IL-2 during in vitro priming promotes subsequent engraftment and successful adoptive tumor immunotherapy by persistent memory phenotypic CD8(+) T cells

Adoptive T cell tumor immunotherapy potentially consists of two protective components by the transferred effector cells, the immediate immune response and the subsequent development of memory T cells. The extent by which adoptively transferred CD8(+) CTL are destined to become memory T cells is ambiguous as most studies focus on the acute effects on tumor shortly following adoptive transfer. In this study we show that a substantial fraction of the input CTL develop into memory cells that reject a s.c. tumor challenge. The use of exogenous IL-2 or a combination of IL-2 and IL-4, but not solely IL-4, during the ex vivo culture for the CTL inoculation was necessary for efficient development of CD8(+) memory T cells. Thus, an important component of adoptive immunotherapy using CTL is the production of CD8(+) Ag-specific memory cells which is primarily favored by IL-2 receptor signaling during ex vivo generation of the effector CTL.     

Dynamic regulation of IFN-gamma signaling in antigen-specific CD8+ T cells responding to infection

IFN-gamma plays a critical role in the CD8(+) T cell response to infection, but when and if this cytokine directly signals CD8(+) T cells during an immune response is unknown. We show that naive Ag-specific CD8(+) T cells receive IFN-gamma signals within 12 h after in vivo infection with Listeria monocytogenes and then become unresponsive to IFN-gamma throughout the ensuing Ag-driven expansion phase. Ag-specific CD8(+) T cells regain partial IFN-gamma responsiveness throughout the contraction phase, whereas the memory pool exhibits uniform, but reduced, responsiveness that is also modulated during the secondary response. The responsiveness of Ag-specific CD8(+) T cells to IFN-gamma correlated with modulation in the expression of IFN-gammaR2, but not with IFN-gammaR1 or suppressor of cytokine signaling-1. This dynamic regulation suggests that early IFN-gamma signals participate in regulation of the primary CD8(+) T cell response program, but that evading or minimizing IFN-gamma signals during expansion and the memory phase may contribute to appropriate regulation of the CD8(+) T cell response.     

Blockade of CTLA-4 decreases the generation of multifunctional memory CD4+ T cells in vivo

CTLA-4 is known as a central inhibitor of T cell responses. It terminates T cell activation and proliferation and induces resistance against activation induced cell death. However, its impact on memory formation of adaptive immune responses is still unknown. In this study, we demonstrate that although anti-CTLA-4 mAb treatment during primary immunization of mice initially enhances the number of IFN-γ-producing CD4(+) T cells, it does not affect the size of the memory pool. Interestingly, we find that the CTLA-4 blockade modulates the quality of the memory pool: it decreases the amount of specialized "multifunctional" memory CD4(+) T cells coproducing IFN-γ, TNF-α, and IL-2 in response to Ag. The reduction of these cells causes an immense decrease of IFN-γ-producing T cells after in vivo antigenic rechallenge. Chimeric mice expressing CTLA-4-competent and -deficient cells unmask, which these CTLA-4-driven mechanisms are mediated CD4(+) T cell nonautonomously. In addition, the depletion of CD25(+) T cells prior to the generation of Ag-specific memory cells reveals that the constitutively CTLA-4-expressing natural regulatory T cells determine the quality of memory CD4(+) T cells. Taken together, these results indicate that although the inhibitory molecule CTLA-4 damps the primary immune response, its engagement positively regulates the formation of a high-quality memory pool equipped with multifunctional CD4(+) T cells capable of mounting a robust response to Ag rechallenge.     

CD25-expressing CD8+ T cells are potent memory cells in old age

We have recently described an IL-2/IL-4-producing CD8+CD25+ non-regulatory memory T cell population that occurs in a subgroup of healthy elderly persons who characteristically still have a good humoral response after vaccination. The present study addresses this specific T cell subset and investigates its origin, clonal composition, Ag specificity, and replicative history. We demonstrate that CD8+CD25+ memory T cells frequently exhibit a CD4+CD8+ double-positive phenotype. The expression of the CD8 alphabeta molecule and the occurrence of signal-joint TCR rearrangement excision circles suggest a thymic origin of these cells. They also have longer telomeres than their CD8+CD25- memory counterparts, thus indicating a shorter replicative history. CD8+CD25+ memory T cells display a polyclonal TCR repertoire and respond to IL-2 as well as to a panel of different Ags, whereas the CD8+CD25- memory T cell population has a more restricted TCR diversity, responds to fewer Ags, and does not proliferate in response to stimulation with IL-2. Molecular tracking of specific clones with clonotypic primers reveals that the same clones occur in CD8+CD25+ and CD8+CD25- memory T cell populations, demonstrating a lineage relationship between CD25+ and CD25- memory CD8+ T cells. Our results suggest that CD25-expressing memory T cells represent an early stage in the differentiation of CD8+ cells. Accumulation of these cells in elderly persons appears to be a prerequisite of intact immune responsiveness in the absence of naive T cells in old age.     

Functional responses and costimulator dependence of memory CD4+ T cells

To examine the functional characteristics of memory CD4+ T cells, we used an adoptive transfer system to generate a stable population of Ag-specific memory cells in vivo and compared their responses to Ag with those of a similar population of Ag-specific naive cells. Memory cells localized to the spleen and lymph nodes of mice and exhibited extremely rapid recall responses to Ag in vivo, leaving the spleen within 3-5 days of Ag encounter. Unlike their naive counterparts, memory cells produced effector cytokines (IFN-gamma, IL-4, IL-5) within 12-24 h of Ag exposure and did not require multiple cycles of cell division to do so. Memory cells proliferated at lower Ag concentrations than did naive cells, were less dependent on costimulation by B7 molecules, and independent of costimulation by CD40. Furthermore, effector cytokine production by memory cells also occurred in the absence of either B7 or CD40 costimulation. Lastly, memory cells were resistant to tolerance induction. Together, these findings suggest that the threshold for activation of memory CD4+ cells is lower than that of naive cells. This would permit memory cells to rapidly express their effector functions in vivo earlier in the course of a secondary immune response, when the levels of Ag and the availability of costimulation may be relatively low.     

In vivo obtained antigen presented by germinal center B cells to T cells in vitro

Shortly after secondary immunization germinal center (GC) B cells obtain antigen from follicular dendritic cells (FDC) in the form of immune complexes. This antigen appears to be degraded by the GC B cells and may be processed for presentation to T cells. The present study was undertaken to determine whether GC B cells can process and present antigen obtained from FDC in vivo to appropriate T cells in vitro. GC B cells were isolated from immune mice with the use of Percoll density separation followed by a panning procedure which utilizes the ability of the plant lectin, peanut agglutinin (PNA), to selectively bind to GC B cells. The enriched GC B cells were approximately 80% highly positive for PNA, 97% positive for Ia and surface IgM, but less than 0.01% positive for Thy-1.2 or esterase. In some experiments, this population was further purified to near 100% highly PNA-positive cells with the use of fluoresceinated PNA and a fluorescence-activated cell sorter. Cell sorting analysis indicated that the antigen (125I-labeled ovalbumin (OVA)) was restricted to the highly PNA-positive cell fraction. The capacity of these highly PNA-positive B cells to present antigen was assessed by monitoring interleukin 2 (IL-2) production by the OVA-specific T cell hybridoma, 3DO-54.8. GC B cells obtained from mice 3 wk or more after secondary immunization did not elicit IL-2 production in the absence of added OVA. However, GC B cells isolated as early as 1 day and for over 1 wk after a challenge with OVA, were able to stimulate high levels of IL-2 production, in the absence of adding OVA to the cell cultures. This response was maximal on day 5 and corresponded precisely with the kinetics of the ultrastructural studies which document the uptake of antigen by GC B cells in vivo. The FDC-derived antigen was remarkably immunogenic when compared with exogenous antigen. These findings demonstrated that antigen obtained in vivo by GC B cells could be processed and presented to T cells. In vivo, GC B cells may induce the T cell help needed for the germinal center reaction, generate B memory cells, and help induce the high titers of antibody associated with the secondary antibody response.     

Subsets of memory cytotoxic T lymphocytes elicited by vaccination influence the efficiency of secondary expansion in vivo

Vaccine-elicited cytotoxic T lymphocytes (CTL) should be long-lived memory cells that can rapidly expand in number following re-exposure to antigen. The present studies were initiated to analyze the ability of plasmid interleukin-12 (IL-12) to augment CTL responses in mice when delivered during the peak phase of an immune response elicited by a plasmid human immunodeficiency virus type 1 gp120 DNA vaccine. Delivery of plasmid IL-12 on day 10 postimmunization resulted in a robust expansion of gp120-specific CD8+ T cells, as measured by tetramer, gamma interferon ELISPOT, and functional-killing assays. Interestingly, this delayed administration of plasmid IL-12 had no significant effect on antigen-specific CD4(+)-T-cell and antibody responses. Phenotypic analyses suggested that administration of plasmid IL-12 near the time of the peak CTL response activated and expanded antigen-specific effector cells, preventing their loss through apoptosis. However, this IL-12-augmented population of gp120-specific CD8+ T cells did not efficiently expand following gp120 boost immunization, suggesting that these effector cells would be of little utility in expanding to contain a viral infection. Analyses of the phenotypic profile and anatomic distribution of the plasmid IL-12-augmented CTL population indicated that these lymphocytes were primarily effector memory rather than central memory T cells. These observations suggest that CTL-based vaccines should elicit central memory rather than effector memory T cells and illustrate the importance of monitoring the phenotype and functionality of vaccine-induced, antigen-specific CTL.     

Impaired accumulation and function of memory CD4 T cells in human IL-12 receptor beta 1 deficiency

Defects in IL-12 production or IL-12 responsiveness result in a vulnerability to infection with non-viral intracellular organisms, but the immunological mechanisms responsible for this susceptibility remain poorly understood. We present an immunological analysis of a patient with disseminated Salmonella enteritidis and a homozygous splice acceptor mutation in the IL-12Rbeta1-chain gene. This mutation resulted in the absence of IL-12Rbeta1 protein on PBMC and an inability of T cells to specifically bind IL-12 or produce IFN-gamma in response to either IL-12 or IL-23. The accumulation of memory (CD45R0(high)) CD4 T cells that were CCR7(high) (putative central memory cells) was normal or increased for age. Central memory CD4 T cells of the patient and age-matched controls were similar in having a low to undetectable capacity to produce IFN-gamma after polyclonal stimulation. In contrast, the patient had a substantial decrease in the number of CCR7(neg/dull) CD45R0(high) memory CD4 T cells (putative effector memory cells), and these differed from control cells in having a minimal ability to produce IFN-gamma after polyclonal stimulation. Importantly, tetanus toxoid-specific IFN-gamma production by PBMC from the patient was also significantly reduced compared with that in age-matched controls, indicating that signaling via the IL-12Rbeta1-chain is generally necessary for the in vivo accumulation of human memory CD4 T cells with Th1 function. These results are also consistent with a model in which the IL-12Rbeta1 subunit is necessary for the conversion of central memory CD4 T cells into effector memory cells.     

Differential outcome of IL-2/anti-IL-2 complex therapy on effector and memory CD8+ T cells following vaccination with an adenoviral vector encoding EBV epitopes

IL-2/anti-IL-2 complex-based therapy has been proposed as a potential adjunct therapeutic tool to enhance in vivo efficacy of T cell-based immunotherapeutic strategies for chronic viral infections and human cancers. In this study, we demonstrate that IL-2 complex therapy can have discerning effects on CD8(+) T cells depending on their stage of differentiation. To delineate the underlying mechanism for these opposing effects on CD8(+) T cells, we examined the effects of IL-2 therapy during early priming, effector, and memory phases of T cell responses generated following immunization with an adenoviral vector encoding multiple EBV CD8(+) epitopes. IL-2 complex treatment during the early priming phase, which coincided with low levels of IL-2Rβ (CD122) and higher levels of IL-2Rα (CD25) on CD8(+) T cells, did not induce the expansion of effector T cells. In contrast, IL-2 complex treatment following the establishment of memory enhanced the expansion of Ag-specific T cells. Additionally, central memory T cells preferentially expanded following treatment at the expense of effector memory T cell populations. These studies demonstrate how differentiation status of the responding CD8(+) T cells impacts on their responsiveness to IL-2 complexes and highlight that timing of treatment should be considered before implementing this therapy in a clinical setting.     

Acellular pertussis booster in adolescents induces Th1 and memory CD8+ T cell immune response

In a number of countries, whole cell pertussis vaccines (wcP) were replaced by acellular vaccines (aP) due to an improved reactogenicity profile. Pertussis immunization leads to specific antibody production with the help of CD4(+) T cells. In earlier studies in infants and young children, wcP vaccines selectively induced a Th1 dominated immune response, whereas aP vaccines led to a Th2 biased response. To obtain data on Th1 or Th2 dominance of the immune response in adolescents receiving an aP booster immunization after a wcP or aP primary immunization, we analyzed the concentration of Th1 (IL-2, TNF-α, INF-γ) and Th2 (IL-4, IL-5, IL-10) cytokines in supernatants of lymphocyte cultures specifically stimulated with pertussis antigens. We also investigated the presence of cytotoxic T cell responses against the facultative intracellular bacterium Bordetella pertussis by quantifying pertussis-specific CD8(+) T cell activation following the aP booster immunization. Here we show that the adolescent aP booster vaccination predominantly leads to a Th1 immune response based on IFNgamma secretion upon stimulation with pertussis antigen, irrespective of a prior whole cell or acellular primary vaccination. The vaccination also induces an increase in peripheral CD8(+)CD69(+) activated pertussis-specific memory T cells four weeks after vaccination. The Th1 bias of this immune response could play a role for the decreased local reactogenicity of this adolescent aP booster immunization when compared to the preceding childhood acellular pertussis booster. Pertussis-specific CD8(+) memory T cells may contribute to protection against clinical pertussis.     

TGF-β Sensitivity Restrains CD8+ T Cell Homeostatic Proliferation by Enforcing Sensitivity to IL-7 and IL-15

The pleiotropic cytokine TGF-β has been implicated in the regulation of numerous aspects of the immune response, including naïve T cell homeostasis. Previous studies found that impairing TGF-β responsiveness (through expression of a dominant-negative TGF-β RII [DNRII] transgene) leads to accumulation of memory phenotype CD8 T cells, and it was proposed that this resulted from enhanced IL-15 sensitivity. Here we show naïve DNRII CD8 T cells exhibit enhanced lymphopenia-driven proliferation and generation of “homeostatic” memory cells. However, this enhanced response occurred in the absence of IL-15 and, unexpectedly, even in the combined absence of IL-7 and IL-15, which were thought essential for CD8 T cell homeostatic expansion. DNRII transgenic CD8 T cells still require access to self Class I MHC for homeostatic proliferation, arguing against generalized dysregulation of homeostatic cues. These findings suggest TGF-β responsiveness is critical for enforcing sensitivity to homeostatic cytokines that limit maintenance and composition of the CD8 T cell pool. (154 words).     

Capacities of migrating CD1b+ lymph dendritic cells to present Salmonella antigens to naive T cells

Dendritic cells (DCs) are well known as professional antigen-presenting cells (APC) able to initiate specific T-cell responses to pathogens in lymph nodes (LN) draining the site of infection. However, the respective contribution of migratory and LN-resident DCs in this process remains unclear. As DC subsets represent important targets for vaccination strategies, more precise knowledge of DC subsets able to present vaccine antigens to T cells efficiently is required. To investigate the capacities of DCs migrating in the lymph (L-DCs) to initiate a specific T-cell response, we used physiologically generated DCs collected from a pseudoafferent lymphatic cannulation model in sheep. The CD1b+ L-DCs were assessed for presenting antigens from the vaccine attenuated strain of Salmonella enterica serovar Abortusovis. CD1b+ L-DCs were able to phagocytose, process and to present efficiently Salmonella antigens to effector/memory T cells in vitro. They were shown to be efficient APC for the priming of allogeneic naive T cells associated with inducing both IFN-γ and IL-4 responses. They were also efficient in presenting Salmonella antigens to autologous naive T cells associated with inducing both IFN-γ and IL-10 responses. The capacities of L-DCs to process and present Salmonella antigens to T cells were investigated in vivo after conjunctival inoculation of Salmonella. The CD1b+ L-DCs collected after inoculation were able to induce the proliferative response of CD4+ T cells suggesting the in vivo capture of Salmonella antigens by the CD1b+ L-DCs, and their potential to present them directly to CD4+ T cells. In this study, CD1b+ L-DCs present potential characteristics of APC to initiate by themselves T cell priming in the LN. They could be used as target cells for driving immune activation in vaccinal strategies.     

Generation of CD8 T cell memory is regulated by IL-12

Various signals during infection influence CD8 T cell memory generation, but these factors have yet to be fully defined. IL-12 is a proinflammatory cytokine that has been shown to enhance IFN-gamma-producing T cell responses and has been widely tested as a vaccine adjuvant. In this study, we show that IL-12-deficient mice generate a weaker primary CD8 T cell response and are more susceptible to Listeria monocytogenes infection, but have substantially more memory CD8 T cells and greater protective immunity against reinfection. Kinetic analyses show that in the absence of IL-12 there is a reduced contraction of Ag-specific CD8 T cells and a gradual increase in memory CD8 T cells as a result of increased homeostatic renewal. By signaling directly through its receptor on CD8 T cells, IL-12 influences their differentiation to favor the generation of fully activated effectors, but hinders the formation of CD8 T cell memory precursors and differentiation of long-term CD8 T cell memory(.) These results have implications for understanding memory T cell development and enhancing vaccine efficacy, and offer new insight into the role of IL-12 in coordinating the innate and adaptive immune response.     

The role of B cells in the development of CD4 effector T cells during a polarized Th2 immune response

Previous studies have suggested that B cells promote Th2 cell development by inhibiting Th1 cell differentiation. To examine whether B cells are directly required for the development of IL-4-producing T cells in the lymph node during a highly polarized Th2 response, B cell-deficient and wild-type mice were inoculated with the nematode parasite, Nippostrongylus brasiliensis. On day 7, in the absence of increased IFN-gamma, IL-4 protein and gene expression from CD4 T cells in the draining lymph nodes were markedly reduced in B cell-deficient mice and could not be restored by multiple immunizations. Using a DO11.10 T cell adoptive transfer system, OVA-specific T cell IL-4 production and cell cycle progression, but not cell surface expression of early activation markers, were impaired in B cell-deficient recipient mice following immunization with N. brasiliensis plus OVA. Laser capture microdissection and immunofluorescent staining showed that pronounced IL-4 mRNA and protein secretion by donor DO11.10 T cells first occurred in the T cell:B cell zone of the lymph node shortly after inoculation of IL-4-/- recipients, suggesting that this microenvironment is critical for initial Th2 cell development. Reconstitution of B cell-deficient mice with wild-type naive B cells, or IL-4-/- B cells, substantially restored Ag-specific T cell IL-4 production. However, reconstitution with B7-1/B7-2-deficient B cells failed to rescue the IL-4-producing DO11.10 T cells. These results suggest that B cells, expressing B7 costimulatory molecules, are required in the absence of an underlying IFN-gamma-mediated response for the development of a polarized primary Ag-specific Th2 response in vivo.     

Memory CD8+ T cells protect dendritic cells from CTL killing

CD8(+) T cells have been shown to be capable of either suppressing or promoting immune responses. To reconcile these contrasting regulatory functions, we compared the ability of human effector and memory CD8(+) T cells to regulate survival and functions of dendritic cells (DC). We report that, in sharp contrast to the effector cells (CTLs) that kill DCs in a granzyme B- and perforin-dependent mechanism, memory CD8(+) T cells enhance the ability of DCs to produce IL-12 and to induce functional Th1 and CTL responses in naive CD4(+) and CD8(+) T cell populations. Moreover, memory CD8(+) T cells that release the DC-activating factor TNF-alpha before the release of cytotoxic granules induce DC expression of an endogenous granzyme B inhibitor PI-9 and protect DCs from CTL killing with similar efficacy as CD4(+) Th cells. The currently identified DC-protective function of memory CD8(+) T cells helps to explain the phenomenon of CD8(+) T cell memory, reduced dependence of recall responses on CD4(+) T cell help, and the importance of delayed administration of booster doses of vaccines for the optimal outcome of immunization.     

Memory CD4 T cells induce selective expression of IL-27 in CD8+ dendritic cells and regulate homeostatic naive T cell proliferation

Naive T cells undergo robust proliferation in lymphopenic conditions, whereas they remain quiescent in steady-state conditions. However, a mechanism by which naive T cells are kept from proliferating under steady-state conditions remains unclear. In this study, we report that memory CD4 T cells are able to limit naive T cell proliferation within lymphopenic hosts by modulating stimulatory functions of dendritic cells (DC). The inhibition was mediated by IL-27, which was primarily expressed in CD8(+) DC subsets as the result of memory CD4 T cell-DC interaction. IL-27 appeared to be the major mediator of inhibition, as naive T cells deficient in IL-27R were resistant to memory CD4 T cell-mediated inhibition. Finally, IL-27-mediated regulation of T cell proliferation was also observed in steady-state conditions as well as during Ag-mediated immune responses. We propose a new model for maintaining peripheral T cell homeostasis via memory CD4 T cells and CD8(+) DC-derived IL-27 in vivo.     

Mucosally delivered dendritic cells activate T cells independently of IL-12 and endogenous APCs

In vitro manipulated dendritic cells (DC) have increasingly been used as a promising vaccine formulation against cancer and infectious disease. However, improved understanding of the immune mechanisms is needed for the development of safe and efficacious mucosal DC immunization. We have developed a murine model of respiratory mucosal immunization by using a genetically manipulated DC vaccine. Within 24 h of intranasal delivery, the majority of vaccine DCs migrated to the lung mucosa and draining lymph nodes and elicited a significant level of T cells capable of IFN-gamma secretion and CTL in the airway lumen as well as substantial T cell responses in the spleen. And such T cell responses were associated with enhanced protection against respiratory mucosal intracellular bacterial challenge. In comparison, parenteral i.m. DC immunization did not elicit marked airway luminal T cell responses and immune protection regardless of strong systemic T cell activation. Although repeated mucosal DC delivery boosted Ag-specific T cells in the airway lumen, added benefits to CD8 T cell activation and immune protection were not observed. By using MHC-deficient vaccine DCs, we further demonstrated that mucosal DC immunization-mediated CD8 and CD4 T cell activation does not require endogenous DCs. By using IL-12-deficient vaccine DCs, we also observed that IL-12(-/-) DCs failed to migrate to the lymph nodes but remained capable of T cell activation. Our observations indicate that mucosal delivery of vaccine DCs represents an effective approach to enhance mucosal T cell immunity, which may operate independent of vaccine IL-12 and endogenous DCs.     

Durable human memory T cells quantifiable by cultured enzyme-linked immunospot assays are induced by heterologous prime boost immunization and correlate with protection against malaria

Immunological memory is a required component of protective antimalarial responses raised by T cell-inducing vaccines. The magnitude of ex vivo IFN-gamma T cell responses is widely used to identify immunogenic vaccines although this response usually wanes and may disappear within weeks. However, protection in the field is likely to depend on durable central memory T cells that are not detected by this assay. To identify longer-lived memory T cells, PBMC from malaria-naive vaccinated volunteers who had received prime boost vaccinations with a combination of DNA and/or viral vectors encoding the multiepitope string-thrombospondin-related adhesion protein Ag were cultured in vitro with Ag for 10 days before the ELISPOT assay. Ex vivo T cell responses peaked at 7 days after the final immunization and declined substantially over 6 mo, but responses identified after T cell culture increased over the 6-mo period after the final immunization. Moreover, individual cultured ELISPOT responses at the day of challenge time point correlated significantly with degree of protection against malaria sporozoite challenge, whereas ex vivo responses did not, despite a correlation between the peak ex vivo response and magnitude of memory responses 6 mo later. This cultured assay identifies long-lasting protective T cell responses and therefore offers an attractive option for assessments of vaccine immunogenicity.     

IL-15 transpresentation augments CD8+ T cell activation and is required for optimal recall responses by central memory CD8+ T cells

Although the adaptive immune system has a remarkable ability to mount rapid recall responses to previously encountered pathogens, the cellular and molecular signals necessary for memory CD8(+) T cell reactivation are poorly defined. IL-15 plays a critical role in memory CD8(+) T cell survival; however, whether IL-15 is also involved in memory CD8(+) T cell reactivation is presently unclear. Using artificial Ag-presenting surfaces prepared on cell-sized microspheres, we specifically addressed the role of IL-15 transpresentation on mouse CD8(+) T cell activation in the complete absence of additional stimulatory signals. In this study we demonstrate that transpresented IL-15 is significantly more effective than soluble IL-15 in augmenting anti-CD3epsilon-induced proliferation and effector molecule expression by CD8(+) T cells. Importantly, IL-15 transpresentation and TCR ligation by anti-CD3epsilon or peptide MHC complexes exhibited synergism in stimulating CD8(+) T cell responses. In agreement with previous studies, we found that transpresented IL-15 preferentially stimulated memory phenotype CD8(+) T cells; however, in pursuing this further, we found that central memory (T(CM)) and effector memory (T(EM)) CD8(+) T cells responded differentially to transpresented IL-15. T(CM) CD8(+) T cells undergo Ag-independent proliferation in response to transpresented IL-15 alone, whereas T(EM) CD8(+) T cells are relatively unresponsive to transpresented IL-15. Furthermore, upon Ag-specific stimulation, T(CM) CD8(+) T cell responses are enhanced by IL-15 transpresentation, whereas T(EM) CD8(+) T cell responses are only slightly affected, both in vitro and in vivo. Thus, our findings distinguish the role of IL-15 transpresentation in the stimulation of distinct memory CD8(+) T cell subsets, and they also have implications for ex vivo reactivation and expansion of Ag-experienced CD8(+) T cells for immunotherapeutic approaches.