John T. Harty

Programmed contraction of CD8 + T cells after infection

The extent of infection and rate of pathogen clearance are thought to determine both the magnitude of antigen-specific CD8+ T cell expansion and the ensuing contraction to a stable number of memory cells. We show that CD8+ T cell expansion after Listeria monocytogenes infection was primarily dependent on the initial infection dose or amount of antigen displayed, and was also influenced by the rate of pathogen clearance. However, the onset and kinetics of CD8+ T cell contraction after L. monocytogenes and lymphocytic choriomeningitis virus infections were independent of the magnitude of expansion, dose and duration of infection or amount of antigen displayed. Thus, major features of antigen-specific CD8+ T cell homeostasis, including the contraction phase of an immune response, may be programmed early after infection.

Specific immunity to listeria monocytogenes in the absence of IFNγ

Cytokine and cytokine receptor gene knockout mice provide powerful experimental systems to characterize the functions of these molecules in resistance to infectious disease. Such mice may also provide unique models of immune deficiency to learn whether manipulation of the immune response can overcome the specific dysfunction. We demonstrate that resistance of IFN gamma gene knockout (GKO-/-) mice to the intracellular bacterium Listeria monocytogenes is severely impaired compared with wild-type mice. However, immunization of GKO-/- mice with an attenuated L. monocytogenes strain generates antigen-specific CD8 T cell responses that can transfer immunity to naive hosts. Furthermore, vaccinated GKO-/- mice themselves exhibit 20,000-fold increased resistance to challenge with virulent L. monocytogenes and this resistance appears to be CD8 T cell mediated. These studies demonstrate that vaccination-induced immunity can overcome the absence of a cytokine that is critical for resistance to acute infection.

Shaping and reshaping CD8 + T-cell memory

The ability to develop and sustain populations of memory T cells after infection or immunization is a hallmark of the adaptive immune response and a basis for protective vaccination against infectious disease. Technical advances that allow direct ex vivo identification and characterization of antigen-specific CD8+ T cells at various stages of the response to infection or vaccination in mouse models have fuelled efforts to characterize the factors that control memory CD8+ T-cell generation. Here, we dissect the input signals that shape the characteristics of the memory CD8+ T-cell response and discuss how manipulation of these signals has the potential to reshape CD8+ T-cell memory and improve the efficacy of vaccination.

Accelerated CD8+ T-cell memory and prime-boost response after dendritic-cell vaccination.

Efficient boosting of memory T-cell numbers to protective levels generally requires a relatively long interval between immunizations. Decreasing this interval could be crucial in biodefense and cancer immunotherapy, in which rapid protective responses are essential. Here, we show that vaccination with peptide-coated dendritic cells (DCs) generated CD8+ T cells with the phenotype and function of memory cells within 4–6 d. These early memory CD8+ T cells underwent vigorous secondary expansion in response to a variety of booster immunizations, leading to elevated numbers of effector and memory T cells and enhanced protective immunity. Coinjection of CpG oligodeoxynucleotides, potent inducers of inflammation that did not alter the duration of DC antigen display, prevented the rapid generation of memory T cells in wild-type mice but not in mice lacking the interferon (IFN)-γ receptor. These data show that DC vaccination stimulates a pathway of accelerated generation of memory T cells, and suggest that events of inflammation, including the action of IFN-γ on the responding T cells, control the rate of development of memory CD8+ T cells.

CD8+ T cell contraction is controlled by early inflammation

Pathogen-specific CD8+ T cells expand in number after infection and then their numbers invariably contract by 90–95%, leaving a stable memory cell pool. The chief features of this response are programmed early after infection; however, the factors regulating contraction are mostly undefined. Here we show that antibiotic treatment before Listeria monocytogenes infection induced numbers of protective memory CD8+ T cells similar to those in control infected mice, by a pathway without contraction. The absence of contraction correlated with decreased early inflammation and interferon-γ production and an increased fraction of CD8+ T cells expressing the interleukin 7 receptor at the peak of the response. Thus, contraction is controlled by early inflammation but is not essential for the generation of protective memory CD8+ T cells after infection.

Therapeutic blockade of PD-L1 and LAG-3 rapidly clears established blood-stage Plasmodium infection

Infection of erythrocytes with Plasmodium species induces clinical malaria. Parasite-specific CD4+ T cells correlate with lower parasite burdens and severity of human malaria and are needed to control blood-stage infection in mice. However, the characteristics of CD4+ T cells that determine protection or parasite persistence remain unknown. Here we show that infection of humans with Plasmodium falciparum resulted in higher expression of the inhibitory receptor PD-1 associated with T cell dysfunction. In vivo blockade of the PD-1 ligand PD-L1 and the inhibitory receptor LAG-3 restored CD4+ T cell function, amplified the number of follicular helper T cells and germinal-center B cells and plasmablasts, enhanced protective antibodies and rapidly cleared blood-stage malaria in mice. Thus, chronic malaria drives specific T cell dysfunction, and proper function can be restored by inhibitory therapies to enhance parasite control.

Differentiation and Persistence of Memory CD8+ T Cells Depend on T Cell Factor 1

T cell factor 1 (TCF-1) is a transcription factor known to act downstream of the canonical Wnt pathway and is essential for normal T cell development. However, its physiological roles in mature CD8(+) T cell responses are unknown. Here we showed that TCF-1 deficiency limited proliferation of CD8(+) effector T cells and impaired their differentiation toward a central memory phenotype. Moreover, TCF-1-deficient memory CD8(+) T cells were progressively lost over time, exhibiting reduced expression of the antiapoptotic molecule Bcl-2 and interleukin-2 receptor beta chain and diminished IL-15-driven proliferation. TCF-1 was directly associated with the Eomes allele and the Wnt-TCF-1 pathway was necessary and sufficient for optimal Eomes expression in naive and memory CD8(+) T cells. Importantly, forced expression of Eomes partly protected TCF-1-deficient memory CD8(+) T cells from time-dependent attrition. Our studies thus identify TCF-1 as a critical player in a transcriptional program that regulates memory CD8 differentiation and longevity.

Compartmentalization of Bacterial Antigens: Differential Effects on Priming of CD8 T Cells and Protective Immunity

Bacterial pathogens synthesize numerous proteins that are either secreted or localized within bacterial cells. To address the impact of antigen compartmentalization on T cell immunity, we constructed recombinant Listeria monocytogenes that express a model CD8T cell epitope as a secreted or nonsecreted fusion protein. Both forms of the antigen, either secreted into the host cell cytoplasm or retained within bacterial cells, efficiently prime CD8 T cell responses. However, epitope-specific CD8 T cells confer protection only against bacteria secreting the antigen but not against the bacteria expressing the nonsecreted form of the same antigen. This dichotomy as a result of antigen compartmentalization suggests that bacterial antigens are presented by multiple MHC class I pathways to prime CD8 T cells, but only the endogenous pathway provides target antigens for CD8 T cell-mediated protective immunity.

Memory CD8 T cell responses exceeding a large but definable threshold provide long-term immunity to malaria

Infection of mice with sporozoites of Plasmodium berghei or Plasmodium yoelii has been used extensively to evaluate liver-stage protection by candidate preerythrocytic malaria vaccines. Unfortunately, repeated success of such vaccines in mice has not translated readily to effective malaria vaccines in humans. Thus, mice may be used better as models to dissect basic parameters required for immunity to Plasmodium-infection than as preclinical vaccine models. In turn, this basic information may aid in the rational design of malaria vaccines. Here, we describe a model of circumsporozoite-specific memory CD8 T cell generation that protects mice against multiple P. berghei sporozoite challenges for at least 19 months. Using this model we defined a threshold frequency of memory CD8 T cells in the blood that predicts long-term sterilizing immunity against liver-stage infection. Importantly, the number of Plasmodium-specific memory CD8 T cells required for immunity greatly exceeds the number required for resistance to other pathogens. In addition, this model allowed us to identify readily individual immunized mice that exceed or fall below the protective threshold before infection, information that should greatly facilitate studies to dissect basic mechanisms of protective CD8 T cell memory against liver-stage Plasmodium infection. Furthermore, the extremely large threshold in memory CD8 T cell frequencies required for long-term protection in mice may have important implications for development of effective malaria vaccines.

Impaired Assembly yet Normal Trafficking of MHC Class I Molecules in Tapasin Mutant Mice

Loading of peptides onto major histocompatibility complex class I molecules involves a multifactorial complex that includes tapasin (TPN), a membrane protein that tethers empty class I glycoproteins to the transporter associated with antigen processing. To evaluate the in vivo role of TPN, we have generated Tpn mutant mice. In these animals, most class I molecules exit the endoplasmic reticulum (ER) in the absence of stably bound peptides. Consequently, mutant animals have defects in class I cell surface expression, antigen presentation, CD8+ T cell development, and immune responses. These findings reveal a critical role of TPN for ER retention of empty class I molecules. Tpn mutant animals should prove useful for studies on alternative antigen-processing pathways that involve post-ER peptide loading.