Primary Memory CD8 Research Articles

Cutting Edge: Antitumor Immunity by Pathogen-Specific CD8 T Cells in the Absence of Cognate Antigen Recognition.

Cancer prognosis often correlates with the number of tumor-infiltrating CD8 T cells, but many of these cells recognize pathogens that commonly infect humans. The contribution of pathogen-specific "bystander" CD8 T cells to antitumor immunity remains largely unknown. Inflammatory cytokines are sufficient for memory CD8 T cell activation and gain of effector functions, indicating tumor-derived inflammation could facilitate pathogen-specific CD8 T cells to participate in tumor control. In this study, we show in contrast to tumor-specific CD8 T cells that pathogen-specific primary memory CD8 T cells inside tumor were not able to exert their effector functions and influence tumor progression. However, infection-induced memory CD8 T cells with defined history of repeated Ag encounters (i.e., quaternary memory) showed increased sensitivity to tumor-derived inflammation that resulted in activation, gain of effector functions, and better control of tumor growth. Thus, memory CD8 T cells with heightened ability to recognize environmental inflammatory stimuli can contribute to antitumor immunity in the absence of cognate Ag recognition.

Repeated Antigen Exposure Extends the Durability of Influenza-Specific Lung-Resident Memory CD8+ T Cells and Heterosubtypic Immunity

Lung-resident primary memory CD8+ Tcell populations (Trm) induced by a single influenza infection decline within months, rendering the host susceptible to new heterosubtypic influenza infections. Here, we demonstrate that, relative to single virus exposure, repeated antigen exposure dramatically alters the dynamics of influenza-specific lung Trm populations. Lung Trm derived from repeatedly stimulated circulating memory CD8+ Tcells exhibit extended durability and protective heterosubtypic immunity relative to primary lung Trm. Parabiosis studies reveal that the enhanced durability of lung Trm after multiple antigen encounters resulted from the generation of long-lasting circulating effector memory (Tem) populations, which maintained the ability to be recruited to the lung parenchyma and converted to Trm, in combination with enhanced survival of these cells in the lung. Thus, generating a long-lasting Trm precursor pool through repeated intranasal immunizations might be a promising strategy to establish long-lasting lung Trm-mediated heterosubtypic immunity against influenza.

Antigen-dependent and -independent contributions to primary memory CD8 T cell activation and protection following infection.

Memory CD8 T-cell activation, including expression of IFN-γ and granzymeB, can be induced by antigen (Ag)-dependent signals through the T-cell-receptor, or by pathogen-derived inflammatory cytokines in an Ag-independent manner. Recent studies have come to conflicting results regarding the contributions of Ag and/or inflammation to memory CD8 T-cell activation. Additionally, research has indicated that inflammation-driven CD8 T-cell responses during un-related infections (bystander activation) have the potential to provide protection, but whether protection occurs in immuno-competent hosts is unclear. To investigate these questions, we examined activation of virus-specific memory CD8 T-cells following infection with L. monocytogenes either expressing or not cognate Ag. We show that Ag and inflammation act synergistically in vitro to induce memory activation. In vivo, we found that when memory CD8 T-cells significantly contribute to clearance of infection, early activation and continued responses by these cells are enhanced by cognate Ag recognition. Mechanistically, we show that bystander responses by memory are dependent upon the dose of infection and the amount of inflammation elicited following infection and are able to provide protection in IFN-γ deficient mice, but not in immuno-competent hosts. The data elucidate the requirements for memory CD8 T-cell activation and the protective role of bystander responses.

Phenotypic and Functional Alterations in Circulating Memory CD8 T Cells with Time after Primary Infection.

Memory CD8 T cells confer increased protection to immune hosts upon secondary viral, bacterial, and parasitic infections. The level of protection provided depends on the numbers, quality (functional ability), and location of memory CD8 T cells present at the time of infection. While primary memory CD8 T cells can be maintained for the life of the host, the full extent of phenotypic and functional changes that occur over time after initial antigen encounter remains poorly characterized. Here we show that critical properties of circulating primary memory CD8 T cells, including location, phenotype, cytokine production, maintenance, secondary proliferation, secondary memory generation potential, and mitochondrial function change with time after infection. Interestingly, phenotypic and functional alterations in the memory population are not due solely to shifts in the ratio of effector (CD62Llo) and central memory (CD62Lhi) cells, but also occur within defined CD62Lhi memory CD8 T cell subsets. CD62Lhi memory cells retain the ability to efficiently produce cytokines with time after infection. However, while it is was not formally tested whether changes in CD62Lhi memory CD8 T cells over time occur in a cell intrinsic manner or are due to selective death and/or survival, the gene expression profiles of CD62Lhi memory CD8 T cells change, phenotypic heterogeneity decreases, and mitochondrial function and proliferative capacity in either a lymphopenic environment or in response to antigen re-encounter increase with time. Importantly, and in accordance with their enhanced proliferative and metabolic capabilities, protection provided against chronic LCMV clone-13 infection increases over time for both circulating memory CD8 T cell populations and for CD62Lhi memory cells. Taken together, the data in this study reveal that memory CD8 T cells continue to change with time after infection and suggest that the outcome of vaccination strategies designed to elicit protective memory CD8 T cells using single or prime-boost immunizations depends upon the timing between antigen encounters.

Diet-induced obesity does not impact the generation and maintenance of primary memory CD8 T cells.

The extent to which obesity compromises the differentiation and maintenance of protective memory CD8 T cell responses and renders obese individuals susceptible to infection remains unknown. In this study, we show that diet-induced obesity did not impact the maintenance of pre-existing memory CD8 T cells, including acquisition of a long-term memory phenotype (i.e., CD27(hi), CD62L(hi), KLRG1(lo)) and function (i.e., cytokine production, secondary expansion, and memory CD8 T cell-mediated protection). Additionally, obesity did not influence the differentiation and maintenance of newly evoked memory CD8 T cell responses in inbred and outbred hosts generated in response to different types of systemic (LCMV, L. monocytogenes) and/or localized (influenza virus) infections. Interestingly, the rate of naive-to-memory CD8 T cell differentiation after a peptide-coated dendritic cell immunization was similar in lean and obese hosts, suggesting that obesity-associated inflammation, unlike pathogen- or adjuvant-induced inflammation, did not influence the development of endogenous memory CD8 T cell responses. Therefore, our studies reveal that the obese environment does not influence the development or maintenance of memory CD8 T cell responses that are either primed before or after obesity is established, a surprising notion with important implications for future studies aiming to elucidate the role obesity plays in host susceptibility to infections.

Characterization of IL-15 independent memory CD8 T cell subsets (P1438)

Abstract IL-15 is required for the maintenance and homeostatic proliferation of conventionally characterized memory CD8 T cells, and thus sets a carrying capacity for the size of the memory CD8 T cell compartment. However, we recently demonstrated that heterologous prime boost (HPB) vaccine regimens that establish very abundant memory CD8 T cell populations result in an enlargement of the memory CD8 compartment, which is almost entirely due to the addition of vaccine-specific memory T cells that lack CD62L expression. Here, we test the hypothesis that memory CD8 T cells resulting from HPB are less dependent on IL-15. We demonstrate that boosted memory cells express decreased levels of IL15Rβ and undergo little homeostatic proliferation. When co-transferring primary and boosted memory CD8 T cells into IL15KOs, boosted memory CD8 T cells that lacked CD62L expression were better able to persist compared to primary memory CD8 T cells (regardless of CD62L expression). We extended these studies by demonstrating that primary memory CD8 T cells that are resident in several peripheral tissues were also maintained in the absence of IL-15, and in some cases, exhibited IL-15 independent homeostatic proliferation. These observations point to a distinct lineage, not wholly defined by lack of CD62L expression, that harbors less dependence on IL-15 for maintenance. This has ramifications for regulation of memory CD8 T cell quantity following certain vaccine modalities and at front line tissues.

The timing of stimulation of primary memory CD8 T cells impacts the rate of secondary memory generation and differentiation (P1402)

Abstract Memory CD8 T cell responses are essential in establishing long-lasting immunity, as these responses confer protection from repeated infections. Various factors have shown to influence the generation of memory CD8 T cells, suggesting that the process of memory differentiation is not fixed and can be manipulated. Here, we asked whether the timing of stimulation (or recruitment) of primary memory CD8 T cells impacts the generation and differentiation of secondary memory CD8 T cell responses after bacterial and viral infections. We show that despite decreased secondary expansion, the differentiation of primary memory CD8 T cells into populations with the long-term secondary memory phenotype (ex. CD62Lhi, CD27hi) and function (ex. tissue distribution, IL-2 production, antigen-driven proliferative expansion) is facilitated by late recruitment into the immune response. Additionally, we show that the expression of survival factors and transcription factors known to be involved in memory CD8 T cell differentiation (ex. Bcl2, Eomes, Tcf1) is increased in late-stimulated secondary effector CD8 T cells. Thus, these data suggest that the timing of antigen-stimulation modulates the program of secondary memory CD8 T cell differentiation, which may have relevance for the design of future prime-boost vaccine protocols.

T cell exhaustion in protozoan disease

Protozoan parasites cause severe morbidity and mortality in humans worldwide, especially in developing countries where access to chemotherapeutic agents is limited. Although parasites initially evoke a robust immune response, subsequent immunity fails to clear infection, ultimately leading to the chronic stage. This enigmatic situation was initially addressed in chronic viral models, where T cells lose their function, a phenomenon referred to as 'exhaustion'. However, recent studies demonstrate that this paradigm can be extended to protozoan diseases as well, although with notable differences. These studies have revealed that T cell responses generated against Toxoplasma gondii, Plasmodium sp., and Leishmania sp. can become dysfunctional. This review discusses T cell exhaustion in parasitic infection, mechanisms of development, and a possible role in disease outcome.

Population Dynamics of Naive and Memory CD8 T Cell Responses after Antigen Stimulations In Vivo

The extent to which the progeny of one primary memory CD8 T cell differs from the progeny of one naive CD8 T cell of the same specificity remains an unresolved question. To explore cell-autonomous functional differences between naive and memory CD8 T cells that are not influenced by differences in the priming environment, an experimental model has been developed in which physiological numbers of both populations of cells were cotransferred into naive hosts before Ag stimulation. Interestingly, naive CD8 T cells undergo greater expansion in numbers than do primary memory CD8 T cells after various infections or immunizations. The intrinsic ability of one naive CD8 T cell to give rise to more effector CD8 T cells than one memory CD8 T cell is independent of the number and quality of primary memory CD8 T cells present in vivo. The sustained proliferation of newly activated naive CD8 T cells contributed to their greater magnitude of expansion. Additionally, longitudinal analyses of primary and secondary CD8 T cell responses revealed that on a per-cell basis naive CD8 T cells generate higher numbers of long-lived memory cells than do primary memory CD8 T cells. This enhanced "memory generation potential" of responding naive CD8 T cells occurred despite the delayed contraction of secondary CD8 T cell responses. Taken together, the data in this study revealed previously unappreciated differences between naive and memory CD8 T cells and will help further define the functional potential for both cell types.

Increased Numbers of Preexisting Memory CD8 T Cells and Decreased T-bet Expression Can Restrain Terminal Differentiation of Secondary Effector and Memory CD8 T Cells

Memory CD8 T cells acquire effector memory cell properties after reinfection and may reach terminally differentiated, senescent states ("Hayflick limit") after multiple infections. The signals controlling this process are not well understood, but we found that the degree of secondary effector and memory CD8 T cell differentiation was intimately linked to the amount of T-bet expressed upon reactivation and preexisting memory CD8 T cell number (i.e., primary memory CD8 T cell precursor frequency) present during secondary infection. Compared with naive cells, memory CD8 T cells were predisposed toward terminal effector (TE) cell differentiation because they could immediately respond to IL-12 and induce T-bet, even in the absence of Ag. TE cell formation after secondary (2°) or tertiary infections was dependent on increased T-bet expression because T-bet(+/-) cells were resistant to these phenotypic changes. Larger numbers of preexisting memory CD8 T cells limited the duration of 2° infection and the amount of IL-12 produced, and consequently, this reduced T-bet expression and the proportion of 2° TE CD8 T cells that formed. Together, these data show that over repeated infections, memory CD8 T cell quality and proliferative fitness is not strictly determined by the number of serial encounters with Ag or cell divisions, but is a function of the CD8 T cell differentiation state, which is genetically controlled in a T-bet-dependent manner. This differentiation state can be modulated by preexisting memory CD8 T cell number and the intensity of inflammation during reinfection. These results have important implications for vaccinations involving prime-boost strategies.

Protective Capacity of Memory CD8 + T Cells Is Dictated by Antigen Exposure History and Nature of the Infection

Infection or vaccination confers heightened resistance to pathogen rechallenge because of quantitative and qualitative differences between naive and primary memory T cells. Herein, we show that secondary (boosted) memory CD8+ T cells were better than primary memory CD8+ T cells in controlling some, but not all acute infections with diverse pathogens. However, secondary memory CD8+ T cells were less efficient than an equal number of primary memory cells at preventing chronic LCMV infection and are more susceptible to functional exhaustion. Importantly, localization of memory CD8+ T cells within lymph nodes, which is reduced by antigen restimulation, was critical for both viral control in lymph nodes and for the sustained CD8+ T cell response required to prevent chronic LCMV infection. Thus, repeated antigen stimulation shapes memory CD8+ T cell populations to either enhance or decrease per cell protective immunity in a pathogen-specific manner, a concept of importance in vaccine design against specific diseases.

Molecular profiling of memory CD8 T cells after repeated antigen stimulations (39.5)

Abstract Repetitive antigen stimulation by prime-boost vaccination or pathogen re-encounter increases memory CD8 T cell numbers, however the impact on memory T cell differentiation is unknown. Here we show that repetitive antigen stimulations induce the selective accumulation of CD8 T cells with a uniform effector memory phenotype. In contrast to this, genome-wide microarray analysis reveals that each additional antigen challenge results in the differential regulation of several hundred genes in memory CD8 T cells and therefore in stepwise diversification of T cell transcriptomes. As a consequence, repeatedly stimulated memory CD8 T cells and primary memory CD8 T cells differ significantly in their molecular signature and share expression of only a small but preserved group of memory signature genes. Our results provide new insight into the complex regulation of memory CD8 T cell differentiation and identify potential new molecular targets to dissect and manipulate the function of memory T cells in human diseases.

Multiple antigen encounters decrease memory CD8 T cell-mediated protection against chronic viral infection (132.8)

Abstract While multiple studies demonstrate the enhanced protective capacity of primary memory compared to naive CD8 T cells, much less is known about the function and properties of memory CD8 T cells that have received multiple antigen exposures through either recurring infections or from booster immunizations. Therefore, we analyzed the ability of primary and secondary memory CD8 T cells to protect against a variety of pathogens expressing a common antigen. Our findings demonstrate that secondary memory CD8 T cells provide better per cell protective capacity than primary memory CD8 T cells against acute infections with both LCMV Armstrong and Listeria monocytogenes. In contrast, while primary memory CD8 T cells were able to effectively prevent a chronic infection with LCMV clone 13, an equal number of secondary memory T cells were unable to clear virus and became functionally exhausted. Delayed re-expression of CD62L on the secondary memory CD8 T cell population affected control of LCMV clone 13, as sorted CD62Lhi primary and secondary memory CD8 T cells were equally capable in preventing chronic infection. Overall, these studies demonstrate that multiple antigen encounters impact the ability of memory CD8 T cells to protect against different pathogens, information that should be considered during vaccine design. Supported by NIH (RO1-AI42767 to JTH and T32-AI07343 to JCN)

Secondary memory CD8+ T cells are more protective but slower to acquire a central–memory phenotype

The formation of memory CD8 T cells is an important goal of vaccination. However, although widespread use of booster immunizations in humans generates secondary and tertiary CD8 T cell memory, experimental data are limited to primary CD8 T cell memory. Here, we show that, compared with primary memory CD8 T cells, secondary memory CD8 T cells exhibit substantially delayed conversion to a central–memory phenotype, as determined by CD62L expression and interleukin (IL)-2 production. This delayed conversion to a central–memory phenotype correlates with reduced basal proliferation and responsiveness to IL-15, although in vitro coculture with a high concentration of IL-15 is capable of inducing proliferation and CD62L upregulation. Functionally, secondary memory CD8 T cells are more protective in vivo on a per cell basis, and this may be explained by sustained lytic ability. Additionally, secondary memory CD8 T cells are more permissive than primary memory CD8 T cells for new T cell priming in lymph nodes, possibly suggesting a mechanism of replacement for memory T cells. Thus, primary and secondary memory CD8 T cells are functionally distinct, and the number of encounters with antigen influences memory CD8 T cell function.