and discuss the implications of incorporating costimulatory molecules into viral vector vaccine strategies.

IL-15-dependent induction of 4-1BB promotes antigen-independent CD8 memory T cell survival.

Mice lacking CD137L (4-1BBL) show normal primary expansion and contraction of the CD8+ T cell response to influenza virus, but exhibit a defect in Ag-specific CD8+ T cell numbers at 3-6 wk postinfection. Previous results showed that the decrease in CD8+ T cell numbers in this model is not due to a programming defect during primary expansion. Thus, it appears that 4-1BB/4-1BBL interactions control the number of surviving CD8+ effector memory cells, late in the primary response. In this report, we asked how 4-1BB on T cells could play a role after Ag has apparently been cleared from the host. We show that IL-15, a cytokine involved in regulation of CD8+ memory T cell survival, induces the expression of 4-1BB on CD8+CD44(high) memory phenotype T cells, but not on CD4+ T cells. The Ag-independent induction of 4-1BB by IL-15 was dependent on MAPK p38 and ERK activation. Transfer of in vitro-generated OT-I CD8+ memory T cells into unimmunized wild-type or 4-1BBL-deficient hosts revealed a 2- to 3-fold survival advantage when 4-1BBL was present, recapitulating the effect seen in the endogenous response to influenza in mice. Decreases in the overall number of memory CD8+ T cells were also observed in the bone marrow of unmanipulated 4-1BBL-deficient mice. These data suggest a model whereby 4-1BB expression on memory CD8+ T cells, perhaps due to encounter with IL-15 in the bone marrow, allows 4-1BB/4-1BBL interactions to maintain memory CD8 T cell survival in the absence of Ag.

Role for inducible costimulator in control of Salmonella enterica serovar Typhimurium infection in mice.

Inducible costimulator (ICOS) is expressed on activated T cells and plays a key role in sustaining and enhancing the effector function of CD4 T cells. Given the function of this molecule in sustaining T-cell responses, we reasoned that ICOS might play an important role in a prolonged infection model, such as Salmonella infection of mice. To test this hypothesis, wild-type (WT) and ICOS-deficient (ICOS-/-) mice were infected systemically with a Salmonella enterica serovar Typhimurium strain expressing the chicken ovalbumin gene (Salmonella-OVA). ICOS-/- mice exhibited greater splenomegaly than WT mice and showed delayed bacterial clearance. The acquired immune response in this model was slow to develop. Maximal T-cell responses to Salmonella-OVA were detected at 3 weeks postinfection in both WT and ICOS-/- mice. CD4 T-cell-dependent gamma interferon production and a class switch to immunoglobulin G2a were severely reduced in ICOS-/- mice. ICOS-/- mice also exhibited a substantial defect in antigen-specific CD8 T-cell responses. In vitro, the effect of anti-ICOS on CD8 T-cell division was greater when CD8 T cells rather than CD4 T cells expressed ICOS, suggesting that the in vivo effects of ICOS on CD8 T cells could be direct. Taken together, these studies show that ICOS plays a critical role in control of Salmonella infection in mice, with effects on antibody, Th1, and CD8 T-cell responses.

Cooperation between 4-1BB and ICOS in the immune response to influenza virus revealed by studies of CD28/ICOS-deficient mice.

CD28, ICOS, and 4-1BB each play distinct roles in the CD8 T cell response to influenza virus. CD28-/- mice are severely impaired in primary CD8 T cell expansion and fail to mount a secondary response to influenza. Influenza-specific CD8 T cells expand normally in ICOS-/- mice, with only a small and transient defect late in the primary response and an unimpaired secondary response. Conversely, 4-1BB/4-1BBL interaction is dispensable for the primary CD8 T cell response to influenza, but maintains CD8 T cell survival and controls the size of the secondary response. Previous results showed that a single dose of agonistic anti-4-1BB Ab at priming allowed partial restoration of primary CD8 T cell expansion and full recovery of the secondary CD8 T cell responses to influenza in CD28-/- mice. In this study we show that anti-4-1BB fails to correct the CD8 T cell defect in CD28-/-ICOS-/- mice, suggesting that ICOS partially compensates for CD28 in this model. In support of this hypothesis, we found that anti-4-1BB enhances ICOS expression on both T cell subsets and that anti-4-1BB and anti-ICOS can synergistically activate CD4 and CD8 T cells. Furthermore, ICOS and 4-1BB can cooperate to directly stimulate isolated CD28-/- CD8 T cells. These results reveal a novel interaction between the ICOS and 4-1BB costimulatory pathways as well as unexpected redundancy between CD28 and ICOS in primary CD8 T cell expansion. These findings have implications for costimulation of human T cell responses in diseases such as AIDS or rheumatoid arthritis, in which CD28- T cells accumulate.